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Following the fusion and drug selection, the next step is to isolate the hybridomas producing antibodies of interest through the screening process. The choice of the appropriate screening strategy is one of the most important parts of hybridoma production. This screening strategy needs to be determined prior to the fusion. Each assay in the screening strategy should be fully developed and when possible, should be tested using the sera from an immunized animal, because there will not be enough time to eliminate problems with the assay during the screening of the fusion. The initial assays used to screen the fusion must be able to screen a large number of samples. Many hundreds or thousands of samples will have to be assayed, and unless the initial screening assay can be done on a large number of samples, and at a reasonable cost and effort, the isolation of interesting hybridomas is likely to fail. In addition, the assay must be sensitive in order to be able to detect low levels of specific antibodies present in the fusion supernants. Since a monoclonal antibody that works in one assay may not function in another, the eventual use of the antibody must be considered when designing the screening strategy. As soon as possible, the antibodies need to be tested in the assay in which they are intended to be used. The immunology core facility has an automated microtiter plate washer which allows for highly controlled washing of large numbers of assay plates. The core will screen fusions by microtiter ELISAs. Standard ELISA assays done by the facility include IgG detection assays and specificity assays using purified proteins or peptides. The facility may assist in other assays (i.e. flow cytometry, immunohistochemistry, etc.) depending on the specific requirements of the screening strategy that the immunology core works out with the investigator. FEES:
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