Western Blot Analysis of Recombinant Proteins

The integrity of the purified recombinant proteins was confirmed by Western blot analysis. Western blotting was performed according to the standard protocol [14 (link)] using sera from rabbit immunized against Vibrio cholerae and negative sera (as the negative controls) as the primary antibody at 1 : 100 dilution and HRP-conjugated (horseradish peroxidase) goat anti-rabbit IgG (Abcam, United Kingdom) at 1 : 2500 dilution in 1x TBST buffer (10x: 15 mMNaCl, 10 mMTris-HCl (pH = 7.4), 0.1% Tween 20) as secondary antibody. The reactions were developed by diaminobenzidine (DAB) solution (Roche, Germany).

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Molaee N., Mosayebi G., Amozande-Nobaveh A., Soleyman M.R, & Abtahi H. (2017). Evolution of the Immune Response against Recombinant Proteins (TcpA, TcpB, and FlaA) as a Candidate Subunit Cholera Vaccine. Journal of Immunology Research, 2017, 2412747.

Publication 2017

diaminobenzidine (DAB) solution

Anti igg Antibody Buffer Dilution Goat Horseradish peroxidase Rabbit Recombinant proteins Sera Tween 20 Vibrio cholerae Western blot analysis

Corresponding Organization : Arak University of Medical Sciences

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Variable analysis

independent variables

Sera from rabbit immunized against Vibrio cholerae

dependent variables

Integrity of the purified recombinant proteins

control variables

Negative sera (as the negative controls)

controls

Sera from rabbit immunized against Vibrio cholerae
Negative sera

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