Centrifugal cytology. III. The utilization of centrifugal cytology for the preparation of fixed stained dispersions of cells separated by bovine serum albumin bouyant density centrifugation.

Abstract

This paper describes the modification of Centrifugal Cytology for the preparation of permanent, fixed, stained dispensions for both light and scanning electron microscopy of cells which have been isolated on bovine serum albumin (BSA) boyant density gradients. The principal problem with BSA gradient fractions is that the albumin which is present even after dilution is precipitated by the glutaraldehyde fixative. This problem has been solved by the layering of an intermediate D2O solution under the BSA and subsequent removal of the BSA solution and the underlaying with D2O containing glutaraldehyde. A special layering machine facilitates and expedites these operations. This technique has also been applied to BSA-seperated guinea pig and chicken bone marrow cells, as well as Ehrlich ascites tumor cells, hen and human blood cells. The number of celll present in each area of the slide is maintained at a constant value by utlizing a table of dilution factors. This table was generated by a computer program which calculates the concentration of cells present in the rractions and divides it by the number of celll desired.