Design of flow chamber with electronic cell volume capability and light detection optics for multilaser flow cytometry

Abstract

A multibeam optical detection system has been developed with a high optical efficiency, achieved through a reduction in the number of optical interfaces employed in the system. This reduction is made possible by a combination of employing simple lenses, gluing the objective lens directly upon the face of the flow cuvette and the extraction of only one fluorescence signal from each laser beam. A This report describes the design and construction of a flow chamber and associated optical fluorescence detection components that permit the simultaneous use of up to three laser beams for flow cytometric investigations. By employing more than one laser beam, different fluorochromes may be excited sequentially with a spatial separation between excitation locations. This permits separate apertures to be used for each fluorochrome, thus reducing the crosstalk between channels and providing for an increased number of analytical options for the investigator (1,2,5,6,8,9). The new design makes efficient light collection possible by gluing the objective lens directly upon the flow cuvette (4) and by extracting only one fluorescent signal from each laser beam employed. Although the Los Alamos system (7) that we had been using included electronic cell volume capability, we had experienced difficulties with the extensive amount of electrical isolation and shielding required for satisfactory operation. In the new design the shielding and isolation requirements were minimized by the incorporation of fluidic electrical isolation resistance elements located in a small shielded enclosure mounted on top of the flow chamber.

Effects of Voltage-Gated K+Channel on Cell Proliferation in Multiple Myeloma

“…We examined the effect of inhibition of K + channel activity by 4-AP on cell volume (see Figure 5 and Table 2). Cell volume is in direct proportion to the intensity of forward side scatter (FSC), which is used to measure the cell volume [7]. RPMI-8226 cells were cultured in the presence of 2 mM 4-AP for 24 h. The suppression of K + channel activity resulted in 12 ± 3.2%, 19.3 ± 1.6%, and 28 ± 4.3% ( n = 3) increase in cell volume compared with control cells, respectively (Table 1).…”

“…Ten thousand events were collected and the data was analyzed with CellQuest software. Forward side scatter (FSC) describing cell size was used to measure the cell volume [7]. …”

Trypanosoma cruzi: Analysis of the Population Dynamics of Heterogeneous Mixtures1

“…Flow cytometric analysis was performed on PARACYT-I , a custom-built research and development instrument designed in the Laboratory of Parasitic Diseases in collaboration with the Biomedical Engineering and Instrumentation Branch, Division of Research Services of the National Institutes of Health. The instrument utilizes a twosheath flow chamber (42). Two signals/cell, mithramycin-induced fluorescence and forward angle light scatter, were collected.…”