This experimental method was obtained from the official website of the Fourth Military Medical University

Operation method

Flow cytometry sample preparation experiments by indirect fluorescent staining of micro whole blood

Move

1. Take heparin or EDTA anticoagulated whole blood (100μl/portion) and place it in a 12×75mm special plastic test tube.

2. Add 50μl of specific monoclonal antibody (primary antibody) and incubate for 30min at room temperature.

3. Lysing red blood cells, stabilizing and fixing white blood cells on a Q-PREP instrument.

4. Centrifuge (800~1000rpm, 5min) and discard the supernatant, and wash the cells with PBS twice.

5. Add 50μl of fluorescent (FITC or PE)-labeled secondary antibody, and stain the cells for 30min at room temperature, protected from light.

6. Detect on flow cytometer.

Caveat

1. Fresh whole blood can be used within 8 hours at room temperature; longer time will reduce the activity and affect the results.

2. Anticoagulated blood should be free of clots.

3. When whole blood is added to the test tube, it should be avoided as much as possible to be added to the wall of the test tube, if it is stained on the wall of the tube, it must be wiped off with a cotton swab, otherwise it will affect the separation effect of the cell population of the cell two-dimensional dot plot.

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