Free-floating cryostat sections for immunoelectron microscopy: Bridging the gap from light to electron microscopy.
Frozen skin sections are routinely used for light microscopic immunohistochemical study of the skin basement membrane zone for two reasons: some skin basement membrane zone proteins are labile to routine chemical fixation, and skin is not amenable to vibratome sectioning. However, inherent limitations of conventional frozen sections, including compromised morphology and a requirement for glass slide-mounting, usually limit immunohistochemical study to the light microscopy level. In the present study, we introduce use of unfixed, free-floating cryostat sections for characterization of immunolocalizations of selected skin basement membrane proteins at both the light and electron microscopy level. The new procedure employs free-floating cryostat sections that can be processed as routine tissue specimens and can be subjected to a variety of special staining procedures including immunohistochemistry. Especially useful is the ease of progressive processing of the same tissue specimen from light microscopy to electron microscopy. In this regard, the method renders itself useful when results of immunolabeling experiments need to be elucidated quickly at histological and ultrastructural levels as required for diagnostic and accelerated investigative strategies.
Kan
R K
RK
Comparative Pathology Branch, Comparative Medicine Division, U.S. Army Medical Research Institute of Chemical Defense, Aberdeen Proving Ground, Maryland 21010, USA. Robert.Kan@APG.AMEDD.ARMY.MIL
Pleva
C M
CM
Backof
D R
DR
Hamilton
T A
TA
Petrali
J P
JP
D015703:Antigens, CD / Q000032:analysis / Q000276:immunology; D001485:Basement Membrane / Q000737:chemistry / Q000166:cytology / Q000276:immunology / Q000648:ultrastructure; D024141:Collagen Type IV / Q000032:analysis / Q000276:immunology; D024143:Collagen Type VII / Q000032:analysis / Q000276:immunology; D016616:Cryoultramicrotomy / Q000379:methods*; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D039503:Integrin alpha6; D007797:Laminin / Q000032:analysis / Q000276:immunology; D008853:Microscopy / Q000379:methods; D016253:Microscopy, Immunoelectron / Q000379:methods*; D012867:Skin / Q000737:chemistry* / Q000166:cytology / Q000276:immunology / Q000648:ultrastructure*
D015703:Antigens, CD; D024141:Collagen Type IV; D024143:Collagen Type VII; D039503:Integrin alpha6; D007797:Laminin