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[Preparation, epitope analysis and clinical application of monoclonal antibodies against heart-type fatty acid binding protein].

[Article in Chinese]

Author information

  • 1Department of Clinical Biochemistry, College of Medical Laboratory, Third Military Medical University, Chongqing 400038, China. hanqi688@163.com

Abstract

OBJECTIVE:

To prepare and characterize the monoclonal antibody (mAb) against recombinant human heart-type fatty acid binding protein (H-FABP) and apply it to the clinical analysis.

METHODS:

BALB/c mice were immunized with recombinant H-FABP (rH-FABP) from prokaryotic expression or synthesized peptide fragment. After common fusion and screening, the subtypes, titer and affinity of mAbs were detected respectively. After purification, the specificity of mAbs was tested by indirect ELISA and Western blotting. ELISA system was established by pair mapping and applied in the detection of clinical samples. Two epitope peptides were designed by bioinformatics and used to detect the epitope of 1-F10 through Western blotting.

RESULTS:

Four different hybridoma clones secreting anti-H-FABP antibodies were developed, with high titres of 1:51 200-1:1024 000. Immunoglobulin types of these mAbs were found to be IgG2a or IgG2b, respectively. The affinity of the mAb 1-F10 even reached 9.02×10(9); mol/L. ELISA and Western blotting showed that these mAbs could identify H-FABP specifically. In addition, the pair mapping of monoclonal antibodies 3-H5-1-F10 could recognize H-FABP in human serum samples. Furthermore, the specific target recognized by 1-F10 mAb was located within amino acid 86-133 of H-FABP.

CONCLUSION:

Four highly specific mAbs against H-FABP were successfully obtained and the established ELISA system could be used to detect the H-FABP in clinical serum samples, and the epitope of 1-F10 mAb was also verified.

PMID:
23746246
[PubMed - indexed for MEDLINE]
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