Hydrogel encapsulation of a designed fluorescent protein biosensor for continuous measurements of sub-100 nanomolar nicotine
The reinforcing and addictive properties of nicotine result from concentration- and time-dependent activation, desensitization, and upregulation of nicotinic acetylcholine receptors. However, time-resolved [nicotine] measurement in people who consume nicotine is challenging, as current approaches are expensive, invasive, tedious, and discontinuous. To address the challenge of continuous nicotine monitoring in human biofluids, we report the encapsulation of a purified, previously developed fluorescent biosensor protein, iNicSnFR12, into acrylamide hydrogels and polyethylene glycol diacrylate (PEGDA) hydrogels. We optimized the hydrogels for optical clarity and straightforward slicing. With fluorescence photometry of the hydrogels in a microscope and an integrated miniscope, [nicotine] is detected within a few min at the smoking- and vaping-relevant level of 10 - 100 nM (1.62 - 16.2 ng/ml), even in a 250 {micro}m thick hydrogel at the end of 400 {micro}m dia multimode fiber optic. Concentration-response relations are consistent with previous measurements on isolated iNicSnFR12. Leaching of iN-icSnFR12 from the hydrogel and inactivation of iNicSnFR12 are minimal for several days, and nicotine can be detected for at least 10 months after casting. This work provides the molecular, photophysical, and mechanical bases for personal, wearable continuous [nicotine] monitoring, with straightforward extensions to existing, homologous "iDrugSnFR" proteins for other abused and prescribed drugs.