Liquid propane jet-freezing, freeze-drying and rotary replication of the cytoskeleton and plasma membrane associated structures in human monocytes.
We have studied the cytoskeleton and plasma membrane associated structures of human monocytes using liquid propane jet-freezing, freeze-drying and rotary replication. We find that the cytoskeleton of these cells consists mainly of actin filaments. The actin filaments were identified by their ability to bind myosin sub-fragment S1. A large fraction of these actin filaments radiates from dense cytoskeletal foci. The presence of such actin filament rich foci also in unfrozen cells was demonstrated using immunofluorescence light microscopic techniques. In addition to coated pits a network of thin fibrils (diameter 4-5 nm) were observed on the protoplasmic side of the plasma membrane facing the substrate. These results indicate that liquid propane jet-freezing may represent a simple, inexpensive alternative to impact freezing at liquid helium temperature for obtaining the high cooling rate necessary for successful freeze-drying and rotary replication of cytoskeletal structures.
D000199:Actins / Q000032:analysis; D002462:Cell Membrane / Q000648:ultrastructure; D003034:Coated Pits, Cell-Membrane / Q000648:ultrastructure; D005455:Fluorescent Antibody Technique; D005612:Freeze Drying; D005615:Freezing; D006801:Humans; D008854:Microscopy, Electron / Q000379:methods; D009000:Monocytes / Q000648:ultrastructure*; D015879:Myosin Subfragments; D009218:Myosins / Q000032:analysis; D010446:Peptide Fragments / Q000032:analysis
D000199:Actins; D015879:Myosin Subfragments; D010446:Peptide Fragments; D009218:Myosins