Hi,
I am currently using Bio-Rad Automated Droplet generator. I am happy with the result. I get on average 19K to 21K droplets per reaction. However, I have also encountered 1 to 2 samples out of 96 with less than 10K droplets and 1 in 300 samples have droplets above 25K. In both cases, the analysis fails. How can I avoid this?
Jing Zhu Thanks, I got you but here we are talking about an automated droplet generator which shouldn't fail. We buy all the consumables from Bio-Rad and they are all ddPCR grade. Thank you
Hi, in theory there can be a bit more then 25K droplets. You can overrule the analysis with a manual threshold there.
Under 10K droplets can happen due to:
- not enough mixing (unlikely for one well)
- samples type (but if it is pretty random, probably not)
- sometimes a piece of plastic that comes loose from a pipet tip somewhere in the workflow, a bead from extraction, ....
- sealing protocol is very important. Be sure to seal 1x 5" at 180°C for the QX200/QX600 workflow.
- ....
It is possible to overrule the analysis here too. (Off course, the plot still needs to look normal and you will also miss a bigger proportion of your sample.)
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