Separation of mononuclear cells (MNCs) by density gradient separation Reagents and materials: experimental method: Rayon Swab ,Micro Head Foam Swab ,Lint Free Polyester Swab,Oral Swab Sticks Hunan Kangfutai Medical Devices Co., Ltd. , https://www.3kfut.com
1. A suitable medium or frozen solution;
2. PBSA;
3. Ficoll-Hypague;
4. Pasteur pipette;
5. 50ml centrifuge tube;
1. Dilute the cord blood sample with PBSA in a 1:1 ratio;
2. Inhale 15ml Ficoll-Hypague into a 50ml centrifuge tube;
3. Slowly add 30 ml of PBSA diluted cord blood sample to Ficoll-Hypague;
4. Centrifuge at room temperature 450g for 20-30min;
5. After centrifugation, a layer of cells is clearly visible on the Ficoll-Hypague liquid surface, which is a mononuclear cell layer with a lower density than Ficoll-Hypague solution;
6. Using a Pasteur pipette, aspirate the mononuclear cell layer and transfer it to another centrifuge tube;
7. Wash the cells with PBSA and centrifuge at room temperature for 200 min at room temperature for 10 min;
8. Pour off the supernatant, resuspend the cells with PBSA, wash the cells, repeat step 7 above;
9. Finally, resuspend the cells (ice or medium) with a suitable medium;