Development of a sensitive micro-magnetic chemiluminescence enzyme immunoassay for the determination of carcinoembryonic antigen |
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Authors: | Wijitar Dungchai Weena Siangproh Jin-Ming Lin Orawon Chailapakul Si Lin Xitang Ying |
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Affiliation: | (1) The Key Laboratory of Bioorganic Phosphorus Chemistry & Chemical Biology, Ministry of Education, Department of Chemistry, Tsinghua University, Beijing, 100084, China;(2) Department of Chemistry, Faculty of Science, Chulalongkorn University, Patumwan, Bangkok, 10330, Thailand;(3) Beijing Chemclin Biotech Co., Ltd., Beijing Academy of Science and Technology, Haidian, Beijing, 100094, China |
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Abstract: | A micro-magnetic chemiluminescence (CL) enzyme immunoassay with high sensitivity, selectivity, and reproducibility was developed for the determination of the tumor marker, carcinoembryonic antigen (CEA) in human serum. A sandwich scheme assay has been utilized with fluorescein isothiocyanate antibody (FITC)-labeled anti-CEA antibody and alkaline phosphate (ALP)-labeled anti-CEA antibody being used in the CL detection. The CL signal produced by the emission of photons from 4-methoxy-4-(3-phosphate-phenyl)-spiro-(1,2-dioxetane-3,2′-adamantane) (AMPPD) was directly proportional to the amount of analyte present in a sample solution. The influences of the reaction time of antigen with antibody, the reaction time of substrate with label, the dilution ratio of ALP-labeled anti-CEA antibody, the concentration of FITC-labeled anti-CEA antibody, and other relevant variables upon the CL signal were examined and optimized. The CL responses depended linearly on the CEA concentration over the range from 2 to 162 ng mL−1 in a logarithmic plot. Assay sensitivity as low as 0.69 ng mL−1 was achieved. A coefficient of variance of less than 13% was obtained for intra- and inter-assay precision. This method has been successfully applied to the analysis of CEA in human serum. According to the procedure based on spiked standards, the recoveries obtained were 80–110%. Comparison experiments were carried out with the commercially available CEA chemiluminescence immunoassay. Satisfactory results were obtained according to a paired t-test method (t value < t critical at the 95% confidence level). |
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Keywords: | Micro-magnetic beads Carcinoembryonic antigen Tumor Chemiluminescence enzyme immunoassay Marker |
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