The Application Of Protein Biochips To Identify The Type And The Subtype Of Interferon

Objective To establish the monoclonal antibodys against IFN, study their charactorization. Adopting the 6mAb which are given or we have prepared, we establish a protein biochip flat to identify the subtype of interfon. Methods Using hybridoma techique, we prepare IFN as antigen. The hybridoma cells are obtained by fusing spleen cells of Balb/c mice to myeloma cells SP2/0 and culured in the HAT selective medium. The positive clones are screened by indirect ELISA and the limited dilution methods are used to select anti-IFN McAbs. We finally get the hybridoma strain which secrete antibody against IFN. McAb class and subclass are identified by Immunotype? kit. The Westen blotting method is analysed the specificity of McAb. Recombinant human interferon are immobilized onto the NHS modified gold biochips, then the protein biochips are incubated with the 6mAb. Cy3 conjugated sheep anti-mouse IgG is used to detect the antigen-antibody. Arrays are imaged using a fluorescence scanner (GenePix4100A) at 532nm for cy3. Compare this method with the traditional ones: ELISA, Western Blot. Results One stain of hybridoma cells secreting IFN mAbs is obtained. It has high speciality. Its agglutinaton titer is more than 1: 5000 and its subclass is IgG2b. The results getting from the protein biochips show that this flat has speciality to identify the subtype of interfon. Conclutions The antibodies against human IFN have been prepared successfully. The protein biochip plat can provide a practical method to identify the subtype of interfon. It also has some advantages: high quantities; simple operation; low sample dosage and high repetition.