Subcellular fractionation by organelle electrophoresis: Separation of phagosomes containing heat-killed yeast particles

Uptake of foreign material and its subsequent lysosomal degradation is an important function of macrophages. The mechanisms involved in the binding, uptake and delivery of such material to lysosomal organelles, are, however, poorly understood. Here we describe a method using organelle electrophoresis to study the uptake and trafficking of heat-killed yeast particles within murine macrophages. Such yeast particles, which were fluorescently labeled, could be readily detected in intact cells as well as in subcellular fractions. Organelle electrophoresis of a homogenate from macrophages that had internalized yeast particles resulted in the separation of yeast- containing organelles from most other subcellular membranes. In addition, this method was used to follow the kinetics of yeast particle transport within macrophages could be followed readily. Organelle electrophoresis may be a valuable tool for the analysis of phagocytosis.