MICROPLITIS-CROCEIPES TERATOCYTES - IN-VITRO CULTURE AND BIOLOGICAL-ACTIVITY OF TERATOCYTE SECRETED PROTEIN

Teratocytes originate from the dissociation of the extraembryonic sero sal membrane in some Braconidae and Scelionidae. Methods used to cultu re teratocytes in vitro are described and the yield of teratocyte secr eted proteins (TSP) was measured. Although 90% are viable after 6 days , in vitro teratocytes reached only half the diameter (32 mu m) of the same age teratocytes obtained in vivo. Teratocytes cultured in vitro secrete as much as 0.7 mu g of protein per day per larval equivalent ( approximate to 900 cells). Presence of parasitoid larvae enhanced tera tocyte viability while periodic exchange of medium did not. However, m edium exchange significantly increased the total amount of protein sec reted. Size and viability were improved with the addition of 10% FBS t o the Ex-cell 400 culture medium. Non-denaturing PAGE showed at least 15 proteins with molecular sizes estimated to be between 24 to 347 kDa in medium containing teratocytes. An in vitro fat body assay was deve loped to measure the effect of TSP on protein synthesis and juvenile h ormone esterase (JHE) activity. Crude TSP inhibited in vitro incorpora tion of [S-35]-methionine into protein synthesized by the fat body. Th e amount of JHE released from in vitro fat body treated with crude TSP was significantly less than controls, most likely caused by the inhib ition of general protein synthesis. The active fraction of TSP passed through a 30 kDa molecular weight cutoff filter but was retained by a 3 kDa filter. SDS-PAGE revealed four proteins with molecular weights b etween 8 and 20 kDa not present in control medium incubated without te ratocytes. (C) 1998 Elsevier Science Ltd. All rights reserved.