Abstract
We present a sensitive and low-cost immunoassay, based on a customized open-source quartz crystal microbalance coupled with graphene biointerface sensors (G-QCM), to quantify antibodies in undiluted patient serum. We demonstrate its efficacy for a specific antibody against the phospholipase A2 receptor (anti-PLA2R), which is a biomarker in primary membranous nephropathy. A novel graphene–protein biointerface was constructed by adsorbing a low concentration of denatured bovine serum albumin (dBSA) on the reduced graphene oxide (rGO) sensor surface. The dBSA film prevents the denaturation of the protein receptor on the rGO surface and serves as the cross-linker for immobilization of the receptor for anti-PLA2R antibodies on the surface. The detection limit and selectivity of this G-QCM biosensor was compared with a commercial QCM system. The G-QCM immunoassay exhibited good specificity and high sensitivity toward the target, with an order of magnitude better detection limit (of 100 ng/mL) compared to the commercial system, at a fraction of the cost and with considerable time saving. The results obtained from patient sera compared favorably with those from enzyme-linked immunosorbent assay, validating the feasibility of use in clinical applications. The multifunctional dBSA-rGO platform provides a promising biofunctionalization method for universal immunoassay and biosensors. With the advantages of inexpensive, rapid, and sensitive detection, the G-QCM sensor and instrument form an effective autoimmune disease screening tool.
Highlights
We present a sensitive and low-cost immunoassay, based on a customized opensource quartz crystal microbalance coupled with graphene biointerface sensors (G-quartz-crystal microbalance (QCM)), to quantify antibodies in undiluted patient serum
FET biosensors suffer from lack of reproducibility of the signal when experimenting in concentrated biological samples like serum due to the high sensitivity to charges in the solutions.[23−25] the manufacturing complexities of graphene FETs remains a major challenge to mass-market adoption as a medical diagnostic device.[15,23,26]
Eight different functional surfaces were used in this comparative study, namely, (1) Au, (2) GO, (3) reduced graphene oxide (rGO), (4) Au with denatured bovine serum albumin (BSA) coating (Au-denatured bovine serum albumin (dBSA)), (5) rGO with dBSA coating, (6) rGO with nondenatured BSA coating, (7) Au with selfassembled monolayer (SAM) with COOH terminal groups (Au-SAM), and (8) rGO-dBSA activated with EDC/NHS for amine coupling
Summary
We present a sensitive and low-cost immunoassay, based on a customized opensource quartz crystal microbalance coupled with graphene biointerface sensors (G-QCM), to quantify antibodies in undiluted patient serum. Several methods have been reported to functionalize graphene-based materials with protein receptors[12−15] such that their tertiary structure and binding sites are preserved and presented Noncovalent linkers, such as 1-pyrenebutanoic acid succinimide ester, N-hydroxysuccinimide (NHS) ester tripod, bovine serum albumin (BSA), and pyrene butyric acid (PBA), have been successfully used to construct a biointerface in graphene-based biosensors to detect glucose,[16] DNA,[14] proteins,[15,17] and other biomolecules.[18,19] Harnessing BSA as a biointerface layer offers various advantages including preventing non-specific binding,[17,20] improving biocompatibility,[21] ease of use,[22] and low cost.[17,22] Zhou et al suggested a one-step method using thermally denatured BSA activated with EDC/NHS to construct a graphene field-effect transistor (GFET) for detecting a cancer biomarker, carcinoembryonic antigen.[22] FET biosensors suffer from lack of reproducibility of the signal when experimenting in concentrated biological samples like serum due to the high sensitivity to charges in the solutions.[23−25] the manufacturing complexities of graphene FETs remains a major challenge to mass-market adoption as a medical diagnostic device.[15,23,26]. PMN progresses slowly, and some patients can enter a spontaneous remission phase, 30−40% of patients eventually develop end-stage renal disease or die.[40−42]
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
