Immunization and Fusion

Immunization

Routinely, we immunize two rats and one mouse. The antigen, together with CpG, is emulsified in Freund’s incomplete adjuvant and injected intraperitoneally and subcutaneously. For peptide immunizations, we perform immunizations with a peptide mixture. The immunization step is followed by one boost injection without adjuvant 6-8 weeks after immunization.

Fusion

Immunized animals are sacrificed 3 days after the final boost injection and splenocytes are collected and prepared for fusion with the mouse myeloma cell line P3X63-Ag8.653 using a 50% PEG solution. In our standard protocol the fused cells are plated in 24 96-well plates in HAT selection media.

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