Controls, standards, and histogram interpretation in DNA flow cytometry.

Publisher Summary This chapter presents technical details for controls, standards, and criteria used to evaluate and interpret DNA flow cytometry measurement (FCM) data that have proved efficient and reproducible. Guidelines proved to be useful in setting up DNA FCM methods in their laboratories and stimulate suggestions, improvements, and eventually standardization. Controls and standards as well as defined criteria to interpret DNA histograms are essential components of DNA FCM methodology. One of these controls is the cytologic or histologic examination of the specimen. This control is necessary for evaluating the quality of the specimen and for assessing the percentage of tumor versus nontumor cells present. Another control is microscopic review of the fluorescently stained cells or nuclei. This control provides information regarding the specific uptake of dye into the nucleus, integrity of cells or nuclei, and the degree of cell aggregation. Essential to the determination of relative DNA content is a DNA reference standard. Both internal and external standards have been used as normal DNA content reference standards from a variety of sources: chicken red blood cells (CRBC), trout red blood cells (TRBC), human peripheral blood lymphocytes (hPBL), and nonmalignant tissue from the same organ as the specimen. The chapter also discusses the evaluation of DNA histograms that involves the incorporation of defined parameters to allow consistency and quality control of data interpretation.