Electrochemical immunosensor nanoarchitectonics with the Ag-rGO nanocomposites for the detection of receptor-binding domain of SARS-CoV-2 spike protein.

As the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) poses a grave threat to human life and health, it is essential to develop an efficient and sensitive detection method to identify infected individuals. This study described an electrode platform immunosensor to detect SARS-CoV-2-specific spike receptor-binding domain (RBD) protein based on a bare gold electrode modified with Ag-rGO nanocomposites and the biotin-streptavidin interaction system. The Ag-rGO nanocomposites was obtained by chemical synthesis and characterized by electrochemistry and scanning electron microscope (SEM). Cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were used to record the electrochemical signals in the electrode modification. The differential pulse voltammetry (DPV) results showed that the limit of detection (LOD) of the immunosensor was 7.2 fg mL-1 and the linear dynamic detection range was 0.015 ~ 158.5 pg mL-1. Furthermore, this sensitive immunosensor accurately detected RBD in artificial saliva with favorable stability, specificity, and reproducibility, indicating that it has the potential to be used as a practical method for the detection of SARS-CoV-2.

[The soluble expression and application of human papillomavirus type 58 (HPV58) major capsid protein L1].

Objective The major capsid protein L1 of human papillomavirus type 58 (HPV58 L1) was obtained and identified by prokaryotic expression. Methods The recombinant expression strain pE-SUMO-58 L1 (BL21) was induced by IPTG. The recombinant protein SUMO-58 L1 was expressed in E.coli and identified by SDS-PAGE and Western blot analysis. Then the recombinant protein SUMO-58 L1 was purified by Ni-column and the SUMO-tag was removed by ubiquitin-like protease 1 (ULP1) digestion. Subsequently, the bioactivity of recombinant protein HPV58 L1 was verified by hemagglutination assay (HA). BALB/c mice were immunized with HPV58 L1, and the antibody titers in sera of the immunized mice were detected by ELISA. And then the reaction between the immune serum and the HPV58 L1 protein transiently expressed by HEK293T cells was detected by indirect immunofluorescence assay (IFA). Results The soluble expression of the recombinant protein SUMO-58 L1 was identified by SDS-PAGE and Western blot analysis, with yields of soluble protein SUMO-58 L1 being about 50% of total soluble bacterial proteins. The relative molecular mass (Mr) of SUMO-58 L1 was about 72 000. After Ni-NTA affinity was purified and the SUMO-tag was removed by ULP1 digestion, Mr of recombinant protein HPV58 L1 reached about 58 000. The recombinant protein HPV58 L1 showed hemagglutination activity similar to that of natural HPV, with hemagglutination value of 1:16. After immunizing BALB/c mice, the titer of immune serum observed was about 1:10 240 by ELISA; and the sera of the immunized mice reacted specifically with HPV58 L1 proteins which were transiently expressed in HEK293T cells by IFA. Conclusion The recombinant protein HPV58 L1 also has hemagglutination activity, which can be successfully obtained from E. coli. The sera of the HPV58 L1 protein immunized mice can be used for immunocytochemical detection of HPV58 L1 protein expressed in eukaryotic cells.

An Electrochemical Immunosensor Based on SPA and rGO-PEI-Ag-Nf for the Detection of Arsanilic Acid.

A sensitive electrochemical immunosensor was prepared for rapid detection of ASA based on arsanilic acid (ASA) monoclonal antibody with high affinity. In the preparation of nanomaterials, polyethyleneimine (PEI) improved the stability of the solution and acted as a reducing agent to generate reduced graphene oxide (rGO) with relatively strong conductivity, thereby promoting the transfer of electrons. The dual conductivity of rGO and silver nanoparticles (AgNPs) improved the sensitivity of the sensor. The synthesis of nanomaterials were confirmed by UV-Vis spectroscopy, X-ray diffraction, transmission electron microscopy and scanning electron microscopy. In the optimal experiment conditions, the sensor could achieve the detection range of 0.50-500 ng mL-1 and the limit of detection (LOD) of 0.38 ng mL-1 (S/N = 3). Moreover, the sensor exhibited excellent specificity and acceptable stability, suggesting that the proposed sensor possessed a good potential in ASA detection. Thus, the as-prepared biosensor may be a potential way for detecting other antibiotics in meat and animal-derived foods.