Microwave incubation improves lipolytic enzyme preservation for ultrastructural cytochemistry.
Standard methods for the ultrastructural detection of lipase and sphingomyelinase activities in the skin result in considerable loss of structural preservation, often interfering with accurate delineation of enzyme localization in association with specific organelles. Moreover, poor preservation occurs, even after extensive aldehyde prefixation, owing to the prolonged incubation times needed to detect residual enzyme activity, which often require non-physiological conditions. A modified incubation protocol is described here, which uses microwave irradiation in conjunction with drastically shortened incubation times, resulting in both superior ultrastructural preservation and excellent localization in mammalian epidermis. This method should be useful generally not only for the study of lipase localization in skin, but also in conjunction with the cytochemical detection of a variety of enzymes in various types of tissue.
Rassner
U A
UA
Dermatology Service, Veterans Affairs Medical Center, San Francisco, CA 94121, USA.
Crumrine
D A
DA
Nau
P
P
Elias
P M
PM
D000818:Animals; D004817:Epidermis / Q000201:enzymology / Q000528:radiation effects; D005976:Glutaral / Q000737:chemistry; D006651:Histocytochemistry; D008049:Lipase / Q000737:chemistry / Q000378:metabolism* / Q000528:radiation effects; D008297:Male; D051379:Mice; D008812:Mice, Hairless; D008854:Microscopy, Electron; D008872:Microwaves; D015388:Organelles / Q000378:metabolism; D012867:Skin / Q000201:enzymology* / Q000528:radiation effects; D013108:Sphingomyelin Phosphodiesterase / Q000737:chemistry / Q000378:metabolism* / Q000528:radiation effects; D016707:Tissue Fixation
D008049:Lipase; D013108:Sphingomyelin Phosphodiesterase; D005976:Glutaral