Rapid-freezing cytochemistry: preservation of tubular lysosomes and enzyme activity.
We show that tubular structures present in phorbol ester-stimulated macrophages are sensitive to commonly used chemical fixatives (i.e., they usually become fragmented during fixation). These structures are well preserved in macrophages that are physically fixed by rapid-freezing and subsequent freeze-substitution in osmium-acetone. We have developed methods that combine rapid-freezing, freeze-substitution, and enzyme cytochemistry for preservation of these tubular structures and for detection of endocytosed material (i.e., horseradish peroxidase). This method of rapid-freeze cytochemistry may be useful in other situations where chemical fixation does not adequately preserve cell structures, particularly of membrane compartments.
Robinson
J M
JM
Department of Cell Biology, Neurobiology, and Anatomy, Ohio State University, Columbus 43210.
Karnovsky
M J
MJ
D000818:Animals; D015925:Cryopreservation; D004705:Endocytosis; D006651:Histocytochemistry; D008247:Lysosomes / Q000378:metabolism* / Q000648:ultrastructure; D008264:Macrophages / Q000201:enzymology* / Q000648:ultrastructure; D051379:Mice; D008813:Mice, Inbred ICR; D008856:Microscopy, Fluorescence; D015388:Organelles / Q000378:metabolism* / Q000648:ultrastructure; D014021:Tissue Preservation