Study On The Determination For Tetanus Antibody Based On Immunochromatographic Test Strip And Nanoparticle

Tetanus is triggered by the action of highly potent toxin which is released by an anaerobic bacteria Clostridium tetani. Since tetanus toxin can damage the body’s neuromuscular tissue directly, the symptoms after onset are more severe,and the mortality rate is relatively high. Therefore, detecting tetanus antibody in human serum quickly and accurately plays a very important role of guidance for the treatment of tetanus. The immunochromatographic strip technology, holding the advantage of simple, rapid, low requirement of laboratory equipment and bedside diagnosis, has good prospects to be used for clinical analysis and medical diagnostics.In this study, we developed two assays for tetanus antibody, one is colloidal gold-based immunochromatographic strip following the principle of double antigen sandwich method, and the other is quantum dots-based fluorescent immunochromatographic strip. The details are as following:1、Preparation of colloidal gold nanoparticles-based immunochromatographic strip for the detection of tetanus antibody.First, colloidal gold nanoparticles with the diameter of about 40 nm were prepared by trisodium citrate reduction. After the exosyndrome of color and UV absorption, these gold nanoparticles were labeled with the tetanus antigen by electrostatic adsorption. Tetanus antigen and tetanus antibody were fixed on nitrocellulose membrane as test line and control line, respectively. The principle of colloidal gold nanoparticles-based test strip method was based on the reaction between tetanus antigen(test line) and colloidal gold-tetanus antigen to combine with target tetanus antibody. The visual detection limit of the test strip for tetanus antibody was found to be 0.01 IU/mL. The test strips could be preserved at 4°C for 6 months without significant loss of activity. Furthermore, the detection results of the colloidal gold nanoparticles-based strips were consistent with those of enzyme linked immunosorbent assay(ELISA).2、Preparation of Cu:Zn-In-S/ZnS quantum dots-based immunochromatographic strip for the detection of tetanus antibody.The oil-soluble Cu:Zn-In-S/ZnS quantum dots(QDs) were prepared and then were transformed to water-soluble quantum dots through mercaptoundecanoic acid’s ligand exchange. IR, UV, fluorescence spectroscopy and stability of optical characterization for the quantum dots were carried out. Then, purified water-soluble Cu:Zn-In-S/ZnS QDs were conjugated with goat anti-human IgG(Fc) using chemical bonding reaction. Test line and control line were zoned with tetanus antigen and human IgG. The principle of this sensor is based on sandwich immunoreaction, where analytes combine with QDs-conjugated Goat Anti-Human IgG(Fc) and then bind to tetanus antigen on a test zone. The detection limit of this strip sensor was 0.001IU/m L, which is 10 times lower than that of gold nanoparticle-based test strip.The result of cross reaction among diphtheria antibody, pertussis antibody, measles antibody and rabies antibody showed a very good specificity. This assay had a good linear response relationship for the detection of spiked serum samples. The method could not only analysize tetanus antibody quantitatively, but also hold the great potential application for point-of-care(POC).