Preparation of Monoclonal Antibodies to Trimethoprim and ELISA Kit for Rapid Detection

[Objective] This study was conducted to find out an approach for determining trimethoprim residues in water. [Method] Trimethoprim antigen was prepared through a series of reactions from trimethoprim hapten which was generated through the reaction between trimethoprim and maleic anhydride. And trimethoprim monoclonal antibodies were prepared by animal immune, and used to prepare ELISA kit to detect trimethoprim residues in water. Finally, the limit of detection （LED） of the ELISA kit was determined. [Result] The standard curve covered a concentration range of 0-80 μg/L. The LeD of trimethoprim in water using the ELISA kit was 2.34 μg/kg; the IC50 （half maximal inhibitory concentration） was 4.8 μg/L; the recovery rate of added trimethoprim standard ranged from 60.5% to 79.7%; within-and among-batches RSD was less than 10%. The trimethoprim monoclonal antibody was specific, as the cross-reactivity rate of trimethoprim antibody and diaveridine was less than 1%. The stability tests revealed that the ELISA kit was stable after being stored at 4 ℃ for 12 months. [Conclusion] The results will provide references for controlling the abuse of trimethoprim.

Development of an Indirect ELISA Using Recombinant Truncated Envelope Glycoprotein for Detection of Antibodies against Japanese Encephalitis Virus

[ Objective] To develop an indirect ELISA assay for detecting antibodies against envelope glycoprotein （ E protein） of Japanese encephalitis virus （JEV）. [ Method] Specific primers were designed according to JEV sequences published in the GenBank. The cDNA of JEV E gene （about 1 000 10p） was amplified by the RT-PCR with the specific primers. After sequencing analysis, the E gene was cloned into pET30a expression vector and expressed in E. coli BL21 （DE3） with the induction of IPTG. After denaturation, purification and renaturation, the recombinant protein was analyzed by the SDS-PAGE and the westem blotting. An indirect ELISA was developed to detect antibodies against JEV. [ Result] The E protein was mainly expressed in inclusion body. With the purified E protein, the indirect ELISA was developed and displayed good specificity, sensitivity and repeatability, [ Conclusion]The developed ELISA using the truncated E protein as antigen is a simple, convenient and rapid serological method for diagnosis, monitoring antibody level and epidemiological investigation of JEV.

Detection of Antibodies against Toxoplasma gondii by ELISA with Recombinant Microneme Protein 3

[Objective] To develop a new method for serodiagnosis of swine toxoplasmosis. [Method] With the purified recombinant microneme protein 3 （rMIC3） as coating antigens, an indirect ELISA was developed for detection of antibodies against Toxoplasma gondii. [ Result] The optimal working concentration of rMIC3 was 3. 40 ug/ml, and the optimal degree of dilution of sera was 1：160. Cross-reaction was not observed between the Toxoplasma gondii-positive sera and the positive sera against classical swine fever virus or some other pathogens. The developed ELISA had 92.56% coincidence rate with latex agglutination test. [ Conclusion] The developed ELISA is sensitive, rapid, specific and reproducible, and thus it can be applied in serodiagnosis and seroprevalence investigation of swine toxoplasmosis.

A MONOCLONAL ANTIBODY RECOGNIZING NON DERIVATIVE 13 HYDROXY GIBBERELLINS AND THEIR GLUCOSIDES *

The production and characterization of a monoclonal antibody (MAb AB10) against GA 3 glucoside as well as GA 3 is described. MAb AB10 was derived from an immunogen in which human serum albumin (HSA) was linked to GA 3 at carbon 3. This antibody showed high affinity for GA 3 glucoside as well as for 13 hydroxy gibberellins (GA 1, GA 3, GA 5, etc). The affinity of MAb AB10 for 13 hydroxy GAs was significantly reduced by methylation of the 7 oic acid but not by glycosylation of 3 hydroxyl group. Based on this antibody, both of competitive enzyme linked immunosorbent assays (ELISAs) for GA 3 glucoside and for GA 3 were developed. These two ELISAs displayed linear detection ranges from 0 2 pmol to 20 pmol. Using these assays, the fluctuation of GA 3 like and GA 3 glucoside like substances in the leaves of Rumex japonicus was investigated. The results indicated that the glycosylation of free GAs was connected with leaf senescence and that the function of 6 benzyl amino purine in retarding the leaf senescence was probably related to delaying the process of glycosylation of free GAs.

兰州地区孕产妇及新生儿中巨细胞病毒感染的研究

为调查兰州地区妊娠妇女中CMV感染状况及CMV母婴传播问题,利用ELISA方法对216例早孕妇女、238例产妇及其中的215例新生儿血清中的CMV特异性抗体IgM、IgG进行检测。结果:产妇静脉血清中CMV抗体阳性率IgM为3.8％,IgG为96％;新生儿脐带血中CMVIgM阳性率为1.9％;早孕妇女血清中CMV抗体阳性率IgM为4.2％,IgG94％。提示:兰州地区妊娠妇女中CMV感染相当普遍,其CMV活动性感染状况与全国其它地区感染水平基本一致。

Potential of soluble CD26 as a serum marker for colorectal cancer detection

Colorectal cancer is characterized by a low survival rate even though the basis for colon cancer development,which involves the evolution of adenomas to carcinoma,is known.Moreover,the mortality rates continue to rise in economically transitioning countries although there is the opportunity to intervene in the natural history of the adenoma–cancer sequence through risk factors,screening,and treatment.Screening in particular accounted for most of the decline in colorectal cancer mortality achieved in the USA during the period 1975-2000.Patients show a better prognosis when the neoplasm is diagnosed early.Among the variety of screening strategies,the methods range from invasive and costly procedures such as colonoscopy to more low-cost and non-invasive tests such as the fecal occult blood test(guaiac and immunochemical).As a non-invasive biological serum marker would be of great benefit because of the performance of the test,several biomarkers,including cytologic assays,DNA and mRNA,and soluble proteins,have been studied.We found that the soluble CD26(sCD26)concentration is diminished in serum of colorectal cancer patients compared to healthy donors,suggesting the potential utility of a sCD26 immunochemical detection test for early diagnosis.sCD26 originates from plasma membrane CD26 lacking its transmembrane and cytoplasmic domains.Some 90%–95%of sCD26 has been associated with serum dipeptidyl peptidase IV(DPPIV)activity.DPP-IV,assigned to the CD26 cluster,is a pleiotropic enzyme expressed mainly on epithelial cells and lymphocytes.Our studies intended to validate this test for population screening to detect colorectal cancer and advanced adenomas are reviewed here.

Study on the Production of Pentachloronitrobenzene Monoclonal Antibody and Its ELISA Kit for Rapid Detection

[Objectives]This study was conducted to develop an enzyme-linked immunoassay kit that can detect the residual amount of pentachloronitrobenzene in Penaeus vannamei.[Methods]This study was conducted to develop an enzyme-linked immunoassay kit that can detect the residual amount of pentachloronitrobenzene in P.vannamei.[Results]The standard curve range of the kit was 0-8.1μg/L;the detection limit for P.vannamei was 0.912μg/kg;the recovery was 80.6%-103.5%;and the relative standard deviation range within batches was 5.3%-10.1%,and the relative standard deviation range between batches was 6.7%-8.1%.The specificity of the pentachloronitrobenzene monoclonal antibody was relatively good,and the cross-reaction rates with pentachlorophenol,hexachlorobenzene,tetrachlorophthalide,and chlorothalonil were low,all of which did not exceed 30%.The ELISA kit could be stored at 4℃for 12 months,showing good stability.[Conclusions]The detection kit has low cost,short time and small deviation,and is an ideal preliminary screening method.