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Hybridoma Techniques

Determining the Volume of a Lymphocyte Suspension
needed to perform a fusion

__________ Appearance of suspension (on hemocytometer under microsocope)

__________ Number of viable lymphocytes/ml of suspension

__________ Total volume of spleen cell suspension (in milliliters)

(Multiply the number of viable lymphocytes/ml x the total volume of cell suspension)

__________ Total number of viable lymphocytes in that suspension (will you have enough?)

Depending on the total number of myeloma cells you will have for a fusion, you will need to determine what volume of your cell suspension would be needed to obtain a specified number of lymphocytes.

(Divide the number of lymphocytes desired by the number of viable lymphocytes/ml)

Volume of cell suspension needed to deliver

__________ 1 x 10⁸ total lymphocytes in that volume

__________ 8 x 10⁷ total lymphocytes in that volume

__________ 6 x 10⁷ total lymphocytes in that volume

__________ 4 x 10⁷ total lymphocytes in that volume