Establishment of a direct competitive ELISA for recombinant human EGF-IL-18 fusion protein
1.School of Laboratory Medicine and Life Science, Wenzhou Medical University, Wenzhou, 325035; 2.Shandong Hong Li Laboratory Animal Experiment Co., Ltd, Jinan, 250101
HONG Defang1,AI Dongdong1,Ding Linchao1, et al. Establishment of a direct competitive ELISA for recombinant human EGF-IL-18 fusion protein[J]. JOURNAL OF WEZHOU MEDICAL UNIVERSITY, 2014, 44(7): 485-.
Abstract：Objective: To establish a direct competitive ELISA for determining recombinant human EGF-IL-18 (rhEGF-IL-18) fusion protein levels in serum samples of macaca fascicularis and perform a validation for the assay which would be used in pharmacokinetics. Methods: The optimal concentrations of coating antibody and enzyme-labelled antigen were determined by chessboard titration crossing test. The direct competitive ELISA for determining rhEGF-IL-18 was established and the methodology performance was evaluated. Results: The optimal concentrations of coating antibody and enzyme-labelled antigen were 4 μg/mL and 1:1 000, respectively. The range of the standard curve was 100~900 ng/mL (R2=0.9898). The RSD of intra-day and inter-day were lower than 2.42% and 2.20%, respectively. The recovery was 95.3%~103.0%. The RSD of the samples standing at room temperature 1 h, freezing and thawing twice, cryopreserved for 20 days were less than 15%. The lower limit of quantification was 20.0 ng/mL. Conclusion: ELISA established in this study is stable and feasible.
