Selective recovery of living microfilariae from Onchocerca volvulus nodules. Determination of optimal conditions for their culture in vitro for excretory/secretory products

A method for the selective recovery of living microfilariae from Onchocerca volvulus nodules is described. The microfilariae migrate through solidified agar gel into overlayering Hanks balanced salt solution (HBSS). The highest recovery rates of the worms were obtained with 0.3 and 0.4% agar. Optimal conditions for in vitro culture of the larvae in HBSS were established: pH range 7.0-7.5, glucose concentration 2-5 mg/ml for long term cultures, osmolality 200-309 mOsmoles/l, temperature 4°C for prolonged cultures and 24-28° C for overall best yield of excretory/secretory products (ESP). Subculturing of the larvae reduced contamination of ESP with human serum protein to minimal amounts after 9 recultures done within 96 hr.