Pair your accounts.

Export articles to Mendeley

Get article recommendations from ACS based on references in your Mendeley library.

Pair your accounts.

Export articles to Mendeley

Get article recommendations from ACS based on references in your Mendeley library.

You’ve supercharged your research process with ACS and Mendeley!

STEP 1:
Click to create an ACS ID

Please note: If you switch to a different device, you may be asked to login again with only your ACS ID.

Please note: If you switch to a different device, you may be asked to login again with only your ACS ID.

Please note: If you switch to a different device, you may be asked to login again with only your ACS ID.

MENDELEY PAIRING EXPIRED
Your Mendeley pairing has expired. Please reconnect
ACS Publications. Most Trusted. Most Cited. Most Read
Antibody-Based Array for Tacrolimus Immunosuppressant Monitoring with Planar Plastic Waveguides Activated with an Aminodextran-Lipase Conjugate
More

OR SEARCH CITATIONS

My Activity
Publications

Figure 1Loading Img
Download Hi-Res ImageDownload to MS-PowerPointCite This:Anal. Chem. 2024, 96, 35, 14142-14149
  • Open Access
Article

Antibody-Based Array for Tacrolimus Immunosuppressant Monitoring with Planar Plastic Waveguides Activated with an Aminodextran-Lipase Conjugate
Click to copy article linkArticle link copied!

Open PDFSupporting Information (1)

Analytical Chemistry

Cite this: Anal. Chem. 2024, 96, 35, 14142–14149
Click to copy citationCitation copied!
https://doi.org/10.1021/acs.analchem.4c02028
Published August 22, 2024

Copyright © 2024 The Authors. Published by American Chemical Society. This publication is licensed under

CC-BY 4.0 .

Abstract

Click to copy section linkSection link copied!
High Resolution Image
Download MS PowerPoint Slide

Cyclic olefin copolymers (COC; e.g., Zeonor, Topas, Arton, etc.) are materials with outstanding properties for developing point-of-care systems; however, the lack of functional groups in their native form makes their application challenging. This work evaluates different strategies to functionalize commercially available Zeonor substrates, including oxygen plasma treatment, photochemical grafting, and direct surface amination using an amino dextran-lipase conjugate (ADLC). The modified surfaces were characterized by contact angle measurements, Fourier transform infrared-attenuated total reflection analysis, and fluorescence assays based on evanescent wave excitation. The bioaffinity activation through the ADLC approach results in a fast, simple, and reproducible approach that can be used further to conjugate carboxylated small molecules (e.g., haptens). The usefulness of this approach has been demonstrated by the development of a heterogeneous fluorescence immunoassay to detect tacrolimus (FK506) immunosuppressant drug using an array biosensor platform based on evanescence wave laser excitation and Zeonor-ADLC substrates. Surface modification with ADLC-bearing FK506 provides a 3D layer that efficiently leads to a remarkably low limit of detection (0.02 ng/mL) and IC50 (0.9 ng/mL) together with a wide dynamic range (0.07–11.3 ng/mL).

This publication is licensed under

CC-BY 4.0 .
  • cc licence
  • by licence
Copyright © 2024 The Authors. Published by American Chemical Society

Subjects

what are subjects
Tacrolimus (FK506) is a hydrophobic macrolide lactone produced by Streptomyces tsukubaensis and is widely used as an immunosuppressant drug after organ transplantation. (1) The free fraction of this drug in serum (1–2%) correlates with its accumulation in blood cells. (2) Due to its narrow therapeutic window, in situ (semi)continuous monitoring of FK506 in transplanted patients would be most helpful in boosting its therapeutic efficiency and avoiding potentially adverse effects. In this context, microdialysis systems interfaced with optical biosensors (3) could achieve the required quantification of FK506 and its metabolites in plasma thanks to their intrinsic high sensitivity, selectivity, and multiplexing capability (sensor arrays).
Point-of-care (POC) microdevices, including miniaturized platforms such as microfluidic devices and biosensors, offer all these features, particularly those fabricated in plastic materials such as cyclic olefin copolymers (COC). (3) In recent years, COC polymers have drawn attention as chip substrates for biomedical diagnostics due to their suitable optical properties (good transmittance in the near-UV region, low autofluorescence), much lower protein binding than glass and other plastic substrates, (4,5) negligible water uptake, low permeability for water vapor, and ease of fabrication. (3,6) Furthermore, they are compatible with bioreagents and biological samples, withstanding polar organic solvents typically used in life sciences (e.g., isopropanol, acetone). (6,7) Notably, some types of COCs, namely Zeonor and Zeonex, have been used for DNA immobilization, nucleic acid purification, and microarray fabrication. (8,9)
However, these materials lack functional groups due to their pure hydrocarbon composition, making surface modification difficult. The hydrophobic nature of the COCs can promote fouling by proteins, which is undesirable in applications for which protein adhesion is not required.
Various surface activation techniques have been proposed to overcome those challenges, including oxygen plasma treatment, UV ozonolysis, or photografting. (9−11) These methods aim to introduce hydrophilic polar functional groups to increase the surface free energy and minimize fouling by proteins. However, during the chemical modification process, the polymer surface may undergo oxidation, degradation, and cross-linking, which can cause structural alterations to its first few molecular layers, (12) thereby affecting the ability of light to propagate through COC substrates and preventing the generation of its evanescent field. (13) Additionally, batch-to-batch reproducibility is conditioned by the instability of these activated surfaces because polymer chains on the surface tend to rearrange stochastically and partially return to the native hydrophobic form. (14) Currently, a large number of POC devices employ waveguides to achieve measurements based on total internal reflection; (15) therefore, it is essential to maintain a correct propagation of the light through the substrate. For these reasons, an extensive effort has been devoted to implement new biomolecule immobilization strategies on COC substrates to improve uniformity, tethering stability, and density of active ligands. (6,12,16)
Lipases are enzymes that contain both hydrophilic and hydrophobic regions on their surface, which allows them to adsorb onto hydrophobic surfaces through a mechanism known as interfacial adsorption. (17) Such enzymes also feature remarkable stability; for instance, BTL2 produced by the thermophilic bacterium Geobacillus thermocatenulatus shows high thermal stability at intermediate temperatures (50 °C) and in alkaline media or organic solvents. (18,19) These properties suggest that lipases might be useful tools for streamlined COC functionalization and serve as scaffolds to further immobilize a variety of molecules, such as haptens or antibodies.
Chemically modified lipases can also be used for surface functionalization. For example, we have previously applied lipases modified with a dextran polysaccharide to develop microarray biosensors. The two-faced Janus material provided a flexible hydrophilic network to immobilize the hepatoxic microcystin LR (MCLR) in a high conjugation degree. The MCLR-dextran-lipase chimeras were immobilized by nonspecific hydrophobic interactions on planar glass substrates, thus improving the interaction with corresponding antibodies and leading to increased sensitivity and a larger dynamic range of the bioassay. (20,21)
In this work, the surface functionalization of commercially available Zeonor substrates was investigated using three strategies: oxygen plasma treatment, photochemical grafting, and direct surface amination using an aminated dextran-BTL2 conjugate (ADLC). Surface modification procedures were characterized by contact angle measurements, Fourier transform infrared-attenuated total reflection spectroscopy (FTIR-ATR), and atomic force microscopy. The suitability of the different approaches for the modification of the COC substrate has been assessed through a heterogeneous competitive fluorescence immunoassay to detect tacrolimus (FK506) (Figure 1).

Figure 1

Figure 1. Workflow of the developed competitive fluorescence immunoassay for the detection of FK506. (Left) Activation of the COC waveguides through the amino dextran-lipase conjugate and subsequent covalent binding of FK506-CO2H. (Right) Measurement and image acquisition: The array was then incubated with the sample containing a mixture of a positive control and anti-FK506 antibodies, followed by an incubation with the labeled detection antibodies.

High Resolution Image
Download MS PowerPoint Slide

Methods

Click to copy section linkSection link copied!

Surface Amination Using an Amino Dextran-Lipase Conjugate

In this novel approach, amino groups are introduced into specific areas of the Zeonor waveguide using a modified BTL2 lipase, which is adsorbed efficiently on the highly hydrophobic polymer surfaces. As previously described, (20) aminated dextran-BTL2 conjugates were hydrophobically adsorbed using a 15-channel PDMS patterning gasket placed over the functionalized waveguide. A 0.05 mg/mL conjugate solution in PBS (pH 7.4) was injected into each channel. After 30 min of incubation at room temperature, the channels were washed with 0.5 mL of PBS. The reaction scheme is depicted in Figure S1c.

Antigen Immobilization onto the Zeonor-Activated Sensing Surface

The functionalization of Zeonor substrates activated with primary amino groups, with the carboxylated FK506 hapten (FK506-CO2H) and the positive control (biotin), was carried out through their acidic groups by the active ester method. (22)
The active ester method was developed in one step in specific waveguide areas. For this purpose, a PDMS mold was arranged on the surface of aminated Zeonor, forming 15 channels parallel to each other. Each channel was filled with about 50 μL of a solution containing FK506-CO2H (100 μg/mL) or biotin (60 μg/mL), EDC (100 mM) and NHS (50 mM) in MES buffer (50 mM, pH 6.0), using hypodermic needles and disposable syringes. The mixtures were incubated on Zeonor substrates for at least 4 h at 4 °C and protected from light. Subsequently, the mold was removed from the surface and rinsed with deionized water. The patterned slides were blocked in PBSPF for 1 h, rinsed with deionized water, dried under argon, and either introduced into the Leopard Array for the measurements or stored at 4 °C.

Assay Protocol

The working principle of the assay is based on a competitive inhibition between FK506-CO2H, immobilized onto the Zeonor surface, and the FK506 in the sample for the antibody binding sites. The assay occurs only in the overlapping regions between those functionalized with the hapten and those of the assay channels, which means 90 sensing regions, 15 for each sample.
As described previously, the patterned modified Zeonor slides were placed in the array biosensor instrument. FK506 standard solutions (390 μL, 0.0001–100 μg/mL) were mixed with 10 μL of Ab solution (0.5 μg/mL IgG anti-FK506 and 0.5 μg/mL IgG antibiotin). After 5 min of preincubation at room temperature, the mixture was loaded into the sample reservoirs, pumped through the channels, and incubated statically over the sensor surface for 20 min. Then, the channels were rinsed twice with 800 μL of buffer, and 400 μL of a solution containing 2.5 μg/mL of labeled Abs in PBS was used to reveal the surface pattern of primary antibodies bound to the patterned antigens. This solution was pumped to the sensor surface through the channels and incubated for 20 min statically. The unbound labeled Abs was removed by rinsing four times the channels with 800 μL of PBS, and the slide was then imaged. A scheme of the immunoassay workflow is depicted in Figure S2a.
After the immunoaffinity reaction, the radiation from the diode laser uniformly illuminates the Zeonor substrate, and the produced evanescent wave excites the fluorescent molecules located in the sensing area. Intensity data were extracted from the CCD images (15 s, 21.5 gain units) and normalized as described elsewhere. (23) The positions and intensities of the fluorescent regions on the Zeonor surface allow for identification and quantification.
The fluorescence data were collected as the B (fluorescence signal in the presence of FK506), B0 (fluorescence signal in the absence of the analyte), and the response normalized using the following expression:
normalizedresponse=(B−B∞)(B0−B∞)
(1)
where B∞ is the background fluorescence obtained in the presence of an excess of FK506. The normalized experimental data were plotted as a function of the FK506 concentration in logarithmic scale and fitted to a four-parameter sigmoidal logistic equation (eq 2) using the Origin 2019 software:
normalizedsignal=Amax−Amin1+([FK506]/IC50)b+Amin
(2)
where Amax and Amin correspond, respectively, to the asymptotic maximum and minimum of the normalized signal, IC50 is the concentration of analyte at the inflection point (concentration that provides 50% inhibition of Amax), and b represents the slope of the curve at the inflection point. The limit of detection (LOD) corresponds to the analyte concentration for which the tracer binding to the antibody was inhibited by 10%, and the dynamic range (DR) of the method was calculated as the analyte concentrations that produced a normalized signal in the 20–80% range defined by the Amax and Amin asymptotes. The fluorescence signals were normalized with positive controls (biotin) to minimize variability between channels and slides. (23)

Results and Discussion

Click to copy section linkSection link copied!

Surface Treatment

As it was mentioned above, this work addresses different strategies to the challenging functionalization of the surface of the COCs, to achieve the most efficient immobilization of FK506-CO2H.

Feasibility of the Oxygen Plasma Treatment

Plasma treatment was carried out at two typical etching times (3 and 6 min), and the oxidation achieved through this protocol was assessed by measuring the degree of hydrophilicity by the water contact angle. Figure S3 shows the contact angles on Zeonor substrates after 0, 3, and 6 min of oxygen plasma treatment. The untreated Zeonor substrate exhibits a large contact angle (>75°) due to its highly hydrophobic nature. Interestingly, an exposure of at least 3 min to the oxygen plasma resulted in a marked decrease of the contact angles (<20°), achieving almost complete wettability in both cases and comparable to a conventional glass waveguide substrate (Figure S3).
The oxidized substrates were treated with 3-aminopropyltriethoxysilane (APTES, 3%, v/v) to introduce amino groups that react with FK506-CO2H and allow its covalent attachment to the substrate through an amide bond. (24) The behavior of the surface after functionalization with different concentrations of FK506-CO2H (90–900 μg/mL) showed a weak response when using the substrates exposed to plasma for 3 min (Figure S4a), and no response was obtained in the case of Zeonor treated for 6 min (Figure S4b).
The relatively low sensitivity obtained under these conditions might be attributed to the deterioration of the Zeonor optical properties upon the plasma treatment. (25) To confirm this adverse effect, the efficiency of the evanescent waves propagated through the Zeonor waveguide was evaluated by exposing a 0.5 μg/mL aqueous solution of AlexaFluor 647-anti-IgG to the evanescent field generated by total internal reflection (TIR) laser illumination at 635 nm. Fluorescence signals were extracted from the CCD images as pixel intensity values. As shown in Figure S5, the loss of excitation efficiency using Zeonor substrates exposed to 3 or 6 min of oxygen plasma was close to 60% with respect to the untreated surface. These results are comparable to those described in the literature for similar cyclo-olefin polymers (e.g., Zeonex, Topas), where extensive loss of light transmission was observed over almost the entire visible wavelength range, probably due to the formation of light-absorbing species and photo-oxidation reactions. (25,26)
Additionally, this analysis lends confirmation to the excellent optical characteristics of the original Zeonor, displaying a 130% better signal-to-noise (S/N) response compared to glass substrates.

Feasibility of the Photochemical Modification

Among the various surface modification methods, photoinduced graft polymerization has been proven to be suitable for introducing acrylic and methacrylic monomers on COC surfaces. (11,27) These processes are based on the use of hydrogen abstraction photoinitiators such as benzophenone, anthraquinones, or the like. (28,29) Therefore, this technique was also evaluated for the introduction of ω-amino groups on the Zeonor surfaces. The latter was achieved in two steps: (i) generation of surface-bonded initiators when UV-excited benzophenone (BP) or azobis(isobutyronitrile) (AIBN) abstract hydrogen atoms from the polymer surface; (ii) grafted polymerization for 30 min of 2-aminoethyl methacrylate (2-AEM) under UV-irradiation (312 nm) using a constant monomer concentration.
The effect of grafting was monitored by ATR-FTIR. Figure S6a shows the ATR-FTIR spectra corresponding to original and different 2-AEM-grafted Zeonor substrates (deoxygenated 0.1 M solution in 90:10 v/v MeOH-water) and increasing concentrations of the benzophenone initiator (0.5–5.0%, w/v). The ATR-FTIR spectra display the typical absorption bands of polyolefins, (30) including the symmetric and asymmetric vibrations of the C–H and C–C bonds at 2917 and 2845 cm–1, respectively, and the peaks around 1470 cm–1 corresponding to the deformation of the methylene group. However, the main absorption bands of the amino group (two bands at ca. 3400 cm–1 corresponding to the N–H stretch vibration) and the strong band of the C═O group (at ca. 1740 cm–1) were not observed, indicating the very low functionalization efficiency of the photochemical method.
To confirm these observations, conventional fluorescence-linked immunoassay (FLI) was performed using the aminated Zeonor substrates and following the procedure described above. As shown in Figure S6b, no response was observed in any of the cases. Similar results were obtained with AIBN as the photoinitiator; therefore, this functionalization strategy was discarded for further experiments.

Feasibility of the Aminated Dextran-Lipase Modification

The lipase selected to carry out the synthesis of the conjugates prepared in this work was BTL2 from Geobacillus thermocatenulatus, because it shows a good thermal stability in alkaline media or in the presence of organic solvents, allowing its chemical modification and conjugation to various molecules and polymers. Lipase was derivatized with dextran in the solid phase, following the steps previously described. (20) The efficient BTL2 modification with the polysaccharide was confirmed by Raman and ATR-FTIR (Figure S7) spectroscopies, and the amount of amino groups in the ADLC conjugates (ca. 0.1 μmol/mg) was assessed by the trinitrobenzenesulfonic acid (TNBS) test. (20,21) Also, the enzyme activity was evaluated after the chemical modification and was not significantly affected, retaining more than 80% of the initial enzymatic activity.
The Zeonor amination was performed in just one step by immobilizing the ADLC conjugate onto the planar waveguide through intermolecular hydrophobic interactions. The BTL2 lipase interacts with any nonpolar surface like they do on fat surfaces, and this interaction is quite specific since it adsorbs on hydrophobic surfaces at low ionic strength, the condition under which most proteins cannot do it. The lipase-hydrophobic surface complex is significantly stable and can be used in aqueous and anhydrous media. (18,19) We have already demonstrated the suitability of ADCL conjugates as tools for low molecular weight ligand immobilization in FLI development. (20) Now, we introduce a novel strategy in which the immobilization process is greatly simplified by using substrates of a hydrophobic nature, avoiding the tedious process of bringing hydrophobicity to surfaces as polar as glass waveguides.
To evaluate the adsorption of the ADLC conjugate on Zeonor surfaces, its topography was studied by AFM. Figure 2a–d shows the images obtained for a planar Zeonor waveguide before and after immobilization of the ADLC (50 μg/mL, 30 min incubation). As can be observed therein, the AFM images evidence the effective immobilization of the ADLC on the Zeonor surface since smoothing of the surface topography is noticed (decrease of the average vertical distance of ca. 11 nm). This observation is compatible with previous results (20) that suggest the formation of a monolayer of enzyme onto the Zeonor surface.

Figure 2

Figure 2. AFM topography of the planar Zeonor surface used as a waveguide: (a) original Zeonor substrate; (b) Zeonor functionalized with the ADLC solution (50 μg/mL, 30 min). (c, d) Topographic profiles along panels (a) and (b), respectively.

High Resolution Image
Download MS PowerPoint Slide
The immobilization procedure was optimized by studying several parameters that affect the efficiency of the FK506-CO2H immobilization on the ADLC-coated Zeonor, namely, ADLC concentration, nature and pH of the incubating solution, 1-ethyl-3-(3-(dimethylamino)propyl) carbodiimide (EDC) concentration, and the viability of using N-hydroxysuccinimide (NHS).

Surface Functionalization with ADLC

The sensor was fabricated by immobilizing the ADLC conjugate onto the Zeonor surface where the amino dextran network acquires an outward orientation, favorable for an accessible covalent immobilization of the FK506-CO2H. The effect of the ADLC concentration in the patterning solution was examined over the 10–100 μg/mL range and tested by the response of the biosensor in the absence of FK506 (B0). As shown in Figure 3a, the best coating efficiency was obtained with a concentration of 50 μg/mL of ADLC, using a fixed incubation time of 30 min, while higher concentrations do not lead to increased absorption of conjugate on the surface.

Figure 3

Figure 3. (a) Comparison of the efficiency of different amino dextran-BTL2 coating solutions at 10, 50, and 100 μg/mL. The conjugate-activated slide was patterned with FK506-CO2H (100 μg/mL) and incubated with 0.5 μg/mL anti-FK506 in the absence of free analyte. A 2.5 μg/mL Alexa Fluor 647-anti-IgM solution was used as the developer agent. (b) Biochip image obtained using two different methods for covalent coupling of FK506-CO2H: the one-step method (EDC) (blue) and the two-step method (EDC/NHS) (green). (c) Competitive calibration curve obtained at 62.5 (black square), 100 (red circle), and 200 μM (blue triangle) of EDC, using a fixed concentration of 50 mM NHS and 100 μg/mL FK506-CO2H. (d) Competitive calibration curve obtained at 50 (black circle), 100 (red diamond), and 200 μg/mL (blue triangle) of FK506-CO2H, with fixed concentrations of 50 mM NHS and 62.5 μM EDC. The assays were performed with FK506 (0.0001–100 μg/L) in the presence of 0.5 μg/mL anti-FK506 and antibiotin antibodies. A mixture of 2.5 μg/mL Alexa Fluor647-anti-IgM and Alexa Fluor 647-anti-IgG was used as the tracer solution. The results are mean signals ± standard errors of the mean (n ≥ 3).

High Resolution Image
Download MS PowerPoint Slide

FK506-CO2H Immobilization

Immobilization of FK506-CO2H was carried out onto ADLC-Zeonor substrates after activation of the hapten with the classical carbodiimide reaction in 2-(N-morpholino)ethanesulfonic acid (MES) buffer. Figure 3b depicts the biosensor response in the absence of FK506 (B0) when the hapten is conjugated to the aminated BTL2-dextran using 0.1 M of EDC in the absence and in the presence of 50 mM of NHS. As it can be observed, the efficiency of the EDC-mediated reaction improves notably in the presence of NHS. This behavior can be attributed to the fact that the ester formed between FK506-CO2H and NHS is more stable against hydrolysis in aqueous media than the one involving EDC and FK506-CO2H. (31)
To find the optimal amount of hapten and EDC for coupling to ADLC-Zeonor substrates, different concentrations of FK506-CO2H (50, 100, and 200 μg/mL) and EDC (62.5, 100, and 200 μM), were investigated. The NHS concentration was fixed at 50 mM because the rate-determining step of the carboxylate activation mechanism is its nucleophilic addition to EDC. The FK506-ADLC-Zeonor waveguides were tested using the FLI assay, and the substrates coupled with 100 μg/mL FK506-CO2H and 100 μM EDC resulted in the highest and most sensitive signal responses (Figure 3c,d). This observation also suggests that the signal responses of FLI could be adjusted by controlling the amount of FK506-CO2H added to the coupling solution. The reproducibility of the coupling protocol was evaluated by performing FLI calibration sets using different batches of activated Zeonor slides using the same protocol. The results showed no statistical variance between different batches prepared by different researchers on different days (RSD < 15.8%).

Selection of the Blocking Agent

In order to minimize the nonspecific binding of bioreagents on the chip patterned surface, which could lead to false positives, four blocking agents were tested: (a) protein-free PBS blocking buffer (PBSPF), (b) StartingBlock TBS blocking buffer (TBSS), (c) StartingBlock PBS blocking buffer (PBSS), and (d) PBS with 0.05% T20 and 0.3% powdered nonfat milk (PBSTM). Overall, the four blocking agents prevent nonspecific binding during the assay. There is no significant difference in the fluorescence intensity obtained in the absence of analyte (B0) when they are included in each assay (Figure 4). However, blocking the Zeonor surface with PBSPF provided the best results in terms of reproducibility (RSD < 20%) and sensitivity (the lowest B/B0 ratio for a B = 100 ng/mL of FK506).

Figure 4

Figure 4. (a) Biochip image using different blocking buffers. Zeonor slide was patterned with 50 μg/mL of aminated dextran-BTL2 conjugate and 100 μg/mL of FK506-CO2H (immunoassay region), 30 μg/mL biotin (as positive control, green “+” symbol), or MES buffer (as negative control, red “–” symbol). B0 (0 ng/mL FK506) and B1 (100 ng/mL FK506) and using 2.5 μg/mL Alexa Fluor 647-anti-IgM as a revealing agent. (b) Comparison of the efficiency of the blocking buffer solutions tested on the B1/B0 ratio (open symbols). The blue bars correspond to the fluorescence signal in the absence of FK506 (B0), the red bars correspond to the signal in the presence of 100 ng/mL of FK506 (B), the green bars correspond to the signal of the background, and the purple circles correspond to the B/B0 ratio. Results are shown as mean ± standard error of the mean (n ≥ 4).

High Resolution Image
Download MS PowerPoint Slide

Analytical Characteristics of Developed Biosensor

Figure 5 shows the competition inhibition curve obtained under the optimized conditions using FK506 standard solutions in the 0.0001–100 μg/L range. The obtained IC50 values and LOD values of the immunoassay were 0.9 and 0.02 ng/mL, respectively. The dynamic range, calculated from 20 to 80% inhibition, spanned from 0.07 to 11.3 ng/mL. Current clinical recommendation for FK506 measurements in transplanted patients involves a limit of quantification of 1 ng/mL to provide reliable amounts of the immunosuppressant drug during low-dose therapy. (32) In this regard, the biosensor developed herein would be sensitive enough to monitor FK506 in real world samples (even with up to 14-fold dilution if required to avoid a matrix effect), matching or improving the LOD of previously described methods (33,34) and opening the door to simplify the current surface modification protocols to manufacture functional POC devices based on COCs such as Zeonor (Table S1).

Figure 5

Figure 5. Competitive calibration curve. The assay was performed with FK506 (0.0001–100 μg/L) in the presence of 0.5 μg/mL of anti-FK506 and antibiotin antibodies. A mixture of 2.5 μg/mL Alexa Fluor 647-anti-IgM and Alexa Fluor 647-anti-IgG was used as the tracer solution. Results are shown as the mean ± standard error of the mean (n ≥ 3).

High Resolution Image
Download MS PowerPoint Slide
Assay selectivity was assessed with two ISDs typically coadministered with FK506 in transplanted patients (Figure S8). In both cases the two drugs exhibited negligible cross reactivities.
Additionally, the feasibility of biosensor regeneration was evaluated (Figure S9). The device can reliably detect FK506 over at least three cycles without significant loss of performance after treatment with a 50 mM NaOH solution at the end of each cycle, thereby confirming its suitability for semicontinuous operation in POC applications.

Conclusions

Click to copy section linkSection link copied!
This work demonstrates the applicability of ADLC conjugates as an efficient alternative for the functionalization of hydrophobic surfaces, such as COCs. The immobilization of custom-made amino-BTL2 conjugates onto planar plastic substrates is fast, straightforward, and reproducible. Further derivatization with FK506 yields a highly sensitive immunoassay for this drug. Therefore, the noncovalent immobilization of immunoreagents by way of their conjugates to a dextran-modified lipase represents an attractive alternative to conventional methods for heterogeneous immunoassays.
The novel aminodextran-lipase-coated COC substrates pave the way to develop on-a-chip immunosensors for point-of-care testing that meet their stringent requirements, as they have shown high reproducibility (mean interslide RSD of 16%), rapid substrate activation protocol (30 min), small volume (390 μL), and preservation of the optical properties of the COC substrate, among others. Furthermore, the method described herein represents a significant step toward the development of evanescent wave-based devices that can be integrated into multiplexed sensor systems.

Supporting Information

Click to copy section linkSection link copied!

The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acs.analchem.4c02028.

  • Details about the COC surface modification by oxygen plasma, photochemical grafting, and amino dextran-lipase conjugate; details about the biosensor development; scheme of the FK506 immunoassay protocol; comparison of the hydrophobicity of Zeonor before and after the plasma treatment; biochip images for the analysis of FK506 using the developed immunoarray; ATR-FTIR results; comparative summary of different methods reported for COC surface activation (PDF)

Terms & Conditions

Most electronic Supporting Information files are available without a subscription to ACS Web Editions. Such files may be downloaded by article for research use (if there is a public use license linked to the relevant article, that license may permit other uses). Permission may be obtained from ACS for other uses through requests via the RightsLink permission system: http://pubs.acs.org/page/copyright/permissions.html.

Author Information

Click to copy section linkSection link copied!
  • Corresponding Authors
    • Elena Benito-Peña - Department of Analytical Chemistry, Faculty of Chemistry, Universidad Complutense de Madrid, Plaza de las Ciencias 2, Madrid 28040, Spain;  Orcidhttps://orcid.org/0000-0001-5685-5559;  Email: [email protected]
    • Guillermo Orellana - Department of Organic Chemistry, Faculty of Chemistry, Universidad Complutense de Madrid, Plaza de las Ciencias 2, Madrid 28040, Spain;  Email: [email protected]
  • Authors
    • Bettina Glahn-Martínez - Department of Analytical Chemistry, Faculty of Chemistry, Universidad Complutense de Madrid, Plaza de las Ciencias 2, Madrid 28040, Spain;  Orcidhttps://orcid.org/0000-0003-0128-3113
    • Sonia Herranz - Department of Analytical Chemistry, Faculty of Chemistry, Universidad Complutense de Madrid, Plaza de las Ciencias 2, Madrid 28040, Spain
    • △Maria C. Moreno-Bondi - Department of Analytical Chemistry, Faculty of Chemistry, Universidad Complutense de Madrid, Plaza de las Ciencias 2, Madrid 28040, Spain;  Orcidhttps://orcid.org/0000-0002-3612-0675
  • Author Contributions

    B.G.-M. and S.H. contributed equally. The manuscript was written through contributions of all authors. All authors have given approval to the final version of the manuscript.

  • Notes
    The authors declare no competing financial interest.
    △Maria C. Moreno-Bondi, deceased (June 2022).

Acknowledgments

Click to copy section linkSection link copied!

This work was funded by the Spanish Ministry of Science and Innovation (MICIN, ref PID2021-127457OB-C21/22). B.G.-M. acknowledges Universidad Complutense de Madrid (UCM) for a research grant. The authors are grateful to Dr. J. M. Guisán (Institute of Catalysis and Petroleochemistry–CSIC) for providing the pT1-BTL2 plasmid and to Microfluidic ChipShop (Germany) for the Zeonor slides.

References

Click to copy section linkSection link copied!

This article references 34 other publications.

  1. 1
    Tanaka, H.; Kuroda, A.; Marusawa, H.; Hatanaka, H.; Kino, T.; Goto, T.; Hashimoto, M.; Taga, T. Structure of FK506, a Novel Immunosuppressant Isolated from Streptomyces. J. Am. Chem. Soc. 1987, 109 (16), 5031– 5033,  DOI: 10.1021/ja00250a050
    Google Scholar
    1
    Structure of FK506, a novel immunosuppressant isolated from Streptomyces
    Tanaka, Hirokazu; Kuroda, Akio; Marusawa, Hiroshi; Hatanaka, Hiroshi; Kino, Toru; Goto, Toshio; Hashimoto, Masashi; Taga, Toru
    Journal of the American Chemical Society (1987), 109 (16), 5031-3CODEN: JACSAT; ISSN:0002-7863.
    The structure I of FK506, a novel macrocyclic lactone isolated from Streptomyces tsukubaensis No. 9993 and which has immunosuppressant activity, was deduced on the basis of chem. and spectroscopic evidence. The relative configuration was detd. by an x-ray crystal anal. of I and the abs. configuration was established by the fact that hydrolysis of I gave L-pipecolic acid. The structure is unique in respect of contg. a hemiketal-masked α,β-diketoamide functionality incorporated in a 23-membered ring.
    >> More from SciFinder ®
    https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADyaL2sXlslGktr8%253D&md5=ca5492f7d8762dffc2e507d0345fa943
  2. 2
    Bittersohl, H.; Schniedewind, B.; Christians, U.; Luppa, P. B. A Simple and Highly Sensitive On-Line Column Extraction Liquid Chromatography-Tandem Mass Spectrometry Method for the Determination of Protein-Unbound Tacrolimus in Human Plasma Samples. J. Chromatogr. A 2018, 1547, 45– 52,  DOI: 10.1016/j.chroma.2018.03.010
    Google Scholar
    There is no corresponding record for this reference.
  3. 3
    Tombelli, S.; Trono, C.; Berneschi, S.; Berrettoni, C.; Giannetti, A.; Bernini, R.; Persichetti, G.; Testa, G.; Orellana, G.; Salis, F.; Weber, S.; Luppa, P. B.; Porro, G.; Quarto, G.; Schubert, M.; Berner, M.; Freitas, P. P.; Cardoso, S.; Franco, F.; Silverio, V.; Lopez-Martinez, M.; Hilbig, U.; Freudenberger, K.; Gauglitz, G.; Becker, H.; Gärtner, C.; O’Connell, M. T.; Baldini, F. An Integrated Device for Fast and Sensitive Immunosuppressant Detection. Anal. Bioanal. Chem. 2022, 414 (10), 3243– 3255,  DOI: 10.1007/s00216-021-03847-x
    Google Scholar
    There is no corresponding record for this reference.
  4. 4
    Fujita, R.; Nagatoishi, S.; Adachi, S.; Nishioka, H.; Ninomiya, H.; Kaya, T.; Takai, M.; Arakawa, T.; Tsumoto, K. Control of Protein Adsorption to Cyclo Olefin Polymer by the Hofmeister Effect. J. Pharm. Sci. 2019, 108 (5), 1686– 1691,  DOI: 10.1016/j.xphs.2018.12.023
    Google Scholar
    There is no corresponding record for this reference.
  5. 5
    ZEONEX® manufacturer; https://www.zeonex.com/pharmaceuticals.aspx.html (accessed 2024–07–29).
    Google Scholar
    There is no corresponding record for this reference.
  6. 6
    Agha, A.; Waheed, W.; Alamoodi, N.; Mathew, B.; Alnaimat, F.; Abu-Nada, E.; Abderrahmane, A.; Alazzam, A. A Review of Cyclic Olefin Copolymer Applications in Microfluidics and Microdevices. Macromol. Mater. Eng. 2022, 307 (8), 2200053,  DOI: 10.1002/mame.202200053
    Google Scholar
    6
    A Review of Cyclic Olefin Copolymer Applications in Microfluidics and Microdevices
    Agha, Abdulrahman; Waheed, Waqas; Alamoodi, Nahla; Mathew, Bobby; Alnaimat, Fadi; Abu-Nada, Eiyad; Abderrahmane, Aissa; Alazzam, Anas
    Macromolecular Materials and Engineering (2022), 307 (8), 2200053CODEN: MMENFA; ISSN:1438-7492. (Wiley-VCH Verlag GmbH & Co. KGaA)
    A review. Cyclic olefin copolymers (COC) are amorphous, transparent thermoplastics composed of cyclic olefin monomers (norbornene) and linear olefins (ethene). They are increasingly utilized as fabrication materials for microsystems and microfluidic devices, owing to their promising features of low water absorption, high elec. insulation, long-term stability of surface treatments, and resistance to a broad variety of acids and solvents. Many manufg. processes for COC-based devices have been developed in recent decades. These methodologies are categorized as replication methods or fast prototyping as common in fabrication of thermoplastic microfluidic devices. This review gives a full discussion of the features of COCs, the various prodn. processes, and the numerous selected applications in microfluidic platforms. The review also explores COCs compn. and fundamental features, as well as fabrication processes and applications in a variety of fields, investigates the materials potential advantages and uses, and attempts to create a comprehensive list of COCs possible benefits. Due to their unique features and simplicity of fabrication, COCs are projected to advance the future of microfluidics, microsystems, and optofluidics.
    >> More from SciFinder ®
    https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BB38XhtFGgtLbK&md5=c4c0b3795bdbabad3fb784d5ecac9e51
  7. 7
    Nunes, P. S.; Ohlsson, P. D.; Ordeig, O.; Kutter, J. P. Cyclic Olefin Polymers: Emerging Materials for Lab-on-a-Chip Applications. Microfluid. Nanofluidics 2010, 9 (2), 145– 161,  DOI: 10.1007/s10404-010-0605-4
    Google Scholar
    There is no corresponding record for this reference.
  8. 8
    Raj, J.; Herzog, G.; Manning, M.; Volcke, C.; MacCraith, B. D.; Ballantyne, S.; Thompson, M.; Arrigan, D. W. M. Surface Immobilisation of Antibody on Cyclic Olefin Copolymer for Sandwich Immunoassay. Biosens. Bioelectron. 2009, 24 (8), 2654– 2658,  DOI: 10.1016/j.bios.2009.01.026
    Google Scholar
    There is no corresponding record for this reference.
  9. 9
    Laib, S.; MacCraith, B. D. Immobilization of Biomolecules on Cycloolefin Polymer Supports. Anal. Chem. 2007, 79 (16), 6264– 6270,  DOI: 10.1021/ac062420y
    Google Scholar
    9
    Immobilization of Biomolecules on Cycloolefin Polymer Supports
    Laib, Stephan; MacCraith, Brian D.
    Analytical Chemistry (Washington, DC, United States) (2007), 79 (16), 6264-6270CODEN: ANCHAM; ISSN:0003-2700. (American Chemical Society)
    Recent trends in the development of microfluidic and biodiagnostic chips favor polymer materials over glass, primarily for optical and economical reasons. Therefore, existing chem. methods to prep. biomol. microarrays on glass slides have to be adapted or replaced to suit polymer substrates. Here the authors present a strategy to immobilize DNA and antibodies on cyclic polyolefin slides, like Zeonor. This polymer represents a class of new polymeric materials with excellent optical and mech. properties. By plasma and liq. chem. treatment followed by coating with polyelectrolytes, the authors have succeeded in immobilizing DNA onto the polymer substrate, yielding stable and versatile biosensor surfaces. The authors demonstrate the stability and usage of the coated Zeonor substrates not only in DNA chip technol. but also in protein chip technol. with DNA-directed immobilization of proteins.
    >> More from SciFinder ®
    https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2sXnsVyms7o%253D&md5=30075423ea8fb8929c3a64720a818ee0
  10. 10
    van Midwoud, P. M.; Janse, A.; Merema, M. T.; Groothuis, G. M. M.; Verpoorte, E. Comparison of Biocompatibility and Adsorption Properties of Different Plastics for Advanced Microfluidic Cell and Tissue Culture Models. Anal. Chem. 2012, 84 (9), 3938– 3944,  DOI: 10.1021/ac300771z
    Google Scholar
    10
    Comparison of Biocompatibility and Adsorption Properties of Different Plastics for Advanced Microfluidic Cell and Tissue Culture Models
    van Midwoud, Paul M.; Janse, Arnout; Merema, Marjolijn T.; Groothuis, Geny M. M.; Verpoorte, Elisabeth
    Analytical Chemistry (Washington, DC, United States) (2012), 84 (9), 3938-3944CODEN: ANCHAM; ISSN:0003-2700. (American Chemical Society)
    Microfluidic technol. is providing new routes toward advanced cell and tissue culture models to better understand human biol. and disease. Many advanced devices have been made from poly(dimethylsiloxane) (PDMS) to enable expts., for example, to study drug metab. by use of precision-cut liver slices, that are not possible with conventional systems. However, PDMS, a silicone rubber material, is very hydrophobic and tends to exhibit significant adsorption and absorption of hydrophobic drugs and their metabolites. Although glass could be used as an alternative, thermoplastics are better from a cost and fabrication perspective. Thermoplastic polymers (plastics) allow easy surface treatment and are generally transparent and biocompatible. This study focuses on the fabrication of biocompatible microfluidic devices with low adsorption properties from the thermoplastics poly(Me methacrylate) (PMMA), polystyrene (PS), polycarbonate (PC), and cyclic olefin copolymer (COC) as alternatives for PDMS devices. Thermoplastic surfaces were oxidized using UV-generated ozone or oxygen plasma to reduce adsorption of hydrophobic compds. Surface hydrophilicity was assessed over 4 wk by measuring the contact angle of water on the surface. The adsorption of 7-ethoxycoumarin, testosterone, and their metabolites was also detd. after UV-ozone treatment. Biocompatibility was assessed by culturing human hepatoma (HepG2) cells on treated surfaces. Comparison of the adsorption properties and biocompatibility of devices in different plastics revealed that only UV-ozone-treated PC and COC devices satisfied both criteria. This paper lays an important foundation that will help researchers make informed decisions with respect to the materials they select for microfluidic cell-based culture expts.
    >> More from SciFinder ®
    https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC38XksFWhu7k%253D&md5=a082cf6db9e72c5dc6a9adff2c033f72
  11. 11
    Peng, X.; Zhao, L.; Du, G.; Wei, X.; Guo, J.; Wang, X.; Guo, G.; Pu, Q. Charge Tunable Zwitterionic Polyampholyte Layers Formed in Cyclic Olefin Copolymer Microchannels through Photochemical Graft Polymerization. ACS Appl. Mater. Interfaces 2013, 5 (3), 1017– 1023,  DOI: 10.1021/am3027019
    Google Scholar
    There is no corresponding record for this reference.
  12. 12
    Hager, R.; Forsich, C.; Duchoslav, J.; Burgstaller, C.; Stifter, D.; Weghuber, J.; Lanzerstorfer, P. Microcontact Printing of Biomolecules on Various Polymeric Substrates: Limitations and Applicability for Fluorescence Microscopy and Subcellular Micropatterning Assays. ACS Appl. Polym. Mater. 2022, 4 (10), 6887– 6896,  DOI: 10.1021/acsapm.2c00834
    Google Scholar
    There is no corresponding record for this reference.
  13. 13
    Khonina, S. N.; Voronkov, G. S.; Grakhova, E. P.; Kazanskiy, N. L.; Kutluyarov, R. V.; Butt, M. A. Polymer Waveguide-Based Optical Sensors─Interest in Bio, Gas, Temperature, and Mechanical Sensing Applications. Coatings 2023, 13 (3), 549,  DOI: 10.3390/coatings13030549
    Google Scholar
    13
    Polymer Waveguide-Based Optical Sensors-Interest in Bio, Gas, Temperature, and Mechanical Sensing Applications
    Khonina, Svetlana N.; Voronkov, Grigory S.; Grakhova, Elizaveta P.; Kazanskiy, Nikolay L.; Kutluyarov, Ruslan V.; Butt, Muhammad A.
    Coatings (2023), 13 (3), 549CODEN: COATED; ISSN:2079-6412. (MDPI AG)
    A review. In the realization of photonic integrated devices, materials such as polymers are crucial. Polymers have shown compatibility with several patterning techniques, are generally affordable, and may be functionalized to obtain desired optical, elec., or mech. characteristics. Polymer waveguides are a viable platform for optical connectivity since they are easily adaptable to on-chip and on-board integration and promise low propagation losses <1 dB/cm. Furthermore, polymer waveguides can be made to be extremely flexible, able to withstand bending, twisting, and even stretching. Optical sensing is an interesting field of research that is gaining popularity in polymer photonics. Due to its huge potential for use in several industries, polymer waveguide-based sensors have attracted a lot of attention. Due to their resilience to electromagnetic fields, optical sensors operate better in difficult situations, such as those found in elec. power generating and conversion facilities. In this review, the most widely used polymer materials are discussed for integrated photonics. Moreover, four significant sensing applications of polymer-waveguide based sensors which include biosensing, gas sensing, temp. sensing and mech. sensing have been debated.
    >> More from SciFinder ®
    https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BB3sXmvFeqtrk%253D&md5=b622aea04cf66d36495384943e629c8d
  14. 14
    Encinas, N.; Díaz-Benito, B.; Abenojar, J.; Martínez, M. A. Extreme Durability of Wettability Changes on Polyolefin Surfaces by Atmospheric Pressure Plasma Torch. Surf. Coat. Technol. 2010, 205 (2), 396– 402,  DOI: 10.1016/j.surfcoat.2010.06.069
    Google Scholar
    There is no corresponding record for this reference.
  15. 15
    Herron, J. N.; Wang, H.-K.; Tan, L.; Brown, S. Z.; Terry, A. H.; Tolley, S. E.; Durtschi, J. D.; Simon, E. M.; Astill, M. E.; Smith, R. S.; Christensen, D. A. Planar Waveguide Biosensors for Point-of-Care Clinical and Molecular Diagnostics. In Fluorescence Sensors and Biosensors; Thompson, R. B., Ed.; CRC Press: Boca Raton, 2005.  DOI: 10.1201/9781420028287 .
    Google Scholar
    There is no corresponding record for this reference.
  16. 16
    Yan, J.; Zhao, C.; Ma, Y.; Yang, W. Covalently Attaching Hollow Silica Nanoparticles on a COC Surface for the Fabrication of a Three-Dimensional Protein Microarray. Biomacromolecules 2022, 23 (6), 2614– 2623,  DOI: 10.1021/acs.biomac.2c00354
    Google Scholar
    There is no corresponding record for this reference.
  17. 17
    Marciello, M.; Bolivar, J. M.; Filice, M.; Mateo, C.; Guisan, J. M. Preparation of Lipase-Coated, Stabilized, Hydrophobic Magnetic Particles for Reversible Conjugation of Biomacromolecules. Biomacromolecules 2013, 14 (3), 602– 607,  DOI: 10.1021/bm400032q
    Google Scholar
    There is no corresponding record for this reference.
  18. 18
    Schmidt-Dannert, C.; Rúa, M. L.; Atomi, H.; Schmid, R. D. Thermoalkalophilic Lipase of Bacillus Thermocatenulatus. I. Molecular Cloning, Nucleotide Sequence, Purification and Some Properties. Biochim. Biophys. Acta BBA - Lipids Lipid Metab. 1996, 1301 (1), 105– 114,  DOI: 10.1016/0005-2760(96)00027-6
    Google Scholar
    There is no corresponding record for this reference.
  19. 19
    Yenenler, A.; Venturini, A.; Burduroglu, H. C.; Sezerman, O. U. Investigating the Structural Properties of the Active Conformation BTL2 of a Lipase from Geobacillus Thermocatenulatus in Toluene Using Molecular Dynamic Simulations and Engineering BTL2 via In-Silico Mutation. J. Mol. Model. 2018, 24 (9), 229,  DOI: 10.1007/s00894-018-3753-1
    Google Scholar
    There is no corresponding record for this reference.
  20. 20
    Herranz, S.; Marciello, M.; Olea, D.; Hernández, M.; Domingo, C.; Vélez, M.; Gheber, L. A.; Guisán, J. M.; Moreno-Bondi, M. C. Dextran–Lipase Conjugates as Tools for Low Molecular Weight Ligand Immobilization in Microarray Development. Anal. Chem. 2013, 85 (15), 7060– 7068,  DOI: 10.1021/ac400631t
    Google Scholar
    There is no corresponding record for this reference.
  21. 21
    Herranz, S.; Marciello, M.; Marco, M.-P.; Garcia-Fierro, J. L.; Guisan, J. M.; Moreno-Bondi, M. C. Multiplex Environmental Pollutant Analysis Using an Array Biosensor Coated with Chimeric Hapten-Dextran-Lipase Constructs. Sens. Actuators B Chem. 2018, 257, 256– 262,  DOI: 10.1016/j.snb.2017.10.134
    Google Scholar
    There is no corresponding record for this reference.
  22. 22
    Hermanson, G. T. The Chemistry of Reactive Groups. In Bioconjugate Techniques (Second ed.); Hermanson, G. T., Ed.; Academic Press: New York, 2008; pp 169– 212.  DOI: 10.1016/B978-0-12-370501-3.00002-3 .
    Google Scholar
    There is no corresponding record for this reference.
  23. 23
    Herranz, S.; Marazuela, M. D.; Moreno-Bondi, M. C. Automated Portable Array Biosensor for Multisample Microcystin Analysis in Freshwater Samples. Biosens. Bioelectron. 2012, 33 (1), 50– 55,  DOI: 10.1016/j.bios.2011.12.016
    Google Scholar
    There is no corresponding record for this reference.
  24. 24
    Glahn-Martínez, B.; Jurado-Sánchez, B.; Benito-Peña, E.; Escarpa, A.; Moreno-Bondi, M. C. Magnetic Janus Micromotors for Fluorescence Biosensing of Tacrolimus in Oral Fluids. Biosens. Bioelectron. 2024, 244, 115796  DOI: 10.1016/j.bios.2023.115796
    Google Scholar
    There is no corresponding record for this reference.
  25. 25
    Munzert, P.; Schulz, U.; Kaiser, N. Transparent Thermoplastic Polymers in Plasma-Assisted Coating Processes. Surf. Coat. Technol. 2003, 174–175, 1048– 1052,  DOI: 10.1016/S0257-8972(03)00414-6
    Google Scholar
    There is no corresponding record for this reference.
  26. 26
    Voigt, J. Photochemische und photooxydative Reaktionen. In Die Stabilisierung der Kunststoffe gegen Licht und Wärme; Springer-Verlag: Berlin, Heidelberg, NY, 1966; pp 67– 73.  DOI: 10.1007/978-3-642-52097-6 .
    Google Scholar
    There is no corresponding record for this reference.
  27. 27
    Jena, R. K.; Yue, C. Y. Cyclic Olefin Copolymer Based Microfluidic Devices for Biochip Applications: Ultraviolet Surface Grafting Using 2-Methacryloyloxyethyl Phosphorylcholine. Biomicrofluidics 2012, 6 (1), 012822  DOI: 10.1063/1.3682098
    Google Scholar
    27
    Cyclic olefin copolymer based microfluidic devices for biochip applications: Ultraviolet surface grafting using 2-methacryloyloxyethyl phosphorylcholine
    Jena, Rajeeb K.; Yue, C. Y.
    Biomicrofluidics (2012), 6 (1), 012822, 13 pp.CODEN: BIOMGB; ISSN:1932-1058. (American Institute of Physics)
    This report studies the surface modification of cyclic olefin copolymer (COC) by 2-methacryloyloxyethyl phosphorylcholine (MPC) monomer using photografting technique for the purpose of biointerface applications, which demonstrate resistance to both protein adsorption and cell adhesion in COC-based microfluidic devices. This is essential because the hydrophobic nature of COC can lead to adsorption of specific compds. from biol. fluids in the microchannel, which can affect the results during fluidic anal. and cause clogging inside the microchannel. A correlation was found between the irradn. time and hydrophobicity of the modified substrate. Static water contact angle results show that the hydrophilicity property of the MPC-grafted substrate improves with increasing irradn. time. The contact angle of the modified surface decreased to 20 ± 5 ° from 88 ± 3 ° for the untreated substrate. The surface characterization of the modified surface was evaluated using XPS and Fourier transform IR spectroscopy (FTIR spectroscopy). Attenuated total reflection-FTIR and XPS results show the presence of the phosphate group (P-O) on modified COC substrates, indicating that the hydrophilic MPC monomer has successfully grafted on COC. Finally, it was demonstrated that cell adhesion and protein adsorption on the MPC modified COC specimen has reduced significantly. (c) 2012 American Institute of Physics.
    >> More from SciFinder ®
    https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC38XhsFSktrjI&md5=6d12e28f8ba6a874e880aac2de4f4e33
  28. 28
    Ma, H.; Davis, R. H.; Bowman, C. N. A Novel Sequential Photoinduced Living Graft Polymerization. Macromolecules 2000, 33 (2), 331– 335,  DOI: 10.1021/ma990821s
    Google Scholar
    28
    A Novel Sequential Photoinduced Living Graft Polymerization
    Ma, Huimin; Davis, Robert H.; Bowman, Christopher N.
    Macromolecules (2000), 33 (2), 331-335CODEN: MAMOBX; ISSN:0024-9297. (American Chemical Society)
    A novel sequential UV-induced living graft polymn. method has been designed and investigated to modify polymeric materials. This method consists of two steps. In the first step, a surface initiator is formed on a substrate under UV irradn. in the presence of benzophenone (BP) solns.; in the second step, the monomers are grafted to the substrate by a living polymn. initiated by the surface photoinitiator. Hydrophobic porous polypropylene (PP) membranes were made hydrophilic and with neg. charged surface by grafting acrylic acid (AA). Exptl. results demonstrated that grafting d. and graft polymer chain length can be controlled by choosing the reaction conditions in the first step and in the subsequent step(s) independently. The amt. of grafted polymer relative to the total amt. of polymer from the novel sequential photoinduced graft polymn. method is 4-fold greater than that of the simultaneous grafting method for the system studied. In addn., a reaction mechanism was proposed and confirmed in the expts. With regard to the surface initiator formation caused by hydrogen abstraction, the kinetic studies show that the reaction rate has a max. value which depends on BP concn. With regard to the graft polymn. in the second step, there is a linear relationship between the graft polymn. rate and the monomer concn.
    >> More from SciFinder ®
    https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADyaK1MXotFKgtbc%253D&md5=6e86cbce87fa8d54d5495adfa46a27b1
  29. 29
    Stachowiak, T. B.; Svec, F.; Fréchet, J. M. J. Patternable Protein Resistant Surfaces for Multifunctional Microfluidic Devices via Surface Hydrophilization of Porous Polymer Monoliths Using Photografting. Chem. Mater. 2006, 18 (25), 5950– 5957,  DOI: 10.1021/cm0617034
    Google Scholar
    There is no corresponding record for this reference.
  30. 30
    Drnovská, H.; Lapčík, L.; Buršíková, V.; Zemek, J.; Barros-Timmons, A. M. Surface Properties of Polyethylene after Low-Temperature Plasma Treatment. Colloid Polym. Sci. 2003, 281 (11), 1025– 1033,  DOI: 10.1007/s00396-003-0871-8
    Google Scholar
    30
    Surface properties of polyethylene after low-temperature plasma treatment
    Drnovska, Hana; Lapcik, Lubomir; Bursikova, Vilma; Zemek, Josef; Barros-Timmons, Ana M.
    Colloid and Polymer Science (2003), 281 (11), 1025-1033CODEN: CPMSB6; ISSN:0303-402X. (Springer-Verlag)
    The effect of oxygen and ammonia plasma treatments on changes of the surface properties of linear high-d. polyethylene (HDPE) was studied. Surface energies of the polymer substrates were evaluated by contact angle measurements using Lifshitz-van der Waals acid-base approach. The surface energy of untreated HDPE is mainly contributed by Lifshitz-van der Waals interactions. After 5 min of plasma treatment, hydrogen bonds are formed on the surface, which is reflected in predominant acid-base interactions. The SEM results obtained demonstrate considerable changes of the surface roughness due to different types of the plasma gas used. Evolution of oxygen- or amino-contg. moieties was detected by XPS and ATR FT IR. The prepd. polyethylene surfaces were used as a basic support for further fabrication of novel hybrid biocomposite sandwich structures.
    >> More from SciFinder ®
    https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD3sXnvFalsLg%253D&md5=2a64635d0b05d24ef83c2ecb35a8a51d
  31. 31
    Salinas Rodríguez, B. Síntesis y funcionalización covalente de nanopartículas superparamagnéticas para imagen biomédica. Tesis Doctoral; Universidad Complutense de Madrid: Madrid, 2014 https://eprints.ucm.es/24586/1/T35165.pdf.
    Google Scholar
    There is no corresponding record for this reference.
  32. 32
    Wallemacq, P.; Armstrong, V. W.; Brunet, M.; Haufroid, V.; Holt, D. W.; Johnston, A.; Kuypers, D.; Meur, Y. L.; Marquet, P.; Oellerich, M.; Thervet, E.; Toenshoff, B.; Undre, N.; Weber, L. T.; Westley, I. S.; Mourad, M. Opportunities to Optimize Tacrolimus Therapy in Solid Organ Transplantation: Report of the European Consensus Conference. Ther. Drug Monit. 2009, 31 (2), 139– 152,  DOI: 10.1097/FTD.0b013e318198d092
    Google Scholar
    32
    Opportunities to Optimize Tacrolimus Therapy in Solid Organ Transplantation: Report of the European Consensus Conference
    Wallemacq, Pierre; Armstrong, Victor W.; Brunet, Merce; Haufroid, Vincent; Holt, David W.; Johnston, Atholl; Kuypers, Dirk; Le Meur, Yannick; Marquet, Pierre; Oellerich, Michael; Thervet, Eric; Toenshoff, Burkhand; Undre, Nas; Weber, Lutz T.; Westley, Ian S.; Mourad, Michel
    Therapeutic Drug Monitoring (2009), 31 (2), 139-152CODEN: TDMODV; ISSN:0163-4356. (Lippincott Williams & Wilkins)
    A review. In 2007, a consortium of European experts on tacrolimus (TAC) met to discuss the most recent advances in the drug/dose optimization of TAC taking into account specific clin. situations and the anal. methods currently available and drew some recommendations and guidelines to help clinicians with the practical use of the drug. Pharmacokinetic, pharmacodynamic, and more recently pharmacogenetic approaches aid physicians to individualize long-term therapies as TAC demonstrates a high degree of both between- and within-individual variability, which may result in an increased risk of therapeutic failure if all patients are administered a uniform dose. TAC has undoubtedly benefited from therapeutic drug monitoring, but interpretation of the blood concn. is confounded by the relative differences between the assays. Single time points, limited sampling strategies, and area under concn.-time curve have all been considered to det. the most appropriate sampling procedure that correlates with efficacy. Therapeutic trough TAC concn. ranges have changed since the initial introduction of the drug, while still maintaining adequate immunosuppression and avoiding drug-related adverse effects. Pharmacodynamic markers have also been considered advantageous to the clinician, which may better reflect efficacy and safety, taking into account the between-individual variability rather than whole blood concns. The choice of method, differences between methods, and potential pitfalls of the method should all be considered when detg. TAC concns. The recommendations of this consensus meeting regarding the anal. methods include the following: encourage the development and promote the use of anal. methods displaying a lower limit of quantification (1 ng/mL), perform careful validation when implementing a new anal. assay, participate in external proficiency testing programs, promote the use of certified material as calibrators in high-performance liq. chromatog. with mass spectrometric detection methods, and take account of the assay and intermethod bias when comparing clin. trial outcomes. It is also important to consider that TAC concns. may also be influenced by other factors such as specific pharmacokinetic characteristics assocd. with the population, drug interactions, pharmacogenetics, adverse events that may alter TAC concns., and any change in the oral formulation that may result in pharmacokinetic changes. This meeting emphasized the importance of obtaining multicenter prospective trials to assess the efficacy of alternative strategies to TAC trough concns. whether it is other single time points or area under the concn.-time curve Bayesian estn. using limited sampling strategies and to select, standardize, and validate routine biomarkers of TAC pharmacodynamics.
    >> More from SciFinder ®
    https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD1MXjsFWqsb0%253D&md5=60985c4ef122f49292866a53826a9b13
  33. 33
    Wu, F.-B.; Yang, Y.-Y.; Wang, X.-B.; Wang, Z.; Zhang, W.-W.; Liu, Z.-Y.; Qian, Y.-Q. A Sample Processing Method for Immunoassay of Whole Blood Tacrolimus. Anal. Biochem. 2019, 576, 13– 19,  DOI: 10.1016/j.ab.2019.04.006
    Google Scholar
    There is no corresponding record for this reference.
  34. 34
    Salis, F.; Descalzo, A. B.; Benito-Peña, E.; Moreno-Bondi, M. C.; Orellana, G. Highly Fluorescent Magnetic Nanobeads with a Remarkable Stokes Shift as Labels for Enhanced Detection in Immunoassays. Small 2018, 14 (20), 1703810,  DOI: 10.1002/smll.201703810
    Google Scholar
    There is no corresponding record for this reference.

Cited By

Click to copy section linkSection link copied!

This article has not yet been cited by other publications.

Download PDF
Open PDF
Go to Analytical Chemistry
Get e-Alerts
Get e-Alerts

Analytical Chemistry

Cite this: Anal. Chem. 2024, 96, 35, 14142–14149
Click to copy citationCitation copied!
https://doi.org/10.1021/acs.analchem.4c02028
Published August 22, 2024

Copyright © 2024 The Authors. Published by American Chemical Society. This publication is licensed under

CC-BY 4.0 .

Article Views

454

Altmetric

-

Citations

-
Learn about these metrics

Article Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.

Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.

The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated.

  • Abstract

    High Resolution Image
    Download MS PowerPoint Slide

    Figure 1

    Figure 1. Workflow of the developed competitive fluorescence immunoassay for the detection of FK506. (Left) Activation of the COC waveguides through the amino dextran-lipase conjugate and subsequent covalent binding of FK506-CO2H. (Right) Measurement and image acquisition: The array was then incubated with the sample containing a mixture of a positive control and anti-FK506 antibodies, followed by an incubation with the labeled detection antibodies.

    High Resolution Image
    Download MS PowerPoint Slide

    Figure 2

    Figure 2. AFM topography of the planar Zeonor surface used as a waveguide: (a) original Zeonor substrate; (b) Zeonor functionalized with the ADLC solution (50 μg/mL, 30 min). (c, d) Topographic profiles along panels (a) and (b), respectively.

    High Resolution Image
    Download MS PowerPoint Slide

    Figure 3

    Figure 3. (a) Comparison of the efficiency of different amino dextran-BTL2 coating solutions at 10, 50, and 100 μg/mL. The conjugate-activated slide was patterned with FK506-CO2H (100 μg/mL) and incubated with 0.5 μg/mL anti-FK506 in the absence of free analyte. A 2.5 μg/mL Alexa Fluor 647-anti-IgM solution was used as the developer agent. (b) Biochip image obtained using two different methods for covalent coupling of FK506-CO2H: the one-step method (EDC) (blue) and the two-step method (EDC/NHS) (green). (c) Competitive calibration curve obtained at 62.5 (black square), 100 (red circle), and 200 μM (blue triangle) of EDC, using a fixed concentration of 50 mM NHS and 100 μg/mL FK506-CO2H. (d) Competitive calibration curve obtained at 50 (black circle), 100 (red diamond), and 200 μg/mL (blue triangle) of FK506-CO2H, with fixed concentrations of 50 mM NHS and 62.5 μM EDC. The assays were performed with FK506 (0.0001–100 μg/L) in the presence of 0.5 μg/mL anti-FK506 and antibiotin antibodies. A mixture of 2.5 μg/mL Alexa Fluor647-anti-IgM and Alexa Fluor 647-anti-IgG was used as the tracer solution. The results are mean signals ± standard errors of the mean (n ≥ 3).

    High Resolution Image
    Download MS PowerPoint Slide

    Figure 4

    Figure 4. (a) Biochip image using different blocking buffers. Zeonor slide was patterned with 50 μg/mL of aminated dextran-BTL2 conjugate and 100 μg/mL of FK506-CO2H (immunoassay region), 30 μg/mL biotin (as positive control, green “+” symbol), or MES buffer (as negative control, red “–” symbol). B0 (0 ng/mL FK506) and B1 (100 ng/mL FK506) and using 2.5 μg/mL Alexa Fluor 647-anti-IgM as a revealing agent. (b) Comparison of the efficiency of the blocking buffer solutions tested on the B1/B0 ratio (open symbols). The blue bars correspond to the fluorescence signal in the absence of FK506 (B0), the red bars correspond to the signal in the presence of 100 ng/mL of FK506 (B), the green bars correspond to the signal of the background, and the purple circles correspond to the B/B0 ratio. Results are shown as mean ± standard error of the mean (n ≥ 4).

    High Resolution Image
    Download MS PowerPoint Slide

    Figure 5

    Figure 5. Competitive calibration curve. The assay was performed with FK506 (0.0001–100 μg/L) in the presence of 0.5 μg/mL of anti-FK506 and antibiotin antibodies. A mixture of 2.5 μg/mL Alexa Fluor 647-anti-IgM and Alexa Fluor 647-anti-IgG was used as the tracer solution. Results are shown as the mean ± standard error of the mean (n ≥ 3).

    High Resolution Image
    Download MS PowerPoint Slide

  • References

    This article references 34 other publications.

    1. 1
      Tanaka, H.; Kuroda, A.; Marusawa, H.; Hatanaka, H.; Kino, T.; Goto, T.; Hashimoto, M.; Taga, T. Structure of FK506, a Novel Immunosuppressant Isolated from Streptomyces. J. Am. Chem. Soc. 1987, 109 (16), 5031– 5033,  DOI: 10.1021/ja00250a050
      1
      Structure of FK506, a novel immunosuppressant isolated from Streptomyces
      Tanaka, Hirokazu; Kuroda, Akio; Marusawa, Hiroshi; Hatanaka, Hiroshi; Kino, Toru; Goto, Toshio; Hashimoto, Masashi; Taga, Toru
      Journal of the American Chemical Society (1987), 109 (16), 5031-3CODEN: JACSAT; ISSN:0002-7863.
      The structure I of FK506, a novel macrocyclic lactone isolated from Streptomyces tsukubaensis No. 9993 and which has immunosuppressant activity, was deduced on the basis of chem. and spectroscopic evidence. The relative configuration was detd. by an x-ray crystal anal. of I and the abs. configuration was established by the fact that hydrolysis of I gave L-pipecolic acid. The structure is unique in respect of contg. a hemiketal-masked α,β-diketoamide functionality incorporated in a 23-membered ring.
      >> More from SciFinder ®
      https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADyaL2sXlslGktr8%253D&md5=ca5492f7d8762dffc2e507d0345fa943
    2. 2
      Bittersohl, H.; Schniedewind, B.; Christians, U.; Luppa, P. B. A Simple and Highly Sensitive On-Line Column Extraction Liquid Chromatography-Tandem Mass Spectrometry Method for the Determination of Protein-Unbound Tacrolimus in Human Plasma Samples. J. Chromatogr. A 2018, 1547, 45– 52,  DOI: 10.1016/j.chroma.2018.03.010
      There is no corresponding record for this reference.
    3. 3
      Tombelli, S.; Trono, C.; Berneschi, S.; Berrettoni, C.; Giannetti, A.; Bernini, R.; Persichetti, G.; Testa, G.; Orellana, G.; Salis, F.; Weber, S.; Luppa, P. B.; Porro, G.; Quarto, G.; Schubert, M.; Berner, M.; Freitas, P. P.; Cardoso, S.; Franco, F.; Silverio, V.; Lopez-Martinez, M.; Hilbig, U.; Freudenberger, K.; Gauglitz, G.; Becker, H.; Gärtner, C.; O’Connell, M. T.; Baldini, F. An Integrated Device for Fast and Sensitive Immunosuppressant Detection. Anal. Bioanal. Chem. 2022, 414 (10), 3243– 3255,  DOI: 10.1007/s00216-021-03847-x
      There is no corresponding record for this reference.
    4. 4
      Fujita, R.; Nagatoishi, S.; Adachi, S.; Nishioka, H.; Ninomiya, H.; Kaya, T.; Takai, M.; Arakawa, T.; Tsumoto, K. Control of Protein Adsorption to Cyclo Olefin Polymer by the Hofmeister Effect. J. Pharm. Sci. 2019, 108 (5), 1686– 1691,  DOI: 10.1016/j.xphs.2018.12.023
      There is no corresponding record for this reference.
    5. 5
      ZEONEX® manufacturer; https://www.zeonex.com/pharmaceuticals.aspx.html (accessed 2024–07–29).
      There is no corresponding record for this reference.
    6. 6
      Agha, A.; Waheed, W.; Alamoodi, N.; Mathew, B.; Alnaimat, F.; Abu-Nada, E.; Abderrahmane, A.; Alazzam, A. A Review of Cyclic Olefin Copolymer Applications in Microfluidics and Microdevices. Macromol. Mater. Eng. 2022, 307 (8), 2200053,  DOI: 10.1002/mame.202200053
      6
      A Review of Cyclic Olefin Copolymer Applications in Microfluidics and Microdevices
      Agha, Abdulrahman; Waheed, Waqas; Alamoodi, Nahla; Mathew, Bobby; Alnaimat, Fadi; Abu-Nada, Eiyad; Abderrahmane, Aissa; Alazzam, Anas
      Macromolecular Materials and Engineering (2022), 307 (8), 2200053CODEN: MMENFA; ISSN:1438-7492. (Wiley-VCH Verlag GmbH & Co. KGaA)
      A review. Cyclic olefin copolymers (COC) are amorphous, transparent thermoplastics composed of cyclic olefin monomers (norbornene) and linear olefins (ethene). They are increasingly utilized as fabrication materials for microsystems and microfluidic devices, owing to their promising features of low water absorption, high elec. insulation, long-term stability of surface treatments, and resistance to a broad variety of acids and solvents. Many manufg. processes for COC-based devices have been developed in recent decades. These methodologies are categorized as replication methods or fast prototyping as common in fabrication of thermoplastic microfluidic devices. This review gives a full discussion of the features of COCs, the various prodn. processes, and the numerous selected applications in microfluidic platforms. The review also explores COCs compn. and fundamental features, as well as fabrication processes and applications in a variety of fields, investigates the materials potential advantages and uses, and attempts to create a comprehensive list of COCs possible benefits. Due to their unique features and simplicity of fabrication, COCs are projected to advance the future of microfluidics, microsystems, and optofluidics.
      >> More from SciFinder ®
      https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BB38XhtFGgtLbK&md5=c4c0b3795bdbabad3fb784d5ecac9e51
    7. 7
      Nunes, P. S.; Ohlsson, P. D.; Ordeig, O.; Kutter, J. P. Cyclic Olefin Polymers: Emerging Materials for Lab-on-a-Chip Applications. Microfluid. Nanofluidics 2010, 9 (2), 145– 161,  DOI: 10.1007/s10404-010-0605-4
      There is no corresponding record for this reference.
    8. 8
      Raj, J.; Herzog, G.; Manning, M.; Volcke, C.; MacCraith, B. D.; Ballantyne, S.; Thompson, M.; Arrigan, D. W. M. Surface Immobilisation of Antibody on Cyclic Olefin Copolymer for Sandwich Immunoassay. Biosens. Bioelectron. 2009, 24 (8), 2654– 2658,  DOI: 10.1016/j.bios.2009.01.026
      There is no corresponding record for this reference.
    9. 9
      Laib, S.; MacCraith, B. D. Immobilization of Biomolecules on Cycloolefin Polymer Supports. Anal. Chem. 2007, 79 (16), 6264– 6270,  DOI: 10.1021/ac062420y
      9
      Immobilization of Biomolecules on Cycloolefin Polymer Supports
      Laib, Stephan; MacCraith, Brian D.
      Analytical Chemistry (Washington, DC, United States) (2007), 79 (16), 6264-6270CODEN: ANCHAM; ISSN:0003-2700. (American Chemical Society)
      Recent trends in the development of microfluidic and biodiagnostic chips favor polymer materials over glass, primarily for optical and economical reasons. Therefore, existing chem. methods to prep. biomol. microarrays on glass slides have to be adapted or replaced to suit polymer substrates. Here the authors present a strategy to immobilize DNA and antibodies on cyclic polyolefin slides, like Zeonor. This polymer represents a class of new polymeric materials with excellent optical and mech. properties. By plasma and liq. chem. treatment followed by coating with polyelectrolytes, the authors have succeeded in immobilizing DNA onto the polymer substrate, yielding stable and versatile biosensor surfaces. The authors demonstrate the stability and usage of the coated Zeonor substrates not only in DNA chip technol. but also in protein chip technol. with DNA-directed immobilization of proteins.
      >> More from SciFinder ®
      https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2sXnsVyms7o%253D&md5=30075423ea8fb8929c3a64720a818ee0
    10. 10
      van Midwoud, P. M.; Janse, A.; Merema, M. T.; Groothuis, G. M. M.; Verpoorte, E. Comparison of Biocompatibility and Adsorption Properties of Different Plastics for Advanced Microfluidic Cell and Tissue Culture Models. Anal. Chem. 2012, 84 (9), 3938– 3944,  DOI: 10.1021/ac300771z
      10
      Comparison of Biocompatibility and Adsorption Properties of Different Plastics for Advanced Microfluidic Cell and Tissue Culture Models
      van Midwoud, Paul M.; Janse, Arnout; Merema, Marjolijn T.; Groothuis, Geny M. M.; Verpoorte, Elisabeth
      Analytical Chemistry (Washington, DC, United States) (2012), 84 (9), 3938-3944CODEN: ANCHAM; ISSN:0003-2700. (American Chemical Society)
      Microfluidic technol. is providing new routes toward advanced cell and tissue culture models to better understand human biol. and disease. Many advanced devices have been made from poly(dimethylsiloxane) (PDMS) to enable expts., for example, to study drug metab. by use of precision-cut liver slices, that are not possible with conventional systems. However, PDMS, a silicone rubber material, is very hydrophobic and tends to exhibit significant adsorption and absorption of hydrophobic drugs and their metabolites. Although glass could be used as an alternative, thermoplastics are better from a cost and fabrication perspective. Thermoplastic polymers (plastics) allow easy surface treatment and are generally transparent and biocompatible. This study focuses on the fabrication of biocompatible microfluidic devices with low adsorption properties from the thermoplastics poly(Me methacrylate) (PMMA), polystyrene (PS), polycarbonate (PC), and cyclic olefin copolymer (COC) as alternatives for PDMS devices. Thermoplastic surfaces were oxidized using UV-generated ozone or oxygen plasma to reduce adsorption of hydrophobic compds. Surface hydrophilicity was assessed over 4 wk by measuring the contact angle of water on the surface. The adsorption of 7-ethoxycoumarin, testosterone, and their metabolites was also detd. after UV-ozone treatment. Biocompatibility was assessed by culturing human hepatoma (HepG2) cells on treated surfaces. Comparison of the adsorption properties and biocompatibility of devices in different plastics revealed that only UV-ozone-treated PC and COC devices satisfied both criteria. This paper lays an important foundation that will help researchers make informed decisions with respect to the materials they select for microfluidic cell-based culture expts.
      >> More from SciFinder ®
      https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC38XksFWhu7k%253D&md5=a082cf6db9e72c5dc6a9adff2c033f72
    11. 11
      Peng, X.; Zhao, L.; Du, G.; Wei, X.; Guo, J.; Wang, X.; Guo, G.; Pu, Q. Charge Tunable Zwitterionic Polyampholyte Layers Formed in Cyclic Olefin Copolymer Microchannels through Photochemical Graft Polymerization. ACS Appl. Mater. Interfaces 2013, 5 (3), 1017– 1023,  DOI: 10.1021/am3027019
      There is no corresponding record for this reference.
    12. 12
      Hager, R.; Forsich, C.; Duchoslav, J.; Burgstaller, C.; Stifter, D.; Weghuber, J.; Lanzerstorfer, P. Microcontact Printing of Biomolecules on Various Polymeric Substrates: Limitations and Applicability for Fluorescence Microscopy and Subcellular Micropatterning Assays. ACS Appl. Polym. Mater. 2022, 4 (10), 6887– 6896,  DOI: 10.1021/acsapm.2c00834
      There is no corresponding record for this reference.
    13. 13
      Khonina, S. N.; Voronkov, G. S.; Grakhova, E. P.; Kazanskiy, N. L.; Kutluyarov, R. V.; Butt, M. A. Polymer Waveguide-Based Optical Sensors─Interest in Bio, Gas, Temperature, and Mechanical Sensing Applications. Coatings 2023, 13 (3), 549,  DOI: 10.3390/coatings13030549
      13
      Polymer Waveguide-Based Optical Sensors-Interest in Bio, Gas, Temperature, and Mechanical Sensing Applications
      Khonina, Svetlana N.; Voronkov, Grigory S.; Grakhova, Elizaveta P.; Kazanskiy, Nikolay L.; Kutluyarov, Ruslan V.; Butt, Muhammad A.
      Coatings (2023), 13 (3), 549CODEN: COATED; ISSN:2079-6412. (MDPI AG)
      A review. In the realization of photonic integrated devices, materials such as polymers are crucial. Polymers have shown compatibility with several patterning techniques, are generally affordable, and may be functionalized to obtain desired optical, elec., or mech. characteristics. Polymer waveguides are a viable platform for optical connectivity since they are easily adaptable to on-chip and on-board integration and promise low propagation losses <1 dB/cm. Furthermore, polymer waveguides can be made to be extremely flexible, able to withstand bending, twisting, and even stretching. Optical sensing is an interesting field of research that is gaining popularity in polymer photonics. Due to its huge potential for use in several industries, polymer waveguide-based sensors have attracted a lot of attention. Due to their resilience to electromagnetic fields, optical sensors operate better in difficult situations, such as those found in elec. power generating and conversion facilities. In this review, the most widely used polymer materials are discussed for integrated photonics. Moreover, four significant sensing applications of polymer-waveguide based sensors which include biosensing, gas sensing, temp. sensing and mech. sensing have been debated.
      >> More from SciFinder ®
      https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BB3sXmvFeqtrk%253D&md5=b622aea04cf66d36495384943e629c8d
    14. 14
      Encinas, N.; Díaz-Benito, B.; Abenojar, J.; Martínez, M. A. Extreme Durability of Wettability Changes on Polyolefin Surfaces by Atmospheric Pressure Plasma Torch. Surf. Coat. Technol. 2010, 205 (2), 396– 402,  DOI: 10.1016/j.surfcoat.2010.06.069
      There is no corresponding record for this reference.
    15. 15
      Herron, J. N.; Wang, H.-K.; Tan, L.; Brown, S. Z.; Terry, A. H.; Tolley, S. E.; Durtschi, J. D.; Simon, E. M.; Astill, M. E.; Smith, R. S.; Christensen, D. A. Planar Waveguide Biosensors for Point-of-Care Clinical and Molecular Diagnostics. In Fluorescence Sensors and Biosensors; Thompson, R. B., Ed.; CRC Press: Boca Raton, 2005.  DOI: 10.1201/9781420028287 .
      There is no corresponding record for this reference.
    16. 16
      Yan, J.; Zhao, C.; Ma, Y.; Yang, W. Covalently Attaching Hollow Silica Nanoparticles on a COC Surface for the Fabrication of a Three-Dimensional Protein Microarray. Biomacromolecules 2022, 23 (6), 2614– 2623,  DOI: 10.1021/acs.biomac.2c00354
      There is no corresponding record for this reference.
    17. 17
      Marciello, M.; Bolivar, J. M.; Filice, M.; Mateo, C.; Guisan, J. M. Preparation of Lipase-Coated, Stabilized, Hydrophobic Magnetic Particles for Reversible Conjugation of Biomacromolecules. Biomacromolecules 2013, 14 (3), 602– 607,  DOI: 10.1021/bm400032q
      There is no corresponding record for this reference.
    18. 18
      Schmidt-Dannert, C.; Rúa, M. L.; Atomi, H.; Schmid, R. D. Thermoalkalophilic Lipase of Bacillus Thermocatenulatus. I. Molecular Cloning, Nucleotide Sequence, Purification and Some Properties. Biochim. Biophys. Acta BBA - Lipids Lipid Metab. 1996, 1301 (1), 105– 114,  DOI: 10.1016/0005-2760(96)00027-6
      There is no corresponding record for this reference.
    19. 19
      Yenenler, A.; Venturini, A.; Burduroglu, H. C.; Sezerman, O. U. Investigating the Structural Properties of the Active Conformation BTL2 of a Lipase from Geobacillus Thermocatenulatus in Toluene Using Molecular Dynamic Simulations and Engineering BTL2 via In-Silico Mutation. J. Mol. Model. 2018, 24 (9), 229,  DOI: 10.1007/s00894-018-3753-1
      There is no corresponding record for this reference.
    20. 20
      Herranz, S.; Marciello, M.; Olea, D.; Hernández, M.; Domingo, C.; Vélez, M.; Gheber, L. A.; Guisán, J. M.; Moreno-Bondi, M. C. Dextran–Lipase Conjugates as Tools for Low Molecular Weight Ligand Immobilization in Microarray Development. Anal. Chem. 2013, 85 (15), 7060– 7068,  DOI: 10.1021/ac400631t
      There is no corresponding record for this reference.
    21. 21
      Herranz, S.; Marciello, M.; Marco, M.-P.; Garcia-Fierro, J. L.; Guisan, J. M.; Moreno-Bondi, M. C. Multiplex Environmental Pollutant Analysis Using an Array Biosensor Coated with Chimeric Hapten-Dextran-Lipase Constructs. Sens. Actuators B Chem. 2018, 257, 256– 262,  DOI: 10.1016/j.snb.2017.10.134
      There is no corresponding record for this reference.
    22. 22
      Hermanson, G. T. The Chemistry of Reactive Groups. In Bioconjugate Techniques (Second ed.); Hermanson, G. T., Ed.; Academic Press: New York, 2008; pp 169– 212.  DOI: 10.1016/B978-0-12-370501-3.00002-3 .
      There is no corresponding record for this reference.
    23. 23
      Herranz, S.; Marazuela, M. D.; Moreno-Bondi, M. C. Automated Portable Array Biosensor for Multisample Microcystin Analysis in Freshwater Samples. Biosens. Bioelectron. 2012, 33 (1), 50– 55,  DOI: 10.1016/j.bios.2011.12.016
      There is no corresponding record for this reference.
    24. 24
      Glahn-Martínez, B.; Jurado-Sánchez, B.; Benito-Peña, E.; Escarpa, A.; Moreno-Bondi, M. C. Magnetic Janus Micromotors for Fluorescence Biosensing of Tacrolimus in Oral Fluids. Biosens. Bioelectron. 2024, 244, 115796  DOI: 10.1016/j.bios.2023.115796
      There is no corresponding record for this reference.
    25. 25
      Munzert, P.; Schulz, U.; Kaiser, N. Transparent Thermoplastic Polymers in Plasma-Assisted Coating Processes. Surf. Coat. Technol. 2003, 174–175, 1048– 1052,  DOI: 10.1016/S0257-8972(03)00414-6
      There is no corresponding record for this reference.
    26. 26
      Voigt, J. Photochemische und photooxydative Reaktionen. In Die Stabilisierung der Kunststoffe gegen Licht und Wärme; Springer-Verlag: Berlin, Heidelberg, NY, 1966; pp 67– 73.  DOI: 10.1007/978-3-642-52097-6 .
      There is no corresponding record for this reference.
    27. 27
      Jena, R. K.; Yue, C. Y. Cyclic Olefin Copolymer Based Microfluidic Devices for Biochip Applications: Ultraviolet Surface Grafting Using 2-Methacryloyloxyethyl Phosphorylcholine. Biomicrofluidics 2012, 6 (1), 012822  DOI: 10.1063/1.3682098
      27
      Cyclic olefin copolymer based microfluidic devices for biochip applications: Ultraviolet surface grafting using 2-methacryloyloxyethyl phosphorylcholine
      Jena, Rajeeb K.; Yue, C. Y.
      Biomicrofluidics (2012), 6 (1), 012822, 13 pp.CODEN: BIOMGB; ISSN:1932-1058. (American Institute of Physics)
      This report studies the surface modification of cyclic olefin copolymer (COC) by 2-methacryloyloxyethyl phosphorylcholine (MPC) monomer using photografting technique for the purpose of biointerface applications, which demonstrate resistance to both protein adsorption and cell adhesion in COC-based microfluidic devices. This is essential because the hydrophobic nature of COC can lead to adsorption of specific compds. from biol. fluids in the microchannel, which can affect the results during fluidic anal. and cause clogging inside the microchannel. A correlation was found between the irradn. time and hydrophobicity of the modified substrate. Static water contact angle results show that the hydrophilicity property of the MPC-grafted substrate improves with increasing irradn. time. The contact angle of the modified surface decreased to 20 ± 5 ° from 88 ± 3 ° for the untreated substrate. The surface characterization of the modified surface was evaluated using XPS and Fourier transform IR spectroscopy (FTIR spectroscopy). Attenuated total reflection-FTIR and XPS results show the presence of the phosphate group (P-O) on modified COC substrates, indicating that the hydrophilic MPC monomer has successfully grafted on COC. Finally, it was demonstrated that cell adhesion and protein adsorption on the MPC modified COC specimen has reduced significantly. (c) 2012 American Institute of Physics.
      >> More from SciFinder ®
      https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC38XhsFSktrjI&md5=6d12e28f8ba6a874e880aac2de4f4e33
    28. 28
      Ma, H.; Davis, R. H.; Bowman, C. N. A Novel Sequential Photoinduced Living Graft Polymerization. Macromolecules 2000, 33 (2), 331– 335,  DOI: 10.1021/ma990821s
      28
      A Novel Sequential Photoinduced Living Graft Polymerization
      Ma, Huimin; Davis, Robert H.; Bowman, Christopher N.
      Macromolecules (2000), 33 (2), 331-335CODEN: MAMOBX; ISSN:0024-9297. (American Chemical Society)
      A novel sequential UV-induced living graft polymn. method has been designed and investigated to modify polymeric materials. This method consists of two steps. In the first step, a surface initiator is formed on a substrate under UV irradn. in the presence of benzophenone (BP) solns.; in the second step, the monomers are grafted to the substrate by a living polymn. initiated by the surface photoinitiator. Hydrophobic porous polypropylene (PP) membranes were made hydrophilic and with neg. charged surface by grafting acrylic acid (AA). Exptl. results demonstrated that grafting d. and graft polymer chain length can be controlled by choosing the reaction conditions in the first step and in the subsequent step(s) independently. The amt. of grafted polymer relative to the total amt. of polymer from the novel sequential photoinduced graft polymn. method is 4-fold greater than that of the simultaneous grafting method for the system studied. In addn., a reaction mechanism was proposed and confirmed in the expts. With regard to the surface initiator formation caused by hydrogen abstraction, the kinetic studies show that the reaction rate has a max. value which depends on BP concn. With regard to the graft polymn. in the second step, there is a linear relationship between the graft polymn. rate and the monomer concn.
      >> More from SciFinder ®
      https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADyaK1MXotFKgtbc%253D&md5=6e86cbce87fa8d54d5495adfa46a27b1
    29. 29
      Stachowiak, T. B.; Svec, F.; Fréchet, J. M. J. Patternable Protein Resistant Surfaces for Multifunctional Microfluidic Devices via Surface Hydrophilization of Porous Polymer Monoliths Using Photografting. Chem. Mater. 2006, 18 (25), 5950– 5957,  DOI: 10.1021/cm0617034
      There is no corresponding record for this reference.
    30. 30
      Drnovská, H.; Lapčík, L.; Buršíková, V.; Zemek, J.; Barros-Timmons, A. M. Surface Properties of Polyethylene after Low-Temperature Plasma Treatment. Colloid Polym. Sci. 2003, 281 (11), 1025– 1033,  DOI: 10.1007/s00396-003-0871-8
      30
      Surface properties of polyethylene after low-temperature plasma treatment
      Drnovska, Hana; Lapcik, Lubomir; Bursikova, Vilma; Zemek, Josef; Barros-Timmons, Ana M.
      Colloid and Polymer Science (2003), 281 (11), 1025-1033CODEN: CPMSB6; ISSN:0303-402X. (Springer-Verlag)
      The effect of oxygen and ammonia plasma treatments on changes of the surface properties of linear high-d. polyethylene (HDPE) was studied. Surface energies of the polymer substrates were evaluated by contact angle measurements using Lifshitz-van der Waals acid-base approach. The surface energy of untreated HDPE is mainly contributed by Lifshitz-van der Waals interactions. After 5 min of plasma treatment, hydrogen bonds are formed on the surface, which is reflected in predominant acid-base interactions. The SEM results obtained demonstrate considerable changes of the surface roughness due to different types of the plasma gas used. Evolution of oxygen- or amino-contg. moieties was detected by XPS and ATR FT IR. The prepd. polyethylene surfaces were used as a basic support for further fabrication of novel hybrid biocomposite sandwich structures.
      >> More from SciFinder ®
      https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD3sXnvFalsLg%253D&md5=2a64635d0b05d24ef83c2ecb35a8a51d
    31. 31
      Salinas Rodríguez, B. Síntesis y funcionalización covalente de nanopartículas superparamagnéticas para imagen biomédica. Tesis Doctoral; Universidad Complutense de Madrid: Madrid, 2014 https://eprints.ucm.es/24586/1/T35165.pdf.
      There is no corresponding record for this reference.
    32. 32
      Wallemacq, P.; Armstrong, V. W.; Brunet, M.; Haufroid, V.; Holt, D. W.; Johnston, A.; Kuypers, D.; Meur, Y. L.; Marquet, P.; Oellerich, M.; Thervet, E.; Toenshoff, B.; Undre, N.; Weber, L. T.; Westley, I. S.; Mourad, M. Opportunities to Optimize Tacrolimus Therapy in Solid Organ Transplantation: Report of the European Consensus Conference. Ther. Drug Monit. 2009, 31 (2), 139– 152,  DOI: 10.1097/FTD.0b013e318198d092
      32
      Opportunities to Optimize Tacrolimus Therapy in Solid Organ Transplantation: Report of the European Consensus Conference
      Wallemacq, Pierre; Armstrong, Victor W.; Brunet, Merce; Haufroid, Vincent; Holt, David W.; Johnston, Atholl; Kuypers, Dirk; Le Meur, Yannick; Marquet, Pierre; Oellerich, Michael; Thervet, Eric; Toenshoff, Burkhand; Undre, Nas; Weber, Lutz T.; Westley, Ian S.; Mourad, Michel
      Therapeutic Drug Monitoring (2009), 31 (2), 139-152CODEN: TDMODV; ISSN:0163-4356. (Lippincott Williams & Wilkins)
      A review. In 2007, a consortium of European experts on tacrolimus (TAC) met to discuss the most recent advances in the drug/dose optimization of TAC taking into account specific clin. situations and the anal. methods currently available and drew some recommendations and guidelines to help clinicians with the practical use of the drug. Pharmacokinetic, pharmacodynamic, and more recently pharmacogenetic approaches aid physicians to individualize long-term therapies as TAC demonstrates a high degree of both between- and within-individual variability, which may result in an increased risk of therapeutic failure if all patients are administered a uniform dose. TAC has undoubtedly benefited from therapeutic drug monitoring, but interpretation of the blood concn. is confounded by the relative differences between the assays. Single time points, limited sampling strategies, and area under concn.-time curve have all been considered to det. the most appropriate sampling procedure that correlates with efficacy. Therapeutic trough TAC concn. ranges have changed since the initial introduction of the drug, while still maintaining adequate immunosuppression and avoiding drug-related adverse effects. Pharmacodynamic markers have also been considered advantageous to the clinician, which may better reflect efficacy and safety, taking into account the between-individual variability rather than whole blood concns. The choice of method, differences between methods, and potential pitfalls of the method should all be considered when detg. TAC concns. The recommendations of this consensus meeting regarding the anal. methods include the following: encourage the development and promote the use of anal. methods displaying a lower limit of quantification (1 ng/mL), perform careful validation when implementing a new anal. assay, participate in external proficiency testing programs, promote the use of certified material as calibrators in high-performance liq. chromatog. with mass spectrometric detection methods, and take account of the assay and intermethod bias when comparing clin. trial outcomes. It is also important to consider that TAC concns. may also be influenced by other factors such as specific pharmacokinetic characteristics assocd. with the population, drug interactions, pharmacogenetics, adverse events that may alter TAC concns., and any change in the oral formulation that may result in pharmacokinetic changes. This meeting emphasized the importance of obtaining multicenter prospective trials to assess the efficacy of alternative strategies to TAC trough concns. whether it is other single time points or area under the concn.-time curve Bayesian estn. using limited sampling strategies and to select, standardize, and validate routine biomarkers of TAC pharmacodynamics.
      >> More from SciFinder ®
      https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD1MXjsFWqsb0%253D&md5=60985c4ef122f49292866a53826a9b13
    33. 33
      Wu, F.-B.; Yang, Y.-Y.; Wang, X.-B.; Wang, Z.; Zhang, W.-W.; Liu, Z.-Y.; Qian, Y.-Q. A Sample Processing Method for Immunoassay of Whole Blood Tacrolimus. Anal. Biochem. 2019, 576, 13– 19,  DOI: 10.1016/j.ab.2019.04.006
      There is no corresponding record for this reference.
    34. 34
      Salis, F.; Descalzo, A. B.; Benito-Peña, E.; Moreno-Bondi, M. C.; Orellana, G. Highly Fluorescent Magnetic Nanobeads with a Remarkable Stokes Shift as Labels for Enhanced Detection in Immunoassays. Small 2018, 14 (20), 1703810,  DOI: 10.1002/smll.201703810
      There is no corresponding record for this reference.

  • Supporting Information

    Supporting Information

    The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acs.analchem.4c02028.

    • Details about the COC surface modification by oxygen plasma, photochemical grafting, and amino dextran-lipase conjugate; details about the biosensor development; scheme of the FK506 immunoassay protocol; comparison of the hydrophobicity of Zeonor before and after the plasma treatment; biochip images for the analysis of FK506 using the developed immunoarray; ATR-FTIR results; comparative summary of different methods reported for COC surface activation (PDF)


    Terms & Conditions

    Most electronic Supporting Information files are available without a subscription to ACS Web Editions. Such files may be downloaded by article for research use (if there is a public use license linked to the relevant article, that license may permit other uses). Permission may be obtained from ACS for other uses through requests via the RightsLink permission system: http://pubs.acs.org/page/copyright/permissions.html.