| Accurate determination of α-fetoprotein(AFP) in human serum is very important for clinical diagnose and treatment of cancer. Inductively coupled plasma mass spectrometry(ICP-MS) has becoming more and more important in the detection of tumor-ralated proteins in recent years because of it’s outstanding properties, including excellent sensitivity, good detection limits for most elements, multielement analysis and isotope analysis capability. Methods based on rare earth element tag feature a high sensitivity and low determination limit for determination of proteins, in this work, two high sensitivity methods for determination of proteins based on ICP-MS have been discussed, the experiment conditions have also been optimized and the optimization conditions were used for determination of standard proteins and discussing the possibility of this method for determination of low abundant proteins in human serum according to the determination limits. The two methods are as follows:1、Proteins were labeled directly with rare earth element which has low blankground signal and high intensitivity by the double function chelating reagents such as DTPAA and DOTA, then separated by one-dimensional (1D HPLC) or nano two-dimensional high performance liquid chromatography(nano 2D HPLC) and polyacrylamide gel electrophoresis (PAGE), followed by quantitative analysis by ICP-MS or LA-ICP-MS.2、This method have made full used of the specificity of immunoreaction. First of all, the rare earth element was labeled on the antibody by the double function chelating reagents, and then the target proteins were labeled by immunoreaction with the labeled antibody, after that the dissociation solution was uesed for direct injection of solution or quantitative analysis directly by LA-ICP-MS. At the same time, different dissociation solutions such as acetic acid、alcohol and EDTA were choosed to disscuss the enhancement mechanism. Finally, the method has been successfully used for determination of AFP in human serum, and the detection limit has been reduced (0.57μ g·L-1). What’s more, the relationship between the target proteins and the labels in the immunocomplex was been established, and the stability of the relationship has been discussed. The possibility of the relationship used for isotope dilution analysis was proposed, an idea which can reduce the cost of TRFIA-AFP diagnostic kit and simplify the experiment process was presented.Sensitive quantitative methods for detection low abundant proteins based on ICP-MS by rare erath element label proteins have been researched, we found that compared with the methods which based on natrually elements exist in proteins, method based on metal labels has obivious advantages of low detection limit, but it still can not used to determination of low abundant proteins in complex samples because of the process has no specificity and the high separation requireements. However, we finally give the target proteins special labels by making full use of the specificity of immunoreaction, and then dissociated by 5%HNO3 which can improve the signal intensity to a large extent, and introdced to ICP-MS to quantitative analysis. The method has been successfully used to determination of AFP in human serum, the measurable range is 1～600μg·L-1, the relative standard deviation (RSD)of the results is below 10% and the detection limit is 0.57μg·L-1, which is low to the method based on ICP-MS reported in the literature before. The same serum samples have also been measured by TRFIA, the results of the two method were in good agreement, and the accuracy of ICP-MS based method is more excellent. |
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