Production of specific antibodies against human cholesteryl ester transfer protein using C-terminal active peptide obtained by fusional expression of cholesteryl ester transfer protein cDNA

Abstract

Partial (94 bp from 3' end) cDNA for cholesteryl ester transfer protein (CETP), obtained from a full-length cDNA clone isolated from a human heart Xgtll cDNA library, was subcloned into a plasmid, pGEX, for the production of glutathione-S-transferase (GST$/CETP fusion proteins in Escherichia coli. The fusion protein, containing the carboxylic terminus of the CETP (31 amino acids) responsible for substrate binding of CETP, was produced as a soluble form in a large quantity. The soluble GST/CETP protein was further purified by glutathione-Sepha-rose-4B affinity chromatography and used as an antigen for the production of the rabbit polyclonal antibody. The resulting antibody showed good titers, not 이lly against the GST/CETP fusion protein, but also against chemically synthesized CETP-specific peptides (16 amino acids) having the internal sequences of the C-terminal region of CETP, as determined by ELISA. The antiserum would be useful for overcoming the difficulty of CETP purification and as an immunological tool for CETP assay in future studies.