Hrp conjugated goat anti mouse igg2a

To determine the titer and subtype of specific antibodies against NiV in serum, we developed an indirect enzyme-linked immunosorbent assay (iELISA) [25 (link)]. In brief, 0.5 μg/mL of F or G proteins of NiV were coated onto ELISA plates overnight at 4 °C. After three washes with PBST, the plate was blocked with 3% bovine serum albumin (BSA) for 2 h. After three washes with PBST, 50 μL/well of the diluted inactivated serum samples were added and incubated for 1 h at 37 °C. After washing, a 1:10,000 dilution of HRP-conjugated goat anti-mouse IgG (CWBIO, Taizhou China), a 1:5000 dilution of HRP-conjugated goat anti-mouse IgG1 (Southern Biotech, Birmingham, AL, USA), or a 1:5000 dilution of HRP-conjugated goat anti-mouse IgG2a (Southern Biotech, Birmingham, AL, USA) was added and incubated at 37 °C for 1 h. After TMB color development, the OD_450nm value of each well was read. A test sample/negative sample with an OD_450nm greater than 2 was considered positive.

H₂O₂ and CT were obtained from Sigma-Aldrich (St. Louis, MI, USA). Anti-mouse CD3-APC, CD4-FITC, CD8-PE, CD11c-eFluor 450, MHCII-APC, CD5-FITC and B220-PerCP-eFluor™ 710 antibodies were bought from eBioscience (San Diego, CA, USA). Horseradish Peroxidase (HRP)-conjugated goat anti-mouse IgA, HRP-conjugated goat anti-mouse IgG, HRP-conjugated goat anti-mouse IgG1 and HRP-conjugated goat anti-mouse IgG2a were obtained from Southern Biotech (Birmingham, AL, USA). RNA-easy Isolation was purchased from Vazyme Biotech Co., Ltd. (Nanjing, Jiangsu Province, China). SYBR qPCR Master Mix for quantitative real-time PCR (qRT-PCR) was bought from Yeasen Biotechnology Co., Ltd. (Shanghai, China). TMB was purchased from Beyotime Biotechnology (Shanghai, China). Red blood cell lysis buffer was purchased from Solarbio Science & Technology Co., Ltd. (Beijing, China). Silane coupling agent (KH550) and ZnONPs (purity 99.9%) were the products of Sinopharm Chemical Reagent Ltd. (Shanghai, China). The purified AMP was prepared in our laboratory (Yangzhou, Jiangsu Province, China). H₂SO₄, alcohol and HCl were obtained from Hushi Laboratory Equipment Co., Ltd. (Shanghai, China). Elisa plate was bought from cellpro biotechnology Co., Ltd. (Suzhou, Jiangsu Province, China).