Ultrasensitive multiplexed immunoassay of autophagic biomarkers based on Au/rGO and Au nanocages amplifying electrochemcial signal

Abstract

A novel sandwich-assay electrochemical immunosensor for simultaneous determination of autophagic biomarkers was introduced for the first time, the gold-reduced grapheme oxide nanocomposite (Au/r-GO) set as a good conductive platform with super high specific area, and provided more binding sites for the both antibodies of Beclin-1 and LC3B-II. While Au nanocages (AuNCs) served as good conductive platform to encapsulate a large amount of redox probe and secondary antibodies for signal amplification, due to the abundant reactive oxygen functional groups on its surface. Through differential pulse voltammetry (DPV) measurements, two separate signals can be detected directly in a single run, which represent the existence of Belin-1 and LC3B-II. Under optimized conditions, the electrochemical immunosensor exhibited good sensitivity and selectivity for the simultaneous determination of Beclin-1 and LC3B-II with linear ranges of 0.1–100 ng/mL. The detection limit for Beclin-1 and LC3B-II is 0.02 and 0.03 ng/mL respectively. This method was also applied for the analysis of Beclin-1 and LC3B-II levels in experimental cellular protein lysates, and the results were in good agreement with those of enzyme linked immunosorbent assay. This approach gives a promising simple, sensitive and quantitative strategy for the detection of autophagy

β-Asarone Attenuates Aβ-Induced Neuronal Damage in PC12 Cells Overexpressing APPswe by Restoring Autophagic Flux

“…To evaluate the activity of autophagic flux and explore the mechanism of β-asarone’s protective effects, the expression levels of Beclin-1, p62, LC3-Ⅱ, and Aβ 1-42 were assessed. Beclin-1 (BECN1) initiates the formation of autophagosomes and induces the recruitment of other autophagy-related proteins to the autophagosome ( Wang et al, 2017 ). Therefore, Beclin-1 protein can serve as a marker for autophagy, and its expression levels can be used to assess whether autophagosome production is activated or inhibited.…”

Biomimetic gold nanomaterials for biosensing, bioimaging and biotherapy: a mini-review

“…Hybrid materials containing gold nanocages can usually achieve targeted drug delivery, PTT, fluorescence imaging and other purposes, which have a broad application prospect in the field of biomedicine. 77–79…”

Multiplexed Immunosensors and Immunoarrays

“…Antibodies were conjugated into the E-rGO surface for label free detection, with LODs from 0.1 mM to 10 mM for both analytes within response time less than a minute. Wang and Li et al, developed a gold-reduced graphene oxide nanocomposite (Au/r-GO) with gold nanocages serving as carriers for secondary antibodies and redox probes to detect autophagic biomarkers . The assay enabled detecting 2 autophagic biomarkers at clinically useful levels, Beclin-1 and LC3B–II, with LODs ∼0.03 ng/mL.…”

“…oxide nanocomposite (Au/r-GO) with gold nanocages serving as carriers for secondary antibodies and redox probes to detect autophagic biomarkers. 16 The assay enabled detecting 2 autophagic biomarkers at clinically useful levels, Beclin-1 and LC3B−II, with LODs ∼0.03 ng/mL. Graphene oxide has also been used for electrochemical protein detection in microfluidic immunoarrays.…”