Author: Chen, Yixin; Chan, Kwok-Hung; Kang, Yahong; Chen, Honglin; Luk, Hayes KH; Poon, Rosana WS; Chan, Jasper FW; Yuen, Kwok-Yung; Xia, Ningshao; Lau, Susanna KP; Woo, Patrick CY
Title: A sensitive and specific antigen detection assay for Middle East respiratory syndrome coronavirus Document date: 2015_4_22
ID: puv7kg4k_8
Snippet: Western blot analysis was performed according to our published protocol. 20 Briefly, 1.5 mg purified (His) 6 -tagged rNPs of MERS-CoV, human CoV (HCoV) 229E and HCoV-OC43 respectively were subjected to 12% SDS-PAGE and were transferred to 0.22 mm nitrocellulose membranes (GE Healthcare, Little Chalfont, Buckinghamshire, UK). The membranes were blocked with 5% skim milk in Trisbuffered saline and then incubated with mouse anti-MERS-CoV-rNP immune .....
Document: Western blot analysis was performed according to our published protocol. 20 Briefly, 1.5 mg purified (His) 6 -tagged rNPs of MERS-CoV, human CoV (HCoV) 229E and HCoV-OC43 respectively were subjected to 12% SDS-PAGE and were transferred to 0.22 mm nitrocellulose membranes (GE Healthcare, Little Chalfont, Buckinghamshire, UK). The membranes were blocked with 5% skim milk in Trisbuffered saline and then incubated with mouse anti-MERS-CoV-rNP immune sera at 37 6 C for 60 min. Antigen-antibody interaction was detected with 1:4000 diluted horseradish peroxidase (HRP)conjugated goat anti-mouse IgG (Novex, Life Technologies, Grand Island, NY, USA) and visualized by 3,39,5,59-tetramethylbenzidine (TMB) liquid substrates system for membranes (Sigma Aldrich, Saint Louis, MO, USA).
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