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Better diagnostic techniques are needed to delineate the size and location of malignant primary tumors and metastatic deposits. We are developing an in vivo tumor visualization technique utilizing 99mTc-labeled immune lymphoid cells. We have investigated the in vivo organ and tissue distribution and tumor localization of 99m sodium pertechnetate, 99mTc, reduced by SnCl2 - 2H2O and 99mTc-labeled viable immune and nonimmune lymphocytes 32 hrs after injection into tumor-bearing mice. The distribution and tumor localization of the 99mTc-labeled compounds and splenocytes within tumor-bearing mice after 32 hrs was determined by imaging with a gamma camera and was quantitated by counting the various dissected organs and tissues in a gamma counter. The gamma camera images and postmortem dissection and counting showed localization of radioactivity at several sites, including the tumor in mice injected with radiolabeled immune splenocytes whereas studies utilizing other 99mTc-labeled substances or nonimmune cells showed little or no localization of radioactivity in the tumor when compared to normal leg tissue.

A method is described for the inexpensive and easy preparation of colloidal silica--aluminum modified--polyvinylpyrrolidone (CS-AM-PVP) density gradient centrifugation medium. Using density gradient centrifugation, several cell separation and biochemical characteristics were studied: centrifuge tube wall cell adherence, mouse spleen cell density distribution, rebanding properties, mitogen response and cell aggregation. Cell adherence to the centrifuge tube wall using density gradients of CS-AM-PVP was compared with density gradients of bovine serum albumin and Ficoll. Few cells adhered to the centrifuge tube wall when CS-AM-PVP was used as a gradient medium; whereas, significant cell adherence to the centrifuge tube cell wall occurred when bovine serum albumin or Ficoll was used as a gradient medium. The CS-AM-PVP gradient medium did not inhibit the response to mitogens of mouse spleen cells which had been separated into density subpopulations in a discontinuous CS-AM-PVP density gradient, caused a minimum amount of cell aggregation, and was found to be non-toxic.

Mouse-spleen lymphocytes, mouse mammary adenocarcinoma cells (MMA-67) and mouse lymphoma cells (S49.1) were radiolabeled with 111In and their in vivo migration patterns studied over a 24-72 h period after i.v. injection into syngeneic mice. All three cell lines showed different in vivo migration patterns. Initially, some trapping of the cells occurred in the lungs at 15 min. More MMA-67 cells were initially trapped in the lungs than S49.1 cells or lymphocytes. At 24 h, most of the 111In labeled cells had left the lungs and accumulated in various tissues and organs. At 48 and 72 h, only slight changes in the localization patterns of the 111In radiolabeled cells in vivo were noted when compared to the 24 h patterns. About 35% of the injected 111In was excreted from the animals over the 72 h period of these experiments. In vitro studies compared the growth and/or viability of 111In radiolabeled lymphocytes or tumor cells to nonradiolabeled lymphocytes or tumor cells. The percentage of 111In released from the cells in culture was also determined. Similar values for growth and/or viability were obtained between non-radiolabeled cells and MMA-67 cells radiolabeled with 1.35 dpm 111In per cell; S49.1 cells, 0.51 dpm111In per cell; and lymphocytes, 0.04 dpm 111In per cell.

A method for the separation of viable and non-viable P815X2 mastocytoma cells is described using Ficoll discontinuous density gradients. Non-viable P815X2 cells are more dense than viable P,15X2 cells and reach a bouyant density equilibrium at a higher Ficoll concentration than the viable cells. This technique allows the separation of viable P815X2 cells from cells that had been killed by antiserum and complement, or by non-growth supporting medium or by cytotoxic lymphocytes. This method is a new assay to determine cytotoxic lymphocyte killing of P815X2 cells.