The Pre-test Of Bladder Cancer Multi-index Colloidal Gold Protein Chip

Objective:Use human serum IgG, IgM for the target detection protein. Preparing the simultaneously detection multi-target protein chips, as the prophase test preparation of bladder cancer multi-target detection.Methods:On the nitrocellulose membrane according to coubling dilution simultaneously coating two kinds of antibodies, goat anti-human antibody IgG and rabbit anti-human antibody IgM, preparing protein chips. Use different dilution ratio of IgG (standard) as the detection protein antigen specifically crossing with antibodies coated on the nitrocellulose membrane, and then hybridizing with the antibody (goat anti-human antibody IgG) added the colloidal gold label. After completion of staining applying the chip scanner to analysis gray value of the optical signal, and protracted the standard curve of different concentration gradient of IgG in the samples to the gray value of optical signal in order to find the best coated condition, and then improving the silver staining to further reduce the detection lower limit. Therefore, under the same conditions using No.174 serum to hybridize with this chip, and then with the colloidal gold labeled second antibody colored in hybridization, comparing with different detection programs to confirm the best one of multi-target detection, and exploring the best coated condition of multiple antigens.Results:1. the chip could be hybridized specifically with the corresponding antigens, no cross reaction.2. the detection lower limit could be up to 100ng/ml, with good linear and gradient relationship, sensitivity up to 300ng/ml.3. exploring the silver enhance system operated in the daily light conditions, the detection lower limit could further upgrade to 10 to 20 times, which can be used for all kinds of colloidal gold detection antigen or antibody technology.4. Comparison of the colloidal gold blending methods, double antibody mixed colloidal gold labeling method is better, and it had good repeatability.Conclusion:The chip could be used to detect multiple antigen targets simultaneously, and it is an effective new method of antibody detection which can be cooperated with improving the silver enhancement technique to further reduce the detection lower limit.