Time-resolved fluoroimmunoassay of pepsinogen I
Objective To establish a two-site time-resolved fluoroimmunoassay (TRFIA) of pepsinogen Ⅰ (PGⅠ) based on the direct sandwich technique. PGⅠ-TRFIA was used to detect serum PGⅠ concentration. Methods In the equilibrium procedure, the monoclonal antibodies (McAb) against a specific antigenic site on the PGⅠ was 8016#, the europium-labelled McAb was prepared by europium-chelate of N-(p-isothiocyanatobenzyl)- diethylenetriamine-N, N, N, N-tetraacetic acid directed against a different antigenic site on the PGⅠ molecule was 8003#. The luminescent enhancement system was enhancement solution containing 2-naphthoyltrifluoroacetone. PGⅠ in sera from patients and healthy controls were examined by PGⅠ-TRFIA with auto DELFIA1235 system. Results The intra- and inter-assay CV of the PGⅠ-TRFIA were 1.9% and 4.7%, respectively, and the recovery rate was 102.65%, the sensitivity was 0.05μg/L. The McAb provided a linear response from 3.5 to 328μg/L with ED 50 of 38.73μg/L. The cross-reacting rate with pepsinogen Ⅱ was negligible. The europium-labelled 8003# was stable for at lest eight months at -20℃ and the results of the TRFIA with same reagents were stable for eight months. Conclusion The newly developed PGⅠ-TRFIA was high sensitivity and very stable. The method can be used in serum immunoassay such as gastric cancer detection. The detection of PGⅠ in sera will show patient an early warning of developing gastric cancer in future.