Separation of ascites myeloma cells, lymphocytes and macrophages by zonal centrifugation on an isokinetic gradient.

In attempting to quantitate immunoglobulin synthesis by ascites myeloma cells, we were surprised to note that malignant appearing cells never exceeded 35.9 = 28.0% of ascitic cells and exceeded 22.4 +/- 23.8% of ascitic cells on only one day between the transplantation of the tumor and the death of the host. The ascites tumor suspensions were separated primarily according to diameter, using a previously described isokinetic density gradient of Ficoll in tissue culture medium. This separation resulted in four modal populations of cells: red blood cells, lymphoid cells, macrophages and myeloma cells. The modal populations of macrophages and lymphoid cells always contained less than 0.2% myeloma cells. The purified cells were tested for tumorigenicity. The animal which received the largest number of cells from the macrophage zone received 296 times the number of cells which had been determined to be tumorigenic for myeloma cells. The animal which received the largest number of cells from the lymphoid zone received 1600 times the tumorigenic dose for myeloma cells. Neither of these animals has become ill 4 months after receiving the purified cells. We conclude that: a) Experimentalists who use ascites tumors are not justified in assuming that even easily detected quantitative differences between benign and malignant tissues would be reflected in analyses performed using unstandardized unexamined ascites tumor suspensions. b) In the case of the MOPC 104 mouse myeloma, cytologic criteria are adequate for distinguishing malignant cells from the inflammatory cells in the ascites suspension with a high degree of correspondence between cytologic appearance and biologic activity. c) Programmed gradient sedimentation in an isokinetic gradient of Ficoll in tissue culture medium is an effective means of separating malignant cells from benign cells in this particular ascites tumor.