Epitope mapping of SAK by gene-targeted random fragment library / 中国免疫学杂志

ABSTRACT

Objective:To map dominant antigenic determinants on SAK by gene-targeted fragmemt library . Methods:①The PcAbs specificly against SAK were produced by immunning BALB/C mice. Pu rified the antisera through a SAK-Sepharose 4B affinity chromatography column. The purified PcAbs were biotinylated for next step. ② After constructing SAK r andom epitope library we sequenced 12 isolated clones randomly to ensure its int egrity ,capability and randomness.③The library was screening by situ-clone hy bridization.④Constructed mSAK ,the A1 region deleted mutant of SAK ,and used Western-blot assay to identified its immunoreactivity. Results:①Got a dominant epitope at amino acid 71-89,called A1 region; ②Western-bl ot assay suggested that mSAK, a mutant SAK without A1 region., didn't combine to the anti-SAK PcAbs.Conclusion:A dominant epitope of SAK was mapped successfully with a simple,effective method . [