Optical Nanosensor Passivation Enables Highly Sensitive Detection of the Inflammatory Cytokine Interleukin-6

Interleukin-6 (IL-6) is known to play a critical role in the progression of inflammatory diseases such as cardiovascular disease, cancer, sepsis, viral infection, neurological disease, and autoimmune diseases. Emerging diagnostic and prognostic tools, such as optical nanosensors, experience challenges in translation to the clinic in part due to protein corona formation, dampening their selectivity and sensitivity. To address this problem, we explored the rational screening of several classes of biomolecules to be employed as agents in noncovalent surface passivation as a strategy to screen interference from nonspecific proteins. Findings from this screening were applied to the detection of IL-6 by a fluorescent-antibody-conjugated single-walled carbon nanotube (SWCNT)-based nanosensor. The IL-6 nanosensor exhibited highly sensitive and specific detection after passivation with a polymer, poly-l-lysine, as demonstrated by IL-6 detection in human serum within a clinically relevant range of 25 to 25,000 pg/mL, exhibiting a limit of detection over 3 orders of magnitude lower than prior antibody-conjugated SWCNT sensors. This work holds potential for the rapid and highly sensitive detection of IL-6 in clinical settings with future application to other cytokines or disease-specific biomarkers.


Figure S2 .
Figure S2.Change in (7,5) fluorescence peak upon challenging protein passivations with FBS.Successful screening of serum interference was exhibited (A) by none of the mass ratios of BSA passivation, n=3(B) by only 50x and 100x mass ratio for NFDM passivation, n=3, (C) by all mass ratios for casein passivation, n=3, (D) The 50x mass ratio is the lowest common passivation ratio shown to be successful amongst protein passivation, n=3,

Figure S11 .
Figure S11.Change in (7,5) fluorescence peak upon challenging polymer passivation with FBS.Successful screening of serum interference was exhibited by all mass ratios of (A) PLK passivations, n=3 and (B) PEI passivations, n=3.No mass ratios of (C) PEG passivations demonstrated successful screening, n=3, (D) 50x mass ratio shows screening effect for PLK as well as PEI passivation.

Figure S12 .
Figure S12.Change in (9,5) fluorescence peak upon challenging polymer passivation with FBS.Successful screening of serum interference was exhibited by none of the mass ratios (A) for PLK passivations, n=3, all of the mass ratios (B) for PEI passivations, n=3, mean ± SD, and none of the mass ratios (C) for PEG passivations, n=3, mean ± SD. (D) For 50x mass ratio, none of the polymer agents were successful.

Figure S14 .
Figure S14.Change in all fluorescence peaks over time after passivation after FBS addition for PEI-passivated SWCNT.(A) For 5x PEI passivation ratio, (B) For 25x PEI passivation ratio, (C) For 50x PEI passivation ratio, (D) For 100x PEI passivation ratio

Figure
Figure S17 Change in (9,5) fluorescence peak upon challenging phospholipid passivation with FBS.Successful screening of serum interference was exhibited by (A) none of the mass ratios For DSPE PEG (NH2) passivations, n=3, and (B) For 16:0 PE PEG passivations, n=3, (C) For 50x mass ratio, both the phospholipid passivations do not screen serum interference.

Figure S20 .
Figure S20.Change in (7,6) fluorescence peak intensity of SWCNT-(TAT)6 in the presence of 50x mass ratio passivation agents compared to FBS alone.(A) Only NFDM and casein of the proteins passivations screened effect of FBS intereference on fluorescence intensity of

Figure S21 .
Figure S21.Change in 1037 nm absorbance peak of SWCNT-(TAT)6 in presence of 50x mass ratio passivation agents compared to FBS alone.(A) Protein passivation agents, (B) polymer passivation agents, and (C) phospholipid passivation agents were evaluated for their ability to prevent center wavelength shift for this center wavelength.

Figure S22 .
Figure S22.Change in 1270 nm absorbance peak of SWCNT-(TAT)6 in the presence of 50x mass ratio passivation agents compared to FBS. (A) Protein passivation agents, (B) polymer passivation agents, and (C) phospholipid passivation agents were evaluated for their ability to prevent center wavelength shift for this center wavelength.Only NFDM, PLK, and PEI show shift higher than serum.1270 nm absorbance peak includes (9,5) chirality absorbance.

Figure S24 .
Figure S24.Change in center wavelength of 1130 nm absorption peak over time after addition of protein passivation agents.(A) For all BSA mass ratio passivation, (B) For all Non-fat dry milk mass ratio passivations, and (C) For all casein mass ratio passivations.

Figure S25 .
Figure S25.Change in center wavelength of 1130 nm absorption peak over time after addition of polymer passivation agents.(A) For all poly-L-lysine mass ratio passivations, (B) For all polyethylene imine mass ratio passivations, and (C) For all polyethylene glycol mass ratio passivations.

Figure S26 .
Figure S26.Change in center wavelength of 1130 nm absorption peak over time after addition of phospholipid passivation agents.(A) For all DSPE PEG (NH2) mass ratio passivations and (B) For all 16:0 PE 2000 PEG mass ratio passivations.

Figure S30 :
Figure S30: Stability of response over time in human serum.For PLK passivated (8,7) chirality, the lowest IL-6 concentrations, 25 pg/mL and 250 pg/mL show stable response over at least 3 hours, indicating stability of PLK passivation.