Step I: Cell line submission, recovery, and expansion. 1-2 weeks

1. Two vials of frozen hybridoma cells or one flask of live cells (> 2x10⁶) in culture medium
2. Formula of original culture medium
3. Name of the species from which the cell line was derived and production history, if available
4. Upon receipt, the cell line will be expanded to obtain enough cells for mouse inoculation.

Step II: Inoculation. 2-3 weeks

1. The mice are primed with 0.5ml/mouse of Pristane for two weeks
2. Peritoneal cavity injection of the primed mice with 0.5-1 x 10⁶ hybridoma cells each
3. Ascites production in 7-10 days

Step III: Harvest. 1 week

1. Ascites harvested using aseptic techniques
2. Cellular and lipid materials removed by centrifugation
3. Clarified ascites aliquoted into sterile containers
4. Aliquots labeled and immediately frozen