Hi there!
Has anyone faced this problem, when i add the elution buffer to spin columns then centrifuge them I didn’t get the same volume as which I added, why ? does that relate to increasing the centrifuge time ? l do that for 1 minute is this not enough!
How much do you actualise loose ? A few microliters may remain trapped in the filter by capillarity, but if you elute with > 15-20 microliters, it won’t really matter. If you do worry about yield, you could proceed with a second wash with a small volume of elution buffer.
The eluted volume should be equal to the added buffer. There might be calibration errors in the micropipettes which could lead to different volumes. To verify this take the same amount of water (around 20ul) in different 250ul tubes (at least 5).
At what speed are you centrifuging your samples? QIAGEN recommend 13,000 x rpm.
Alternatively, you could look at eluting in two steps: If you are aiming for 50 ul of DNA, add 25 ul first, incubate for 1 min and centrifuge then repeat.
Pierre Béguin Jack Hopkins Abhishek Kumar thanks a bunch all for ur suggestions i will try them.
- Badges
- Science method