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. Author manuscript; available in PMC: 2010 Dec 1.
Published in final edited form as: J Reprod Immunol. 2009 Oct 23;83(1-2):145–150. doi: 10.1016/j.jri.2009.06.260

Figure 1.

Figure 1

Peripheral blood lymphocytes (PBL) were collected from immunoinfertile men and vasectomized men and were activated for seven days in vitro with human sperm extract (HSE), purified human fertilization antigen-1 (FA-1), and synthetic YLP12 peptide. RNA was isolated and reverse transcribed to cDNA by PCR using various VL and VH chain primers as described in detail elsewhere (Samuel and Naz, 2008). The scFv assembly of VH and VL chains, with a (G4S)3 linker between the chains, was performed using the splicing by overlapping extension PCR (SOE-PCR) procedure. The final assembled PCR product was ligated into pCANTAB5E vector to make the scFv phage library. The library was panned against various antigens (HSE/FA-1/YLP12) and the isolated clones were sequenced and analyzed for antibody activity using enzyme-linked immunosorbent assay (ELISA), Western blot procedure, immunoprecipitation procedure, indirect immunofluoroescence technique (IFT), and sperm acrosome reaction.