Autonomous, Real-Time Monitoring Electrochemical Aptasensor for Circadian Tracking of Cortisol Hormone in Sub-microliter Volumes of Passively Eluted Human Sweat

Abstract

The proposed work involves the development of an autonomous, label-free electrochemical sensor for real-time monitoring of cortisol levels expressed naturally in sub-microliter sweat volumes, for prolonged sensing periods of ∼8 h. Highly specific single-stranded DNA (ssDNA) aptamer is used for affinity capture of cortisol hormone eluted in sweat dynamically. The cortisol present in sweat binds to the aptamer capture probe that changes conformation and modulates electrochemical properties at the electrode–buffer interface, which was studied using dynamic light scattering studies for the entire physiological sweat pH. Attenuated total reflectance-Fourier transform infrared spectroscopy and UV–vis spectroscopy were used to optimize the binding chemistry of the elements of the sensor stack. Nonfaradaic electrochemical impedance spectroscopy was used to calibrate the sensor for a dynamic range of 1–256 ng/mL. An R 2 of 0.97 with an output signal range of 20–50% was obtained. Dynamic cortisol level variation tracking was studied using continuous dosing experiments to calibrate the sensor for temporal variation. The sensor did not show significant susceptibility to noise due to cross-reactive interferents and nonspecific buffer constituents. The performance of the developed aptasensor was compared with the previously established cortisol immunosensor in terms of surface charge behavior and nonfaradaic biosensing. The aptamer sensor shows a higher signal-to-noise ratio, better resolution, and has a larger output range for the same input range as the cortisol immunosensor. The feasibility of deploying the developed aptasensing scheme as continuous lifestyle and performance monitors was validated through human subject studies.

Emerging wearable flexible sensors for sweat analysis

“…Their output signals can be fed back to the users either directly through circuits or by wireless communication. Various electrochemical sensing mechanisms, such as potentiometry [60,61], chronoamperometry [46,62], voltammetry [63,64], and electrochemical impedance spectroscopy [65,66], are substantially leveraged toward specific application scenarios where they all have their unique advantages.…”

Wearable electrochemical biosensors to measure biomarkers with complex blood-to-sweat partition such as proteins and hormones

“…6g ) [ 102 ]. The basal-produced sweat at the eccrine and apocrine glands is passively transported to the detection region without the need of neither accumulating sweat nor another iontophoretic-like active methodology for sweat sampling [ 103 , 104 ]. The applied small sinusoidal AC voltage induces a given capacitance over the WE that generates an output in the form of a phase-shifted current, recorded as a change in the impedance modulus (Zmod) (Table 1 ).…”

“…While aptamers as biorecognition elements are smaller than antibodies, they provide greater capacitance given the reached stacking after immobilization. Thus, aptasensor yields a more comprehensive output signal range (30 k − 5 kΩ) [ 104 ] compared with a previously reported cortisol wearable immunosensor (10 k − 5 kΩ) (Fig. 6i ) [ 103 ].…”

“…T: 2.5 μm; Functionalization: Thiol at the 5′ end of cortisol single stranded DNA aptamer is crosslinked with the ZnO SAM using DTSS Passivation: NSB was avoided using microporous substrate Technique: Non-faradaic EIS; 100 Hz; with 10 mV AC amplitude LDR: 1 − 256 ng mL −1 Time of response: 5 min Stability: > 1 week. Remains active after 8 h of use [ 103 , 104 ] Immunesensor (SPE) Sample: Sweat Analyte: Neuropeptide Y (NPY) Electrode: Gold interdigitated WE, RE and CE, printed onto nanoporous polyamide substrate Functionalization: Thiol binding between anti-NPY and WE using DSP linker Passivation: NSB was reduced by the size-based exclusion induced by the nanoporous substrate Technique: a) EIS; 1 Hz – 1 MHz; with 10 mV AC amplitude; b) ChA: DC step potential of 600 mV by 60 s LDR: 10 − 500 pg mL −1 Time of response: 1 min Stability: Not shown [ 102 ] Immunesensor (SPE) Sample: Sweat (10 μL) Analyte: IL-1β, IL-6, IL-8, IL-10, TNF-α, CRP Electrode: Interdigitated RE and WE made of ZnO printed onto nanoporous polyamide substrate Functionalization: Monoclonal antibodies cross-linked by DTSSP through thiol-bond Passivation: Tween 80 Technique: Non-faradaic EIS; 180 Hz, with 10 mV AC amplitude LDR: a) Cytokines: 0.2–200 pg·mL −1 ; b) CRP: 1–10 ng·mL −1 Time of response: Immediately Stability: > 1 month [ 55 , 109 ] MIP-FET Sample: Sweat Analyte: Cortisol Electrode: S, D & G: Ag/Cl; C: PEDOT:PSS; B: SEBS elastomer; T: 100 μm; Functionalization: MAA as Cortisol-selective polymer and EDMA as cross-linker Passivation: Non-required Technique: i) V G : 0.2 V; ii) Output: I SD ; iii) Doping polarity: n LDR: 0.1 – 10 μM Time of response: ~ 6 min without microfluidic system; 1 s with it [ 110 ] BRE-FET …”

“…The records using aptamers suggest a greater signal compared with the use of antibodies (i). Reprinted with permission from [ 55 , 104 , 105 , 109 ] …”