ANTIBODIES BOUND TO NITROCELLULOSE IN ACIDIC BUFFERS RETAIN BIOLOGICAL-ACTIVITY

Citation
Wl. Hoffman et al., ANTIBODIES BOUND TO NITROCELLULOSE IN ACIDIC BUFFERS RETAIN BIOLOGICAL-ACTIVITY, Electrophoresis, 14(9), 1993, pp. 886-891
Citations number
20
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Biochemical Research Methods
Journal title
ISSN journal
0173-0835
Volume
14
Issue
9
Year of publication
1993
Pages
886 - 891
Database
ISI
SICI code
0173-0835(1993)14:9<886:ABTNIA>2.0.ZU;2-V
Abstract
This report compares the binding of proteins to nitrocellulose membran es in acidic buffers (pH 2 and 3) with binding in neutral buffer (pH 7 ). Initially, similar amounts of antibodies and other proteins bound t o the nitrocellulose membrane in both acidic and neutral buffers. Howe ver, the susceptibility of individual proteins to displacement (stripp ing) from the membrane by the milk blocking agent depended on the pH o f the buffer used to bind the proteins to the membrane. Most proteins that bound to nitrocellulose in acidic buffers were relatively resista nt to milk-stripping compared to proteins bound in pH 7 buffer. Acid-b inding of proteins to nitrocellulose also decreased the amount of prot ein that was stripped from the nitrocellulose membrane when Tween 20 w as included in the washing buffer. After correcting for the amount of antibody remaining on the membrane after the milk block, it was found that acid-bound antibodies were unchanged in biological activity when compared with the same antibodies bound at neutral pH. These results s uggest that acid-binding of proteins could increase the sensitivity of nitrocellulose membrane assays that use milk and/or Tween 20.