When the titers to the immunogen in the injected mice are elevated, one or two mice will be selected for spleen harvest. The mice selected will undergo a final I.V.(or I.P.) immunization at least three weeks after the last immunization. These mice will be sacrificed 3-4 days after the final boost and blood will be collected. The spleen will be collected and prepared for fusion. The fusion of the spleen cell with P3X63-Ag8.653 (ATCC CRL-1580) cells using a 50% PEG solution. The fusion will be plated in 96 well plates at a total cell concentration of ~1 x 10⁵ cells per well in the HAT selection media. Media will be added to the plates depending on how the clones are growing up after the fusion. After 12-14 days, the fusion plates or clones will be fed with HT media. Fusion plates are usually screened 10-14 days after the fusion was performed. Initial screening of the fusion plates by ELISA is usually performed by core personnel. ELISA positive samples (100-150 microliters) are given to the customer's lab for screening on other assays (immunoprecipatation, immunofluorescence, Western blot, etc.)

FEE: $1324 for a fusion of 10-25 plates

UNC Immunology Core Facility
CB# 7182 / 444 Taylor Hall, Chapel Hill, NC 27599

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