Alexa fluor 647 affinipure goat anti human igm

The proteome microarrays containing full-length N, full-length E, and truncated S proteins of SARS-CoV-2 and 966 peptides representing SARS-CoV-2 proteins were prepared as described previously [17] . The proteome microarrays were assembled in an incubation tray and blocked with 5% (w/v) milk in PBS containing 0.05% (v/v) Tween-20 (PBST) for 10 min at room temperature before antibody detection. After aspirating the blocking solution, 1:101 diluted serum was added to the array and incubated at room temperature for 30 min. After washing three times with PBST, the array was then incubated for 20 min with a mixture containing Alexa Fluor ®647 Affinipure goat anti-human IgM(Jackson ImmunoResearch, USA, CAT#709–165-149) and Cy™3 Affinipure donkey anti-human IgG(H + L) antibody(Jackson ImmunoResearch, USA, CAT#109–605-043) (4 μg/mL) or only Cy™3 AffiniPure goat anti-Human serum IgA antibody(Jackson ImmunoResearch, USA, CAT#109–165-011) (4 μg/mL). Finally, the array was washed with PBST and deionized water, dissembled from the tray, and dried with a vacuum pump. The proteome microarray was scanned at 532 and 635 nm using a GenePix 4300A microarray scanner(Molecular Devices). The median of fluorescent signal intensity with the deduction of the background was extracted using GenePix Pro7 software.