Abstract:

BACKGROUND: Apoptosis-inducing factor not only has the functional redox and electron transfer function, but also has a pro-apoptotic function, so it plays an important role in maintaining the normal physiological activity of the cells.
OBJECTIVE: To prepare mouse anti-human apoptosis-inducing factor monoclonal antibodies and to identify their biological characteristics.
METHODS: The Ensembl database and DNAstar package were used to analyze amino acid sequence of apoptosis-inducing factor and obtained the epitope polypeptide. Carbodiimide method was used to prepare the complete antigen immunized animal through coupling the polypeptide with a carrier protein. The BALB/c mice were immunized with a designed apoptosis-inducing factor peptide (KLH-coupled), and then the anti-AIF McAb was obtained by means of the B lymphoma hybridoma technique. Hybridoma technique was used to prepare and purify the apoptosis-inducing factor monoclonal antibodies.
RESULTS AND CONCLUSION: The detection of indirect ELISA method showed that titer of apoptosis-inducing factor monoclonal antibodies in ascetic fluid reached 1: 252 400. The result of Western blot demonstrated that there was specific Mr 670 000 brand which consistent with the antigen band. Immunohistochemical detection results showed there was brown positive expression in the colon cancer cells, which indicated that the antibodies can also be used for immunohistochemistry staining. Hyper-activity, specific and purified mouse anti-human apoptosis-inducing factors have been successfully obtained and identified.