Development of a rapid and sensitivity magnetic chemiluminescence immunoassay for DNA methyltransferase 1 in human serum
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摘要
DNA methyltransferase 1(DNMT1) is a useful biomarker for lung cancer in early clinical diagnosis. A rapid magnetic chemiluminescence immunoassay(MCLIA) for DNMT1 in human serum has been developed.Horseradish peroxidase(HRP)-second-Ab was used to labeled polyclonal antibodies of anti-DNMT1. DNMT1 in sample integrates with specific immunomagnetic beads and can constitute a supersandwiched immunoreaction. In magnetic field, nonspecific materials can be separated. After luminescent substrate luminol-H_2O_2-BIP was added, the relative light unit(RLU) of HRP was detected and was discovered to be directly proportional to the content of DNMT1 in sample. The correlative variables involved in the MCLIA value were optimized and the methodological evaluation was carried out. After optimization, in the range of0.5–128 ng/mL, the linear regression equation was y = 0.5014 x + 1.769(x was logCDNMT1, y was relative luminescence units(RLU)/RLU0), and the limit of detection was 0.01 ng/mL. The RSD of intra-and interassays were 15.8%–16.9% and 14.3%–18.1%, respectively. The recovery was from 70.0% to 106.2%.Furthermore, paralleled with purchasable enzyme-linked immunosorbent assay(ELISA) kits, MCLEIA had lower detection limit, wider linear range and shorter detection time. Therefore, the MCLEIA established in this study could be used for the sensitive detection of DNMT1 in serum sample.
DNA methyltransferase 1(DNMT1) is a useful biomarker for lung cancer in early clinical diagnosis. A rapid magnetic chemiluminescence immunoassay(MCLIA) for DNMT1 in human serum has been developed.Horseradish peroxidase(HRP)-second-Ab was used to labeled polyclonal antibodies of anti-DNMT1. DNMT1 in sample integrates with specific immunomagnetic beads and can constitute a supersandwiched immunoreaction. In magnetic field, nonspecific materials can be separated. After luminescent substrate luminol-H_2O_2-BIP was added, the relative light unit(RLU) of HRP was detected and was discovered to be directly proportional to the content of DNMT1 in sample. The correlative variables involved in the MCLIA value were optimized and the methodological evaluation was carried out. After optimization, in the range of0.5–128 ng/mL, the linear regression equation was y = 0.5014 x + 1.769(x was logCDNMT1, y was relative luminescence units(RLU)/RLU0), and the limit of detection was 0.01 ng/mL. The RSD of intra-and interassays were 15.8%–16.9% and 14.3%–18.1%, respectively. The recovery was from 70.0% to 106.2%.Furthermore, paralleled with purchasable enzyme-linked immunosorbent assay(ELISA) kits, MCLEIA had lower detection limit, wider linear range and shorter detection time. Therefore, the MCLEIA established in this study could be used for the sensitive detection of DNMT1 in serum sample.
DNA methyltransferase 1(DNMT1) is a useful biomarker for lung cancer in early clinical diagnosis. A rapid magnetic chemiluminescence immunoassay(MCLIA) for DNMT1 in human serum has been developed.Horseradish peroxidase(HRP)-second-Ab was used to labeled polyclonal antibodies of anti-DNMT1. DNMT1 in sample integrates with specific immunomagnetic beads and can constitute a supersandwiched immunoreaction. In magnetic field, nonspecific materials can be separated. After luminescent substrate luminol-H_2O_2-BIP was added, the relative light unit(RLU) of HRP was detected and was discovered to be directly proportional to the content of DNMT1 in sample. The correlative variables involved in the MCLIA value were optimized and the methodological evaluation was carried out. After optimization, in the range of0.5–128 ng/mL, the linear regression equation was y = 0.5014 x + 1.769(x was logCDNMT1, y was relative luminescence units(RLU)/RLU0), and the limit of detection was 0.01 ng/mL. The RSD of intra-and interassays were 15.8%–16.9% and 14.3%–18.1%, respectively. The recovery was from 70.0% to 106.2%.Furthermore, paralleled with purchasable enzyme-linked immunosorbent assay(ELISA) kits, MCLEIA had lower detection limit, wider linear range and shorter detection time. Therefore, the MCLEIA established in this study could be used for the sensitive detection of DNMT1 in serum sample.
引文
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