Isolation of antibodies against different protein conformations using immunoaffinity chromatography

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• 作者：Mikhail L. Kuravsky^a ; Elena V. Schmalhausen^b ; Natalia V. Pozdnyakova^b ; Vladimir I. Muronetz^a ; ^b ; ^{vimuronets@belozersky.msu.ru}
• 关键词：Polyclonal antibodies ; Immunoaffinity chromatography ; Conformational epitopes ; GAPDS
• 刊名：Analytical Biochemistry
• 出版年：2012
• 出版时间：1 July, 2012
• 年：2012
• 卷：426
• 期：1
• 页码：47-53
• 全文大小：308 K

文摘

A polyclonal antiserum obtained after the immunization of a rabbit with recombinant human sperm-specific glyceraldehyde-3-phosphate dehydrogenase lacking in 68 N-terminal amino acid residues (dN-GAPDS) was purified using different immunosorbents with immobilized dN-GAPDS in the native or denatured states. The procedure resulted in isolation of two types of polyclonal antibodies. The first type interacted with native recombinant dN-GAPDS as well as with native human sperm-specific glyceraldehyde-3-phosphate dehydrogenase, not cross-reacting with muscle glyceraldehyde-3-phosphate dehydrogenase (GAPD). The second type interacted with both native and denatured forms of the sperm-specific proteins, exhibiting some cross-reaction with GAPD. Thus, the suggested approach allows isolation of the antibodies against conformational or linear epitopes from the same polyclonal serum.