Tip-Enhanced Sub-Femtomolar Steroid Immunosensing via Micropyramidal Flexible Conducting Polymer Electrodes for At-Home Monitoring of Salivary Sex Hormones

Abstract

Noninvasive testing and continuous monitoring of ultralow-concentration hormones in biofluids have attracted increasing interest for health management and personalized medicine, in which saliva could fulfill the demand. Steroid sex hormones such as progesterone (P4) and β-estradiol (E2) are crucial for female wellness and reproduction; however, their concentrations in saliva can vary down to sub-pM and constantly fluctuate over several orders of magnitude. This remains a major obstacle toward user-friendly and reliable monitoring at home with low-cost flexible biosensors. Herein we introduce a 3D micropyramidal electrode architecture to address such challenges and achieve an ultrasensitive flexible electrochemical immunosensor with sub-fM-level detection capability of salivary sex hormones within a few minutes. This is enabled by micropyramidal electrode arrays consisting of a poly­(3,4-ethylenedioxythiophene):poly­(4-styrenesulfonate) (PEDOT:PSS) thin film as the coating layer and electrochemically decorated gold nanoparticles (AuNPs) to improve the antibody immobilization. The enhanced mass transport around the 3D tips provided by the micropyramidal architecture is discovered to improve the detection limit by 3 orders of magnitude, pushing it to as low as ∼100 aM for P4 and ∼20 aM for E2, along with a wide linear range up to μM. Accordingly, these hormones down to sub-fM in >1000-fold-diluted saliva samples can be accurately measured by the printed soft immunosensors, thus allowing at-home testing through simple saliva dilution to minimize the interfering substances instead of centrifugation. Finally, monitoring of the female ovarian hormone cycle of both P4 and E2 is successfully demonstrated based on the centrifuge-free saliva testing during a period of 4 weeks. This ultrasensitive and soft 3D microarchitected electrode design is believed to provide a universal platform for a diverse variety of applications spanning from accurate clinical diagnostics and counselling and in vivo detection of bioactive species to environmental and food quality tracing.

Soft Microstructure Arrays: A Multifunctional Microsystem for Mass Transport, Energy Transformation, and Surface Manipulation

“…Reproduced with permission. 103 Copyright 2023, American Chemical Society. (e) Schematic of the workflow for in situ sampling and detection of protein biomarkers in ISF based on the SMA patch, along with a picture of it showing strong fluorescence under mouse interleukin 6.…”

“…117 cantly magnifies the detection limit by greatly improving current and analytical transport. 103 Similarly, to improve the sensitivity of the immunoassay, Singamaneni et al coated the surface of polystyrene-based SMA patches with a layer of ultrabright fluorescent nanolabels called plasmonic fluorescence. The label increased the detection limit of various ISF protein biomarkers by nearly 800-fold and significantly reduced the sampling time compared to that of conventional fluorophores.…”

“…The array, encapsulated in methacrylic acid hyaluronic acid, contains a smart DNA loop system which produces an amplified fluorescent signal in response to miRNA . Xie et al enhanced detection capabilities by employing a 3D microcone tip (Figure d) on an electrochemical sensor array, coated with poly(3,4-ethylenedioxythiophene):poly(4-styrenesulfonate) (PEDOT:PSS) and Au nanoparticles; this geometry significantly magnifies the detection limit by greatly improving current and analytical transport . Similarly, to improve the sensitivity of the immunoassay, Singamaneni et al coated the surface of polystyrene-based SMA patches with a layer of ultrabright fluorescent nanolabels called plasmonic fluorescence.…”

“…(d) PEDOT:PSS-coated pyramidal SMA electrodes for testing ultralow-concentration hormones in biofluids. Reproduced with permission 103. Copyright 2023, American Chemical Society.…”