Preparation of single cell suspensions of cultured cells

Summary

This experiment is based on "Color Atlas of Practical Flow Cytometry", edited by Shukui Wang and Zhenying Zhou.

Operation method

Preparation of single cell suspensions of cultured cells

Materials and Instruments

Trypsin Cells
EDTA-2Na PBS
Centrifuge tube

Move

I. Characterization of Cultured Cells

Generally, cell culture is divided into suspension culture and adherence culture. Due to the proliferation of cells, it is possible to form cell clumps of varying sizes or connect them into pieces. If two or more cells are adhering to one piece or if there are too many cell fragments, the FCM results will be affected, which will lead to the failure of the experiment. Therefore, it is very important to prepare qualified single-cell suspensions of cultured cells.

Procedures for the preparation of cultured cell samples

1. Digest the cultured cells with 0.04% ethylenediaminetetraacetic acid disodium salt (EDTA-2Na) or 0.29% trypsin for 3~7 min (depending on the room temperature) until sieve-like gaps appeared between the adherent cells under the light microscope, discard the digested solution, and add the PBS solution.

2. Blow the cells gently down from the wall of the vial with a pipette and transfer them into a centrifuge tube. 3. Centrifuge the cells into a tube.

3. Centrifuge the cells at 800~1000 r/min for 5 min.

4. Discard the supernatant, add 5~8 ml of PBS with pH 7.4, and centrifuge the cells at 800~1000 r/min for 3~5 min. Repeat the procedure 2~3 times to remove cell debris in the cell suspension.

5. Add a small amount of PBS, and gently blow the precipitated cells to homogeneity. Add fixative or keep at low temperature for use.

For more product details, please visit Aladdin Scientific website.

https://www.aladdinsci.com/