IGC

Protocols


In this section we provide several success ful protocols for sample preparation and cell sorting experiments. We detail the conditions in which samples are prepared for sorting and how the machines are setup and run for cell separation.



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Preparation for Cell Sorting

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Antibody staining of surface antigens

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Intracellular staining - PFA & saponin

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Intracellular staining for Transcription Factors

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Intracellular staining for cytokines


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Cell Cycle protocol for FC


Yet to be finnished. If you have any questions, contact the FCF staff.


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CFSE staining


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Ficoll Gradient
to isolate PBMCs

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Thermofisher's Cell Preparation Protocol

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Thermofisher's Viability Staining Protocol

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Thermofisher's Surface Staining Protocol



Other tools



Viability Dye Assistant (beta version)

As cells die, the membrane becomes permeable. This allows for antibodies to penetrate the cells, which might be misleadingly considered target cells. This is why a viability dye is crucial in any Flow Cytometry experiment that aims to evaluate live cell lineage and functionality.

If your protocol for sample preparation involves the fixation of your cells, you should choose a fixable viability dye! This will ensure the preservation of the staining pattern after fixation.

Here's a small app to help you choose the viability dye that best suits your needs. Do NOT forget to read carefully about the viability dye you choose before using it and check that it is compatible with your experiment.


Compatible Viability Dyes
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Dye Information

This dye can be used in the following cytometers:

Aria
MoFlo
X-20
Fortessa
Calibur

How to use: You have a list of several viability dyes. You can filter the dyes based on which cytometer your planning to use, whether your sample preparation involves fixation or not, and which channels are already occupied by your other dyes.

Click on a dye on the table to know more about it.