I published the following picture in 1965 in a paper entitled Viremia in Friend Leukemia: the electron microscope approach to the problem which appeared in Pathologie-Biologie, vol 13, pp. 125-134. Transmission electron microscopy was used to verify the success of a method for virus purfication which I had developed when working at the Sloan Kettering Institute in New York. The method was as follows: About 20 ml of blood from leukemic DBA/2 mice was collected, blood cells were removed by low speed centrifugation and the plasma was diluted 1/1 with cold heparinized Ringer's solution. The diluted plasma was cleared from contaminating debris by two consecutive steps of Millipore ultrafiltration, using pore size 0.65 µ first and 0.22 µ next. The second filtrate was then spun at high speed at 30000 g for 2 hours. The resulting pellet, about 1 mm in diameter, was immediately fixed with osmium tetroxide, embedded in epoxy resin and prepared for electron microscopy by routine thin sectioning methods. Aliquots of the unfixed pellet were resuspended in Ringer's solution and used for titration of the leukemogenic activity in adult DBA/2 mice, known to be 100% susceptible to the virus. It. was that simple!

The picture shows, at a magnification of 19500 x, an almost pure population of typical "type C" viruses (not yet called retrovirus in 1965....). Three arrows point at contaminating debris and microvesicles. The interpretation of these EM pictures was that virus purification was satisfactory and that contamination rate was extremely low.

Dangerously enough, EM was progressively dismissed in retrovirus research after 1970. Molecular biologists started to rely exclusively on various "markers", and what was sedimenting in sucrose gradient at density 1.16 gm/ml was regarded as "pure virus". It is only in 1997, after fifteen years of intensive HIV research, that elementary EM controls were performed, with the disastrous results recently reviewed in Continuum. How many wasted efforts, how many billions of research dollars gone in smoke...

Horrible.

Errare humanum est sed diabolicum perseverare....

Etienne de Harven, MD

Member of The Group for the Reappraisal of the HIV/AIDS Hypothesis
Prof. Emerit. (Pathology) University of Toronto