靶向人CPNE7可变剪接异构体的鼠源抗血清制备与应用鉴定OA

Objective Aberrant expression of key alternative splice isoforms of calcium-dependent membrane-binding protein 7(CPNE7)has been observed in various cancers.To facilitate functional studies,this study aims to generate a mouse-derived antiserum capable of specifically recognizing both major splice variants,CPNE7-FL and CPNE7-S.Methods Conserved sequences shared among CPNE7 isoforms were identified by bioinformatic a-nalysis,and synthetic peptides were designed and used to immunize eight BALB/c mice to prepare antiserum.The antiserum titer was determined by indirect ELISA.Subsequently,a prokaryotic expression vector for CPNE7 was constructed and an A549 overexpression system was established,and the specificity of the antiserum was ver-ified by Western blot.Results A high-titer antiserum with a dilution of up to 1∶32 000 was successfully ob-tained.Western blot results showed that the antiserum effectively recognized recombinant GST-CPNE7 fusion proteins in E.coli and detected the overexpressed CPNE7-FL and CPNE7-S isoforms in mammalian cells.Conclusion A high-titer and high-specificity mouse antiserum against human CPNE7 was successfully prepared.This antiserum provides a valuable tool for future studies on the expression patterns and biological functions of CPNE7 isoforms,laying a foundation for exploring their potential roles in cancer diagnosis and therapy.