The combined procedures of Ficoll-Paque centrifugation and anion exchange separation for the recovery of trypanosomes from blood and other tissues.

The conventional method of separating African trypanosomes from infected blood using a DEAE-cellulose (Whatman Chromedia, DE-52) column was modified by the inclusion of an initial centrifugation step employing the density gradient medium, Ficoll-Paque. On centrifugation in this medium using diluted infected rodent blood the trypanosomes moved into the buffy coat interphase which consisted mainly of lymphocytes with a few contaminating RBCs and neutrophils. The buffy coat interphase layer was withdrawn, washed, resuspended and the trypanosomes finally separated from the contaminating blood cells by passage through buffer-equilibrated DE-52 packed in a 25 ml syringe barrel, representing a considerable saving of DE-52 over the conventional method. The centrifugation step was applied to the investigation of other tissues of infected rodents, such as bone marrow, brain, spleen, liver, lymph nodes, kidney and testis, for the presence of trypanosomes. Using the combined procedure, high recoveries of purified brucei group trypanosomes were obtained from bone marrow, brain, spleen and lymph nodes.