NUTRITION OF ANIMAL CELLS IN TISSUE CULTURE: VIII. DESOXYRIBONUCLEIC ACID PHOSPHORUS AS A MEASURE OF CELL MULTIPLICATION IN REPLICATE CULTURES

Abstract

A chemical method of estimating the size of cell populations in replicate cultures has been devised for use with certain cell strains from the mouse and rat. The method is based upon the determination of desoxyribonucleic acid phosphorus (DNAP). Although the method may be used as an alternative to the procedure of counting isolated, stained nuclei in a hemocytometer, it actually supplements such measurements and provides useful information on the DNA-content per cell. The method consists of a modified Schmidt and Thannhauser separation of ribonucleic acid (RNA) and desoxyribonucleic acid (DNA) followed by the spectrophotometric estimation of the orthophosphate. The present communication describes the procedures in detail and reports recoveries of RNAP and DNAP from mixtures of highly polymerized nucleic acids and various types of biological material. Good correlation was found between nuclear counts and DNAP values in natural media (containing horse serum and chick embryo extract) and in synthetic (chemically-defined) media. Finally, the efficiency of the dispensing apparatus most frequently employed in preparing replicate cultures from continuously stirred, washed cell suspensions has been determined by measuring the DNAP content of serially dispensed samples.