| Malignant tumor is one disease with high fatality rate. About 8.2 million people worldwide died from cancer in 2012 according to World Cancer Report 2014 published by WHO and International Agency for Research on Cancer(IARC). Detecting tumor markers at early age facilitates diagnosis of cancer and treatment monitoring and significantly improves the treatment efficiency and survival rates. There are lots of methods to detect tumor markers, of which the immunochromatography technology has gained more and more attention. Immunochromatography technology is developing rapidly because it has advantages of minimal manipulation, low cost, small analyte volume, no need of professional as well as the capability of realizing point-of-care testing. However, the most used colloidal gold-based test strip, which mainly realizes qualitative detection, requires two or more test lines when working with multianalyte and its low sensitivity cannot meet the demand for early simultaneous quantitative detection of tumor markers.The material used for signal generation plays a decisive role in the immunosensor, which determines the performance of the sensor. Fluorescent QDs is chosen as fluorescent probe with the advantages of high level of brightness, and exceptional photostability, the emission spectra of QDs are narrow and symmetrical while the absorption spectra are broad and continuous, which allow exciting all colors of QDs simultaneously with only a single excitation light source without substantial emission spectral overlap. Considering the current shortcomings of the immunochromatography product and the excellent characteristics of QDs, we adopt multicolor QDs as fluorescent material to develop a multicolor QDs based immunosensor to realize rapid, simultaneous and quantitative detection of tumor markers.EDC and NHS are used as cross linkers to prepare conjugation of QDs546-anti-AFP McAb and QDs620-anti-CEA McAb. In order to fabricate the multicolor QDs based immunosensor, the two prepared conjugation are mixed at a desired ratio and are applied onto the conjugate pad, the mixture of mouse anti-AFP McAb and mouse anti-CEA McAb are immobilized on NC membrane as test line, and the goat anti-mouse IgG is immobilized as control line. Fluorescence intensity of test line and control line on NC membrane are measured by a fluorescence reader. Simultaneous quantitative detection of AFP and CEA is achieved according to standard curves which are plotted based on the relationship between the T/C ratio and different AFP and CEA concentrations. Under the optimum conditions, AFP and CEA could be detected at a limitation as low as 2ng/ml and 1 ng/ml and no obvious cross-reactivity is observed. The multicolor QDs based immunosensor is validated with clinical samples and it demonstrates high sensitivity, high specificity and high recoveries. The test results exhibites good stability within batches and small difference between the batches.The immunosensor that combined multicolor QDs with immunochromatography assay could achieve simultaneous quantitative detection of multiple tumor markers with only one test line and one control line on the NC membrane. This immunosensor is easy to use and there is no need of professional operation, which facilitates the early screening of tumor in communities, families, and is helpful for detecting instrument miniaturization and intellectualization in the future. |
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