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s.varasteh 10/11/2012
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اصول و کاربرد های فلوسایتومتری تحقیقاتی
اصول و کاربرد های فلوسایتومتری تحقیقاتی دانشگاه علوم پزشکی وخدمات بهداشتی-درمانی استان قزوین
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Application
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What is Flow Cytometry? The measurement of cell in flow stream which delivers the cells in single file past a point of measurement.
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Flow cytometers are engineered with precise light pathway.
Light is routed through three different types of filter.
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Fluorescence and Fluorochromes
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Fluorochromes Fluorochromes absorb energy (light) and their electrons go from a ground state to an exited state. The electrons return to ground state by emitting light of lower energy , therefore longer wavelength.
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Example of Fluorochromes
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The fluorescence intensity measured is proportional to the number to the fluorescent molecules bound to the cell.
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Using scattered light to determine what is real!
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Visualizing Flow Cytometry data
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Flow cytometry unstained control
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Flow cytometry viability control
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Fluorescence Minus One (FMO)
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Compensation
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Compensation FITC signal is leaking into the PE detector.
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Gate on your cell of interest
Gating Gate on your cell of interest
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When there is an obvious positive and negative population it is easy to use a control (A) to set a marker to define positivity (B).
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Control Negative sample (A) and test sample (B) and (C) shows that probably all cells are weakly positive.
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Negative sample (A) and test sample (B)
Negative sample (A) and test sample (B). The overlay (C) shows that some cells are negative and some are positive.
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