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Flow Cytometry Cheat Sheet Cheat Sheet by

Flow Cytometry Cheat Sheet

What is Flow Cytometry?

Flow cytometry is a technique used to analyze and measure charac­ter­istics of cells or particles in a fluid sample.
It provides quanti­tative data about cell popula­tions based on their physical and chemical proper­ties.

Basic Components of a Flow Cytometer

Fluidics System
Controls the flow of the sample through the instru­ment.
Optics System
ncludes lasers, detectors, and filters for detecting and measuring light signals.
Electr­onics System
Converts optical signals into electronic signals for data analysis.

Data Acquis­ition and Analysis

Forward Scatter (FSC): Measures the size of partic­les­/cells.
Side Scatter (SSC): Reflects the internal complexity or granul­arity of partic­les­/cells.
Fluore­scence Detectors: Capture emitted light at specific wavele­ngths from labeled cells.
Compen­sation: Corrects spectral overlap between fluoro­chromes to obtain accurate fluore­scence measur­ements.
Gating: Defines specific popula­tions of interest within the sample based on fluore­scence intensity and scatter charac­ter­istics.

Consid­era­tions and Tips:

Choose approp­riate fluoro­chromes and antibodies for specific targets.
Use approp­riate controls (positive, negative, compen­sation) to ensure accurate results.
Maintain instrument perfor­mance through regular cleaning, calibr­ation, and quality control.
Optimize staining protocols and instrument settings for best results.
Data analysis software skills are crucial for extracting meaningful inform­ation.

Staining and Sample Prepar­ation

Fluore­scent Labels: Use fluore­scent dyes or antibodies to label specific cell components or markers of interest.
Fixation and Permea­bil­iza­tion: Helps preserve cell structure and allows the entry of antibodies into the cell.
Compen­sation Controls: Used to correct for spectral overlap between different fluoro­chr­omes.

Applic­ations of Flow Cytometry

Immuno­phe­not­yping: Identi­fying and charac­ter­izing different cell types in a mixed popula­tion.
Cell Cycle Analysis: Determ­ining the distri­bution of cells in different phases of the cell cycle.
Apoptosis Detection: Assessing cell death and apoptosis using markers like Annexin V and propidium iodide.
Intrac­ellular Protein Analysis: Analyzing protein expression within cells using antibodies and permea­bil­ization methods.
Cell Sorting: Separating specific cell popula­tions based on defined criteria using high-speed drople­t-based sorting.

Read More About Flow Cytometry



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