"In vivo cryotechnique" in combination with replica immunoelectron microscopy for caveolin in smooth muscle cells.
A novel "in vivo cryotechnique" with replica immunoelectron microscopy was developed for detecting caveolin localization on replica membranes prepared directly from living smooth muscle cells. After quick-freezing mouse duodenal walls by our "in vivo cryotechnique", the specimens were prepared for freeze-fracture and deep-etch replica membranes. Then they were treated with 5% SDS and 0.5% collagenase to keep some antigens on the replica membranes. The immunogold method could be used to clarify the localization of the caveolin antigen in relation to three-dimensional ultrastructures of living smooth muscle cells. Our new cryotechnique can provide native organization of functional molecules in living cells.
Takayama
I
I
Department of Anatomy, Yamanashi Medical University, Tamaho, Japan.
Terada
N
N
Baba
T
T
Ueda
H
H
Kato
Y
Y
Fujii
Y
Y
Ohno
S
S
D000818:Animals; D051242:Caveolin 1; D022461:Caveolins; D004386:Duodenum / Q000378:metabolism / Q000648:ultrastructure; D005613:Freeze Etching; D005614:Freeze Fracturing; D008565:Membrane Proteins / Q000378:metabolism*; D051379:Mice; D016253:Microscopy, Immunoelectron; D009130:Muscle, Smooth / Q000378:metabolism* / Q000648:ultrastructure; D016628:Replica Techniques
C495541:Cav1 protein, mouse; D051242:Caveolin 1; D022461:Caveolins; D008565:Membrane Proteins