Continuous Free.FIow Fractionation of Cellular Constituents in Rat Bone Marrow

By HOWARD C. MEL, LINDA T. MITCHELL AND Bo THORELL T HE WELL-KNOWN heterogeneity of bone marrow cells greatly cornplicates biological, biochemical, and biophysical studies on this tissue. In such a situation two developments would appear highly useful: ( 1 ) an unambiguous, reproducible physical classification of the cellular constituents in the heterogeneous mixture, in the tradition established, for proteins, by Svedberg ( sedimentation rate ) and by Tiselius ( electrophoretic mobility); ( 2 ) preparative separation of large numbers of cells of the different types, for direct investigation of their characteristic biological (e.g., transplantation, immunological ) properties.1 A Start has been made on achieving both of these objectives simultaneously. In a previous publication results were presented on apparently anomolous flow-sedimentation of rat bone marrow cells, considering only the two general classes-nucleated and nonnucleated.2 In the present paper we describe the more detailed continuous preparative fractionation of rat bone marrow as judged by morphological criteria. The different fractions will also be char acterized by rough uncorrected sedimentation rates (in svedbergs). No systematic review of the separation literature will be attempted here. Allfrey3 has reviewed literature prior to 1959. Recent remarks by DeDuve,4 and Beaufay and Bertheta are relevant to general biophysical problems of cell separations, particularly those based on centrifugation (the most common approach). In an interesting paper dealing with bone marrow, Good man6 further reviews the literature, and presents her own experimental results. A very recent report is by Stoloff and Weiss.7 The cell separation problem is a difficult one, and Goodman’s observation, that none of the past approaches has yielded a satisfactory separation from the standpoint of purity, high yield, and cell viability, appears still to be true today.