Ultrastructural immunogold localization of some organelle-transport relevant proteins in wholemounted permeabilized nonextracted goldfish xanthophores.
By whole-cell transmission electron microscopy (WCTEM), we recently demonstrated that carotenoid droplets are transported by elongating or retracting endoplasmic reticular cisternae in goldfish xanthophores. Here we report that permeabilized xanthophores demonstrate immunogold reactivity against several proteins involved in organelle translocation. The gold labeling against beta-tubulin and the intermediate filament protein p45a were found on microtubules and intermediate filaments. Labeling with anti-actin was found on nonidentifiable structures, on vesicles of unknown origin, occasional labeling on carotenoid droplets, and on occasional microfilaments. Immunoreactivity was demonstrated with anti-p57 on the carotenoid droplet surface, confirming previous results (Lynch et al., 1986a,b). Labeling with anti-PCD6 subunit (of the inositol trisphosphate/ryanodine receptor) was demonstrated on carotenoid droplets suggesting they possess calcium channels. Anti-MAP 1C (dynein) immunolabeling was generally seen on club-shaped structures in the cytomatrix and on carotenoid droplets. Finally, immunogold labeling with anti-MAP 2a + 2b was seen on a meshwork of microfilaments and intermediate filaments. Finally, this is the first report of a WCTEM technique for permeabilized cells that reveals immunoreactive elements, organelles, and cytomatrix components without the additional requirements of extraction or fracturing.
Kimler
V A
VA
Department of Biological Sciences, Wayne State University, Detroit, Michigan, USA.
Taylor
J D
JD
D000818:Animals; D000911:Antibodies, Monoclonal; D001692:Biological Transport; D015220:Calcium Channels / Q000032:analysis; D002338:Carotenoids / Q000032:analysis; D002856:Chromatophores / Q000737:chemistry / Q000648:ultrastructure*; D004398:Dyneins / Q000032:analysis; D006054:Goldfish; D007150:Immunohistochemistry; D007381:Intermediate Filament Proteins / Q000032:analysis; D007382:Intermediate Filaments / Q000737:chemistry; D016253:Microscopy, Immunoelectron; D008870:Microtubules / Q000737:chemistry; D015388:Organelles / Q000378:metabolism*; D011506:Proteins / Q000032:analysis*; D014404:Tubulin / Q000032:analysis
D000911:Antibodies, Monoclonal; D015220:Calcium Channels; D007381:Intermediate Filament Proteins; D011506:Proteins; D014404:Tubulin; D002338:Carotenoids; D004398:Dyneins