title: Cryosectioning and immunolabeling.
abstract: In this protocol, we describe cryoimmunolabeling methods for the subcellular localization of proteins and certain lipids. The methods start with chemical fixation of cells and tissue in formaldehyde (FA) and/or glutaraldehyde (GA), sometimes supplemented with acrolein. Cell and tissue blocks are then immersed in 2.3 M sucrose before freezing in liquid nitrogen. Thin cryosections, cut in an ultracryotome, can be single- or multiple immunolabeled with differently sized gold particles, contrasted and viewed in an electron microscope. Semi-thin cryosections can be used for immunofluorescence microscopy. We describe the detailed procedures that have been developed and tested in practice in our laboratory during the past decades.
authors.affiliation: Cell Microscopy Center of the Department of Cell Biology, University Medical Center Utrecht, Heidelberglaan 100, 3584 CX Utrecht, The Netherlands.
authors.forename: Jan W
authors.initials: JW
authors.lastname: Slot
authors.forename: Hans J
authors.initials: HJ
authors.lastname: Geuze
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D016616:Cryoultramicrotomy / Q000379:methods*; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods*; D008055:Lipids / Q000032:analysis; D011506:Proteins / Q000032:analysis; D051381:Rats; D016707:Tissue Fixation
chemical_list: D008055:Lipids; D011506:Proteins