Table I.

Distribution of gp96-Au particles in preendosomal structuresa

Competitor% Au Particles in Coated Structures% Au Particles in Noncoated Structures
None3268
BSA4258
gp96892
HSC703367
  • a D2SC/1 cells were incubated with gp96-Au alone or with gp96-Au and a 500-fold molar excess of unlabeled competitor for 60 min at 4°C followed by 30 min at 20°C. Cells were then fixed at 20°C and prepared for electron microscopy. At 20°C, prelysosomal late endosomes no longer fuse with newly formed endosomes. Indeed, at this relatively low temperature, the number of coated pits per cell section was the highest (up to 11) and therefore the reduction observed is more reliable. We did not attempt to compete HSC70-Au10 molecules with unlabeled HSC70 or gp96 molecules because the low number of coated pits observed with HSC70-Au10 did not allow a reliable quantitation of the inhibition experiments. For each experiment, 30 cells were randomly selected and the gold particles in preendosomal structures were counted (about 700 Au-particles for each condition) and converted to % Au particles in coated or noncoated structures.