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Regional Lymph Node Reactivity in Explanted Bladder Cancer of Mice as Measured by Flow Cytometry¹

Urology Service, Department of Surgery [L. G. C., W. F. W.], and Cytology Service, Department of Pathology [Z. D., F. T., T. K. S., M. R. M.], Memorial Sloan-Kettering Cancer Center, New York, New York 10021

The reactivity of lymphocytes in lymph nodes draining the site of a transplantable experimental bladder tumor (MBT2 in C3H/HeJ mice) has been measured in a multiparameter flow cytometry system. Acridine orange was used as a nucleic acid probe. This dye intercalates in helical DNA, emitting green (530 nm) fluorescence upon exposure to blue light; it stacks to single-stranded RNA, emitting red (640 nm) fluorescence. The relative magnitude of the increase of lymphocyte DNA and RNA has been evaluated simultaneously in tumor-draining nodes, in nondraining nodes of the same animal, and in untreated control animals.

Stimulation of the regional node lymphocytes could be observed after 20 days but not after 10 days. It was uniformly high at 35 days. The transcriptive response (increased proportion of lymphocytes with high RNA) was more pronounced than the proliferative (increased proportion of lymphocytes with more than diploid DNA). The histological changes in the stimulated nodes resembled closely those described by others in human tumor-draining nodes. The described method has the advantage of being simple, rapid, and able to measure a representative part of the whole-cell population.

¹ Supported by Grant 126 CA 14134 from the National Cancer Institute through the National Bladder Cancer Program.

² Supported in part by the Emanuel B. Scher Educational Fund, and by the Swedish Society of Medical Sciences.

³ To whom requests for reprints should be addressed, at Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, New York, N. Y. 10021.

Received 11/ 2/78. Accepted 3/ 8/79.