The Preparation Of Polyclonal Antibodies And The Detection Of Some Pollutants Of Human Internal Exposure

With the development of technology and industry,people’s life is becoming more and more convenient,but also,the problem of environmental pollution has been brought about.In recent years,the pollutants such as salbutamol、diethylstilbestrol polychlorinated biphenyl、bisphenol A、tartrazine,have been widely used,that seriously threatens our lives.This issue has caused wide public concern.The detection of pollutants internal exposure can objectively reflect the exposure levels of individuals to pollutants,also can avoid the influence of individual differences.lt is one of the effective methods to prevent the harm of environmental pollution.However,the detection technology of the internal exposure of some pollutants still have some technological bottlenecks,such as the types of the pollutants that should be detected is numerous,the pretreatment of samples is difficult,the quality of pollutants in the biological samples is low,the cost of detection is difficult to control,so that the detection project of pollutants internal exposure is difficult to extent.Therefore,the detection technology of pollutants of internal exposure should be studied.The objective of this study is to establish a rapid、easy、sensitive and special assay method to detect some pollutants of internal exposure,so that the method is suitable to large-scale detecting human samples.According to the references,firstly,the artificial antigen and the polyclonal antibodies of some pollutants (salbutamol、diethylstilbestrol) have been prepared,then a assay method that is rapid、easy、sensitive、special and suitable to large-scale detecting human samples have been developed.The study have laid a concern foundation to overcome the bottlenecks of the technology to detect the pollutants of human internal exposure.1.The preparation of diethylstilbestrol polyclonal antibodies and the development of the detection technology of human internal exposure to detect diethylstilbestrolObjective:To prepare the DES polyclonal antibodies of high titer and sensitivity,further to develop and optimize the detection technology of human internal exposure to diethylstilbestrol.Methods:The nucleophilic substitution reaction and EDC/sulfo-NHS were utilized to prepare the artificial antigen,then the rabbits were immunized with the DES artificial antigen to prepare the DES polyclonal antibodies. The ELISA assay method has been developed and optimized,the ELISA method has also been studied.Then the ELISA assay method have been used to large-scale detect the DES in human urine samples. Results:The DES polyclonal antibodies which have the IC50of7.67ng/ml had been obtained.The indirect ELISA assay method to detect the DES of human urine samples has been developed. And the limit of detection for DES of this method is0.634ng/ml,the recovery of this method is85%～110%,the CV<10%.100normal human urine samples have been detected by this method.Conclusions:The method established by this study has almost the same sensitivity as those methods that have been reported and can be used to monitor and measure the harm that the DES has to human.2.The preparation of salbutamol polyclonal antibodies and the development of the detection technology of human internal exposure to detect salbutamol and clenbuterol simultaneouslyObjective:To prepare the SAL polyclonal antibodies of high titer and sensitivity,further to develop and optimize the simultaneous detection technology of human internal exposure to salbutamol and clenbuterol.Methods:The rabbits were immunized with the salbutamol artificial antigen to prepare the salbutamol polyclonal antibodies.The ELISA assay method has been developed and optimized,the ELISA have also been determined.Then the ELISA assay method has been used to large-scale detect the salbutamol and clenbuterol in human urine samples.Results:The salbutamol polyclonal antibodies which have the IC50of2.208ng/ml had been obtained.The indirect ELISA assay method to detect the salbutamol of human urine samples has been developed. And the limit of detection of this method is0.152ng/ml,the recovery of this method is85%～110%,the CF<10%,as well as the salbutamol polyclonal antibodies have the cross-reactivity ration of98.12%,the method that have been established can also be used to detect salbutamol and clenbuterol simultaneously.100normal human urine samples have been detected by this method.Conclusions:The method established by this study have almost the same sensitivity as those methods that have been reported,can be used to monitor and measure the harm that the salbutamol and clenbuterol simultaneously has to human.To sum up:In my work,the human internal exposure detection technology of some pollutants such as salbutamol and diethylstilbestrol has been explored.The artificial antigens of some pollutants have been synthesized,and the polyclonal antibodies of some pollutants have been prepared.Basing on this,the ELISA method to measure the pollutants in human have been established.Also,the normal human urine have been used as the analyzed samples.Then a large number of samples have been detected by the methods.The high positive rate of pollutants in human urine samples not only indicate the necessity of studying the detection technology of internal exposure to pollutants,but also,have proved that the detection methods is scientific.Because the human urine samples has no harm to human being,in the case of not reducing accuracy,the methods that have been established can reduce the workload effectively,also can be rapid、high-throughput、convenient and low-cost,so it have laid a concern foundation for its popularization.