Microscopy under pressure--an optical chamber system for fluorescence microscopic analysis of living cells under high hydrostatic pressure.
High hydrostatic pressure (HHP) becomes more and more interesting for life science research, since it can be employed to inactivate various cells. To directly monitor "cells under pressure," the development of an optical high-pressure chamber is required. Therefore, an optical pressure chamber that can be used for up to 300 MPa was constructed. This chamber has already been described as a tool for in situ observation of dynamic changes of microscopic structures in bright field as well as phase contrast. In combination with an inverted microscope, we obtained brilliant microscopic color pictures with an optical resolution more than 0.56 microm. Here, we demonstrate the capabilities of the HHP cell, in combination with epifluorescence microscopy. Using a nonadherent human B-cell line (Raji, ATCC CCL 86), stained with the fluorescent dyes propidium iodide, Hoechst 33342, or dihexyloxacarbocyanine iodide, we were able to show that the system is suitable to perform fluorescence microscopic analyses, with pressures up to 300 MPa, with viable mammalian cells.
Frey
Benjamin
B
Institute for Energy and Environmental Technologies of the Food Industry, Center of Life and Food Sciences, Technische Universitaet Muenchen, D-85350 Freising, Germany. benjamin.frey@med3.imed.uni-erlangen.de
Hartmann
Markus
M
Herrmann
Martin
M
Meyer-Pittroff
Roland
R
Sommer
Karl
K
Bluemelhuber
Gerrit
G
D001402:B-Lymphocytes / Q000166:cytology*; D001562:Benzimidazoles; D002232:Carbocyanines; D045744:Cell Line, Tumor; D005456:Fluorescent Dyes; D006801:Humans; D006874:Hydrostatic Pressure; D008856:Microscopy, Fluorescence / Q000379:methods*; D011419:Propidium; D013194:Staining and Labeling
D001562:Benzimidazoles; D002232:Carbocyanines; D005456:Fluorescent Dyes; D011419:Propidium; C007392:3,3'-dihexyl-2,2'-oxacarbocyanine; C017807:bisbenzimide ethoxide trihydrochloride