cytometry protocol

Flow cytometry (FACS) staining protocol (Cell surface staining)

• Harvest, wash the cells (single cell suspension) and adjust cell number to a concentration of 1-5x106 cells/ml in ice cold FACS Buffer (PBS, 0.5-1% BSA or 5-10% FBS, 0.1% NaN3 sodium azide*). ...
• Add 100 μl of cell suspension to each tube.

1. What is flow cytometry used for?
2. What is the principle of flow cytometry?
3. What can flow cytometry measure?
4. How do you stain cells for flow cytometry?
5. Can flow cytometry detect dead cells?
6. Can flow cytometry detect leukemia?
7. Why is laser light used in flow cytometry?
8. How is flow cytometry used in hematology?
9. How do you represent flow cytometry data?
10. Can flow cytometry detect lymphoma?
11. How long do flow cytometry results take?
12. How do you compensate flow cytometry?

What is flow cytometry used for?

Flow cytometry is a laboratory method used to detect, identify, and count specific cells. This method can also identify particular components within cells. This information is based on physical characteristics and/or markers called antigens on the cell surface or within cells that are unique to that cell type.

What is the principle of flow cytometry?

The basic principle of flow cytometry is the passage of cells in single file in front of a laser so they can be detected, counted and sorted. Cell components are fluorescently labelled and then excited by the laser to emit light at varying wavelengths.

What can flow cytometry measure?

Cytometry, in its purest form, is the measurement of cell characteristics, which can include cell size, cell count, cell cycle and more. This technique allows researchers to get highly specific information about individual cells.

How do you stain cells for flow cytometry?

Dilute the appropriate fluorochrome-labeled secondary reagent in 100 µL of Flow Cytometry Staining Buffer and add to the cells. Incubate for at least 30 minutes at 2-8 °C or on ice. Protect from light. Wash the cells by adding Flow Cytometry Staining Buffer.

Can flow cytometry detect dead cells?

Loss of membrane integrity is a definitive indicator of cell death in flow cytometric assays. Cells that exclude a dead cell dye are considered viable, while cells with a compromised membrane allow the dye inside into cell to stain an internal component, thus identifying the cell as dead.

Can flow cytometry detect leukemia?

Flow cytometry immunophenotyping may be performed on blood, bone marrow, or other samples to provide this additional information. It can detect normal cells as well as abnormal cells whose pattern of markers are typically seen with specific types of leukemia and lymphoma.

Why is laser light used in flow cytometry?

Lasers are used in flow cytometry as the excitation source of fluorophore-labeled cell samples. Often, multiple laser colors are needed in order to characterize multiple cellular properties.

How is flow cytometry used in hematology?

Flow cytometry has been used to accurately identify and phenotype the recipient's red cells (44). Flow cytometry is being used increasingly in the blood bank to assess leukocyte contamination in leukocyte-reduced blood products (45)(46).

How do you represent flow cytometry data?

Flow cytometry data is typically represented in one of two ways: histograms, which measure or compare only a single parameter, and dot-plots which compare 2 or 3 parameters simultaneously on a two- or three-dimensional scatter-plot.

Can flow cytometry detect lymphoma?

Flow Cytometry Can Diagnose Classical Hodgkin Lymphoma in Lymph Nodes With High Sensitivity and Specificity.

How long do flow cytometry results take?

These patterns are compared to normal patterns to determine the significance of the results. The test takes approximately three hours and consists of staining the cells, acquiring the cells on a flow cytometer, and then having a skilled technologist analyze the results that have been saved to a computer file.

How do you compensate flow cytometry?

How To Compensate A 4-Color Flow Cytometry Experiment Correctly

1. 4 Steps To Compensating A 4-Color Experiment. ...
2. Choose the correct carrier for compensation. ...
3. Step 2: Collect the data and make sure there is a sufficient number of events. ...
4. Calculate compensation correctly. ...
5. Apply the compensation values and inspect the results.