Analysis of immunogenicity ofΔ42PD1 via yeast surface displaying peptide frag-ments / 中国免疫学杂志

ABSTRACT

Objective:To analyze the immunogenicity of the extracellular region of Δ42PD1.Methods: Six fragments ofΔ42PD1 extracellular region-encoding sequence were amplified by PCR, and were cloned into pCTCON2 vector, a yeast surface displaying vector.Yeast cells were transfected with Δ42PD1 fragment-carrying plasmids, then yeast cells were spread on SDCAA plates.Single cell clones were selected and cultured in SGCAA media to induce expression of the target genes.Mouse anti-humanΔ42PD1 anti-serum were generated by immunization of BALB/c mice via intramuscular injection ofΔ42PD1-carrying plasmid plus in-situ electroporation.The binding of anti-serum with yeast cells surface-displaying Δ42PD1 fragments were analyzed using flowcytometry.Results:Nucleotide sequences analysis indicated that the amplified six fragments ofΔ42PD1 sequence length were 110 bp,and the isolated sequence ofΔ42PD1 fragments were 100%homology with PD1 gene previously registered in GenBank.Results from flowcytometry showed that among the six fragments of Δ42PD1 displaying on the surface of yeast cells,F3 and F2 profoundly boundΔ42PD1-specific polyclonal antibodies.Conclusion:F3 and F2 ofΔ42PD1 is an immunogenic dominant region,which pave the way for generation of Δ42PD1-specific monoclonal antibody and epitope mapping.