Evaluation of methods for isolating human peripheral blood monocytes. Studies on chemotactic locomotion and other...

We have compared six different methods of purifying human blood monocytes for their usefulness in relation to assays of polarization, locomotion and chemotaxis. For polarization assays it is essential to prepare an unstimulated, spherical, cell population in suspension. The techniques compared were based either on density differences between monocytes and lymphocytes using Percoll or Nycodenz, or on the separation of adherent monocytes from non-adherent cells on protein-coated surfaces, i.e., foetal calf serum (FCS); gelatin-FCS; gelatin-plasma; baby hamster kidney (BHK) microexudate coats. The BHK microexudate technique (Ackerman and Douglas, 1978) gave the best yield and purity of monocytes. These were spherical and had not been activated by the separation procedure. This technique provided monocytes in suspension that were functionally normal in locomotion and chemotaxis assays, phagocytosis, chemiluminescence and Fc receptor expression. To achieve a good yield of spherical cells, it was necessary to use tubes to which monocytes did not adhere. Siliconized glass was superior to tissue culture plastic for this purpose.