Cell or Leukocyte Loss During Pelleting

Symptom

Lower than expected DNA yield despite using correct input amounts. Cell pellet may not be visible when working with low cell numbers or certain frozen blood samples.

1. 1 Cell pellet inadvertently aspirated during supernatant removal because pellet location not tracked
2. 2 Leukocyte pellet lost during erythrocyte lysis supernatant removal in blood prep
3. 3 Cell pellet not visible by eye when working with low cell numbers (1–5 × 10⁵ cells)
4. 4 Leukocyte pellet not clearly visible with low blood input (< 500 µl) or certain frozen blood samples

1. 1 Note which side of tube points outward in centrifuge to identify pellet location before removing supernatant
2. 2 Keep pellet side of tube facing downward and remove supernatant from upper side only
3. 3 Leave < 20 µl of supernatant behind to ensure pellet is not disturbed
4. 4 For frozen blood or aged blood with sticky leukocytes: ensure complete resuspension after all pelleting steps