NASA/American Cancer Society high-resolution flow cytometry project - III. Multiparametric analysis of DNA content and electronic nuclear volume in human solid tumors

Background: The NASA/American Cancer Society (ACS) flow cytometer can simultaneously measure electronic nuclear volume (ENV) and DNA content of nuclei. The preceding articles in this volume ("NASA/American Cancer Society High-Resolution Flow Cytometer Project-I") described the schematics, performance, and procedures used for the preparation of nuclei for analysis on this unit. In the present article, we describe the analysis of selected human tumors using the ratio of ENV/DNA content (nuclear packing efficiency [NPE]). Methods: Tumor specimens (frozen) were minced with scalpels and stained with 1-10 μg/ml of 4',6-diamidino-2-phenylindole (DAPI) dihydrochloride at pH 6.0 - 7.2. Trout erythrocytes were used as internal standards. Data on ENV and DNA content were collected in list mode files. Propidium iodide-stained nuclei, analyzed on a Coulter XL cytometer, were used for comparison. Results: Simultaneous measurement of ENV and DNA makes it possible to discriminate between hypodiploid or hyperdiploid tumor cells, as well as to differentiate between near-diploid aneuploid and diploid cells on the basis of their increased ENV. The NPE ratio is a valuable parameter for the detection of small quantities of tumor cells, separating overlapping diploid and aneuploid populations for cell cycle analysis and characterizing the level of differentiation in some tumors. Conclusion: NPE analysis provides unique measuring capabilities for the study of human solid tumors by flow cytometry.