Light scattering properties of murine hemopoietic cells

The light scatter of cells contains information about cell structure and size. Much of this information can be obtained by measuring the light scattered in a flow cytometer in two directions: forward (1.5 degrees-13 degrees) and perpendicular (65 degrees-115 degrees) with respect to the direction of the laser light. For different mouse bone marrow cell types and Sephadex G-25 beads of 10-50 micron diameter, the forward light scatter intensity can be shown to be linearly proportional to the cross-sectional area. The perpendicular light scatter intensity can be shown to depend both on size and degree of structuredness. Therefore, light scatter measurements may be used to obtain overall morphological descriptions of rare cells. By sorting on the basis of light scatter measurements and by subsequent in vivo and in vitro culture assays, it can be shown that the pluripotent hemopoietic stem cell and three committed progenitor cells which represent consecutive stages in the granulocyte/monocyte differentiation series have diameters of 7.1-7.5 micron, and show a complexity of structuredness which increases with differentiation. Since these cells have a low incidence and are only described by their function, such morphological information cannot be obtained by direct microscopic examination of bone marrow. Furthermore, most measurements by flow cytometers can be improved by simultaneous light scatter measurements. Examples are presented which illustrate this in studies of immunofluorescence, leukemic bone marrow, and stem cell purification.