Optical Standard Setup - Parameters And Filters - SYSMEX CyFlow Ploidy Analyser Instrument Operating Manual

Instrument Operating Manual
Scatter Signals
In the CyFlow
single file through the Nd-YAG laser beam focused into the capillary. This causes
scattering of light providing information about different cellular characteristics. The
side scattered light (side scatter, SSC) is emitted in an angle of about 90° and
provides information about the "granularity" or refraction properties of the particles
or cells. Particles/Cells with a high granularity/refraction create a strong signal in
the SSC.
Fluorescence Signals
Fluorescence occurs when a molecule excited by light of one wavelength returns
to a lower energy state by emitting light of a longer wavelength. Cell nuclei can be
stochiometrical labelled by fluorescent probes and fluorescence emission after
sufficient excitation can be correlated to DNA content.
Optical Standard Setup – Parameters and Filters
2.4
The CyFlow® Ploidy Analyser is equipped with one (11-01-1002) or two (11-01-
1003/11-01-1004) optical parameters for the detection of DAPI fluorescence (11-01-1002)
and/or Propidium Iodide fluorescence and side scattered light (11-01-1003/1004). The
fluorescence and side scattered light is collected by a microscope lens in an angle of 90°.
The light is subdivided into two different wavelength bands by optical filter glasses.
Signal processing with Photomultiplier Tubes (PMT)
Photomultiplier tubes (PMT) collect fluorescence or scatter light signals, which
generate an electronic pulse inside the PMT. The pulse amplitude is proportional
to the number of incoming photons. Following linear or logarithmic amplification,
the signals are converted into electronic signals by an Analog/Digital-Converter
(ADC). As a particle of interest passes the focus, its fluorescence is detected by
the PMT, and an electronic pulse is generated. The acquisition of a signal is
triggered when the signal amplitude exceeds a predefined threshold level. The
threshold is primarily used to reject events such as debris particles or noise from
optical and electronic sources.
Optical filtration
Optical filters transmit a specific wavelength band of the visible light and block
other wavelengths. Optical filtration is used to adapt the wavelength reaching the
optical detector to the emission wavelengths of the used fluorochrome.
Sysmex Partec GmbH
Am Flugplatz 13, 02828 Görlitz, Germany · Phone +49 3581 8746-0 · Fax +49 3581 8746-70 · info@sysmex-partec.com ·
®
Ploidy Analyser 11-01-1003 and 11-01-1004 cells pass in a
CyFlow® Ploidy Analyser Rev-03_2104-10-22
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