Studies on human peripheral blood lymphocytes in vitro. I. Biological and physicochemical properties of the pokeweed mitogen

Abstract

The saline extract from the roots of Phytolacca americana (pokeweed) possesses three biological properties; hemagglutinin, leukagglutinin, and mitogen. Fractionation and further purification on calcium phosphate column chromatography revealed that the biologically active substance was eluted in the front moving fraction with 0.05 M phosphate buffer pH 7.5. Analytical separation on polyacrylamide gels in disc electrophoresis yielded a single homogeneous band with an R(f) value of 0.43 containing all three biological activities. This fraction had an ultraviolet absorption spectrum similar to PHA, was stable to both periodate and mercaptoethanol treatment and gave a single band in double diffusion and immunoelectrophoretic analysis against the antibody prepared to the crude PWM saline extract. Absorption studies with red cells or stroma revealed that the hemagglutinin could be selectively removed without significantly altering the mitogen, whereas absorption with leukocytes resulted in loss of both the mitogenic and leukagglutinating activities.