Hi. Is there anyone with experience using 384 well plates doing bead cleanup for NGS protocols (using a passiv magnet on the deck)? Anything I should/must consider?
Thanks!
What liquid handler do you have? For 384-well PCR cleanups I would seriously consider ditching beads and going with a fully tip based solution. DPX makes a product that has a silica media in the tip and the bind/wash/elute all happens in the tip, that replaces the beads/magnet completely.
Thanks for the reply. I have a Hamilton STAR, 8ch, 96 head. We are considering buying a STAR with 384 head for the application. What experience do you have with different liquid handlers and the tips you suggest?
I’m a Bravo guy through and through, but I think a Star would be fine for this application. DPX makes tips for many different platforms. Perhaps contact them and see if they’ll send you some free tips to trial it out.
But for me the workflow is much simpler with the Nix tips. A normal bead-based cleanup would use four boxes of 384 tips (if you’re doing 2 EtOH washes), a Nix tip cleanup would use just one. That makes the logistics of getting this done on a liquid handler much simpler. This isn’t as important on something with a large deck like a Star, but on a Bravo it really, really simplifies the protocol. DPX also priced out the tips such that you pretty much break even on cost between a bead-based cleanup and Nix tips.
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Hi @Dr_TL,
In my work with 384w plate bead clean ups, the biggest consideration was the volume used for each reagent. We found that while simply cutting all reagents down to 25% of the original volume was sufficient to yield the data we wanted, it also led to issues when resuspending beads during the final elution step. The bead pellet would sit higher in the well that the liquid could reach, and the amount of liquid was insufficient to resuspend by pipette mixing. Ultimately, we ended up sealing the plate and taking it off deck to shake vigorously to achieve resuspension.
The other way to resolve this issue is to quarter all reagent volumes except resuspension buffer, which leads to a lower concentration initial elution. This is then followed up by reconcentrating the eluate by one’s method of choice.
Thank you,
Dan
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Thanks a lot for the detailed answer. No side-effect such as drying out of the beads etc noticed?
Best regards
Thomas
Hi @Dr_TL,
No more so than one might see performing a 96w plate clean up. A lot of that comes down to timing, and that gets handled the same way with a 384w plate as it would a 96w plate (when being processed by channels), which is breaking the plate down into batches and recoding the method so each batch is processed in a way that the step following a supernatant removal is immediately adding reagent before moving onto the next batch. For example with a 384w plate using the MPH96, instead of removing ethanol from all wells of the plate before adding the next round of ethanol, remove ethanol from quadrant A1, add ethanol to quadrant A1, then repeat for the remaining quadrants. This significantly reduces air exposure timing for the beads.
Thank you,
Dan
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