RESUMEN
Objective The value of pro-gastrin releasing peptide ( PGRP) which is the tumor marker of small cell lung canc-er has become a hot topic in recent years .The research was to build a new enzyme-linked immune sorbent assay ( ELISA) method ai-ming at detecting the concentration of PGRP in patients′serum. Methods We utilized synthetic PGRP epitopes for the screening of the monoclonal antibodies , labeled the screened monoclonal antibodies with horseradish peroxidase by modified sodium iodide method , and then established double antibody sandwich ELISA which could be used to detect the serum concentrations of PGRP in cancer pa -tients. Results We successfully screened E 12 mAb which could be served as the coating antibody and ED 1 mAb as the labeled anti-body.The standard antibody density range of new ELISA was 33 pg/mL~1.7 ×104 pg/mL.The comparison experiments between our method and the commercially available ELISA kit showed no significant difference ( P>0.05).The specificity of our method was 50%, and the sensitivity was 100%, while IBL kit was 92.2% and 100% respectively. Conclusion New ELISA can be used to detect the serum PGRP concentration in patients with small cell lung cancer .
RESUMEN
Objective To explore the effect of purification on monoclonal antibody (MAb) against PGRP by Protein A-Sepharose affinity chromatography, and to provide some based data for the purification of other antibody using the same method. Methods The ascites which include MAb was purified by Protein A-Sepharose affinity chromatography. The purity and activity of MAb was tested by SDS-PAGE and ELISA. The biological function was identified by flow cytometer and immunohistochemistry. Results The average concentration of protein in ascites before purification is 23.62 mg/ml. Before and after purification, the total protein is 148.79 mg and 146.67 mg, respectively. The recovery coefficient of protein is 98.58%. The concentration of MAb in ascites is 5.21 mg/ml averagely. The MAb purity is more than 95 %. The immunoactivity of purified antibody is higher than that of unpurified antibody. Conclusion The purity of MAb against PGRP purified by Protein A-Sepharose affinity chromatography is very high. The immunoactivity of purified antibody is higher than that of unpurified antibody. So the ProteinA-Sepharose affinity chromatography is a rapid, convenient and reliable method for the purification of MAb Against PGRP.