| Ampicilline coupled onto macromolecular protein such as Bovine Serum Albumin(BSA),Human Serum Albumin(HAS), Hemocyanin succinylated from keyhole limpet (KLH),by glutaraldehyde reaction.After a reaction time of 4 h,the ampicilline-protein conjugate was dialysed against repeated changes of 1000 mL PBS (pH 6.0) using magnetic stirrer ,at 4℃ for 48h.The rotio of ampicilline coupled onto protein was determined by UV ,the rotio of ampicilline- BSA, ampicilline- HSA, ampicilline- KLH was 118,44,2192 respectively. The reslut of SDS-PAGE indicate the ampicilline-protein was prepared succeefully.l mL ampicilline-protein emulsified with 1 mL complete Freund's adjuvant(CFA) or incomplete Freund's adjuvant(IFA), the Balb/c mice with age of eight weeks was immuned with 250 M 1 of the mixture by injected into abdominal cavity, and after about one month the next immunity was done,the Balb/c mice was immuned 5-6 times. Splenocytes of vaccined were fused with mouse myelomas cells SP2/0 by 50% PEG 4000 ,and selected with HAT medium.Hybridoma clone secreting monoclonnal antibodies against ampicilline were screened by indirect ELISA,and positive clones were subcloned three times by limited dilution. Four strain was obtained and was named 2D5 4C5 respectively.The average number of chromosome the four hybridoma cell srains 2D5 4C5 was 95.6 96.9.The ELISA titers of monoclonal antibodies in cell supernatant was 1: 1024, 1: 512,and that of ascite was 1: 102400 1: 204800 respectively; the protein denisity of ascite was 5.16 mg/mL 7.23 mg/mL.The subtype of the four hybridoma cell srains was IgG2b IgG2arespectively.The blocking test demonstrated that it was only the ampicilline that can block the antisrun specifically ,and the penicilline can block in some dgree,but the cephalexin,Streptomycin, Norfloxacin cann't block the antisrun specifically.Strain 2D5 4C5 hybridoma cell can also stably secret McAb after twenty serial passages and freezingand revivfing three times.
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