Laser Scanning Cytometer
The CompuCyte's proprietary core technology, Laser Scanning Cytometry (LSC), uses laser-based opto-electronics and automated analysis capabilities to simultaneously and rapidly measure biochemical constituents and evaluate cell morphologies. LSC technology provides a measure of precision and visualization, combined with flexibility that is available in no other cellular analysis system.
Although LSC technology has a flow cytometry heritage, it is not limited to analyzing cells in fluids. Instead, the technology allows automated analysis of solid-phase samples, including adherent cultured cells, tissue sections, cancer tissue imprints, and cytology smears, preserving the sample along with the exact position of each measured sample. This important feature allows the researcher to automatically return to, visually inspect and interrogate specific cells having defined genetic, biochemical, or morphological properties; or to remeasure specimens after retreating them with reagents or drugs.
ApplicationsLaser Scanning Cytometry allows users to:
- Generate stoichiometric data and analyze heterogeneous cell populations.
- Visualize specimens conventionally with either powerful laser-scanning imaging techniques or with a microscope.
- View any number of individual events in the population data.
- Reanalyze the same cells under varying conditions.
- Measure and localize cellular constituents.
- Detect molecular constituents in the surrounding environment and correlate their presence to cell processes.
- Simultaneously study cultured cells at the individual- and colony-level.
- Perform automated analysis on tissue sections and tissue microarrays.
Lasers and detectorsThe LSC system uses three lasers:
- 405 nm, violet laser
- 488 nm, 20 mW argon ion laser (Cyonics Uniphase Model 2014A-20SL)
- 633 nm, 5 mW red HeNe laser(Cyonics Uniphase)
Fluorescent light is collected and transmitted to the up to four PMT (photomultiplier tube) detectors each measuring different color band. The PMT assemblies process the light selected by each filter. The filters, dichroics and PMTs are in removable modules that can be changed depending on the fluorochromes to be analyzed. Available optical filter tubes (filter ID, wavelength/bandpass (at 50% peak height) and the fluorescent dye typically used with the filter are specified):
- B: 463/39 (DAPI)
- D: 530/30 (FITC, Green Fluorescent Protein)
- J1: 580/30 (PE)
- K1: 625/28 (PI)
- H1: 650/LP (CY5)
Microscope unitThe microscope unit consists of:
- standard Olympus BX-50
- adapted motorized stage (1)
- the manual stage positioning controls (2), located in front of the microscope, allow for coarse or fine operator control of the motorized stage positioning.
- forward scatter detector (3) located under the stage,
- intermediate tube assembly (4) which accepts the scanning laser beam emanating from the microscope housing
- two CCD cameras (5) and monitor
The microscope is equipped with the objective turret containing 10X, 20X and 40X Universal Plan Semi-Apochromatic objectives. An auxiliary position exists for the placement of an objective of your choice. It also contains a crosshair alignment tool. For each objective in the microscope, the nominal beam or spot size at the specimen slide are:
- 10X: 10 microns
- 20X: 5 microns
- 40X: 2.5 microns