Author: Escandon, Paulina; Heatley, J Jill; Berghman, Luc R; Tizard, Ian; Musser, Jeffrey MB
Title: Comparison Of Four Anti-Avian IgY Secondary Antibodies Used In Western Blot And Dot-Blot ELISA To Detect Avian Bornavirus Antibodies In Four Different Bird Species Document date: 2019_11_12
ID: 2sr6ds4r_11
Snippet: Dot-blot ELISAs were performed according to Guo et al (2014) , 27 with the following modifications. Briefly, 2.0 µl of a 0.15 mg/mL recombinant N-protein solution was dotted onto a nitrocellulose (NC) membrane strip and incubated in 5% skim dried milk/0.05% Tween/0.02M PBS overnight at room temperature. The membrane strip was incubated for 5 mins with plasma diluted 1:60 with 3 mL 2% dried skim milk/0.02M PBS/0.05% Tween solution, followed by th.....
Document: Dot-blot ELISAs were performed according to Guo et al (2014) , 27 with the following modifications. Briefly, 2.0 µl of a 0.15 mg/mL recombinant N-protein solution was dotted onto a nitrocellulose (NC) membrane strip and incubated in 5% skim dried milk/0.05% Tween/0.02M PBS overnight at room temperature. The membrane strip was incubated for 5 mins with plasma diluted 1:60 with 3 mL 2% dried skim milk/0.02M PBS/0.05% Tween solution, followed by three 1 min rinses with 3 mL of PBS/ 0.05%Tween. The membrane was incubated for five minutes in 3 mL of one of the diluted secondary antibodies, followed by three 1 min rinses with 3 mL of PBS/0.05% Tween. The membrane was then incubated for 5 mins in a solution containing SigmaFast BCIP/NBT substrate tablet dissolved in 10 mL ultra-pure water. Finally, the membrane was rinsed with ultra-pure water.
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