Abstract
It is a serious problem to relate the results of a flow cytometric analysis of a marine sample to different species. Images of particles selectively triggered by the flow cytometric analysis and picked out from the flowing stream give a valuable additional information on the analyzed organisms. The technical principles and problems of triggered imaging in flow are discussed, as well as the positioning of the particles in the plane of focus, freezing the motion of the quickly moving objects and what kinds of light sources are suitable for pulsed illumination. The images have to be stored either by film or electronically. The features of camera targets and the memory requirements for storing the image data and the conditions for the triggering device are shown. A brief explanation of the features of three realized flow cytometric imaging (FCI) systems is given: the Macro Flow Planktometer built within the EUROMAR MAROPT project, the Imaging Module of the European Plankton Analysis System, supported by the MAST II EurOPA project and the most recently developed FLUVO VI universal flow cytometer including HBO 100- and laser excitation for fluorescence and scatter, Coulter sizing as well as bright field and and phase contrast FCI.
Highlights
Relating the results of flow cytometric measurement to the more or less known different species in a sample represents a serious problem
The time conditions for generating sharp images depend on the flow speed and the optical magnification, which defines the extension of the image on the camera target (Fig. 2)
If the nominal imaging point is in the center of the field of view and the trigger delay is fixed, an overall change of the mentioned parameters by 5 μs means that the particle to be imaged is not yet in or has already left the field of view when the flash is triggered
Summary
SUMMARY: It is a serious problem to relate the results of a flow cytometric analysis of a marine sample to different species. Images of particles selectively triggered by the flow cytometric analysis and picked out from the flowing stream give a valuable additional information on the analyzed organisms. The technical principles and problems of triggered imaging in flow are discussed, as well as the positioning of the particles in the plane of focus, freezing the motion of the quickly moving objects and what kinds of light sources are suitable for pulsed illumination. The features of camera targets and the memory requirements for storing the image data and the conditions for the triggering device are shown. A brief explanation of the features of three realized flow cytometric imaging (FCI) systems is given: the Macro Flow Planktometer built within the EUROMAR MAROPT project, the Imaging Module of the European Plankton Analysis System, supported by the MAST II EurOPA project and the most recently developed.
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