IMPROVED MEMBRANE PRESERVATION OF FLAVIVIRUS-INFECTED CELLS WITH CRYOSECTIONING

Ultra-cryomicrotomy and electron microscopy were used to investigate m embranous structures in dengue virus-infected mammalian and insect cel ls. The cryo-sectioned samples displayed ultrastructure comparable to their resin-embedded counterparts with all previously identified virus -induced structures being observed. Structures not previously identifi ed were also found. In particular, membrane-bound packets of vesicles, 100-200 nm in diameter were seen distributed throughout areas of viru s-induced membrane proliferation. These packets were clearly distinct from virion arrays. Small smooth membrane vesicles, previously found t o contain thread-like enclosures (M.L. Ng, J. Gen. Virol. 68 (1987) 57 7-582), were frequently observed to contain dense staining material, h owever the exact nature of this material remains unclear. Virus-induce d modification of golgi-like and/or ER membranes was also observed and may represent early events in the generation of the smooth membrane v esicles seen during infection. We suggest that cryosectioning is the m ethod of choice to investigate membrane rearrangement induced by this family of viruses and that a diamond knife and modified staining techn iques, as utilised in this report, be employed to enhance morphology a nd section preservation.