Tips for low volume pipetting

Hi all-

I’m working on a method which will need to stamp 3uL of reagent into an empty PCR plate. I’m using 10uL tips and I’ve tried just about everything I can think of (surface empty, jet empty, side touch, different flow rates etc), but can at best only seem to get liquid into about 10% of the wells.

Does anyone have strategies or recommendations to reliably achieve this sort of transfer?
Here are the liquid class parameters I’m starting from:

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Hi @mgildemeister

What is the specific problem(s) you are facing with the transfer? For example, is it not aspirating enough liquid (i.e. you telling the Hamilton to aspirate 3 uL but its only aspirating 1 uL)? Or is it not dispensing properly?

The issue is in the dispense step where it’s not actually dispensing into the majority (~90%) of wells. The wells where I have seen it dispense have been accurate enough, and I’ve confirmed that there is liquid in the tips which don’t dispense. The liquid isn’t abnormal in terms of viscosity/volatility.

Try using the 50 uL tips if you have them. I have tried using the 10 uL tips in the past to stamp 3 uL of aqueous samples but I kept getting inconsistency in both aspiration and dispensation despite my optimization attempts. However, as soon as I switched to the 50 uL tips, even with the default liquid class parameter, I got much better results.

Another suggestion, if you haven’t done it yet, is to teach the instrument where exactly the bottom (and center of it) of the well is. The labware models provided in the software aren’t always accurate so teach the instrument with the 96-head and the same type of tips you will use. Without much context on your protocol, it is possible that the tips (or some of them) are hitting the bottom of the well, preventing the liquid from dispensing. Once you measure where the bottom of the well is, start by setting the pipetting height at 1 mm.

If you haven’t read these yet, I also suggest taking a look at these two links for additional references. :slightly_smiling_face:

Thanks!! I’ll look into all of this.

Of course! Feel free to ask additional questions. Currently, I am downsizing volumes in my protocols so I could give my two cents when it comes to transferring small volumes.

For me it only worked with enlarging the blowout but then it worked down to 1 uL consistently. With 50 uL tips!

It’s always going to be very challenging dispensing low volumes into an empty plate.

Is there a reason why the plate has to be empty? Could you not have a buffer in there first? The buffer will help draw the liquid out from the tip.