Hi!

A dude of mine has a self-made patch-pipette-stabilizer. It's a simple metal stab with a small notch. So that one can "swing" it in and stabilize the pipette from its back. I've a pic somewhere. If I find it, I can mail it to you.

Hi Richard:

Interesting approach. Please post the picture here.

Huong,

I had similar drift issues when I was using Burleigh manipulators.  They were fine for using when you raise the cell off the plate but for primary cultures they drifted all over the place.  My favorite manipulator is the Narishige MLW-3.  I've never had any issues with any drift with it.  People laugh that I'm old fashion by not getting into the techno manipulators but for whole cell attached recordings I find my MLW-3 the best.

Good Luck!

Ann Marie

Thanks all for the info!

@Andrei: It is not at all a dump question! Yes, I am fairly certain it is not the coverslip since I visualize the cells and pipette at the same time, then marked on the screen of my monitor where the pipettes were. After a while, cells were still at the same position but pipettes moved. :(

@Richard: that sounds awesome! I would love to learn more about how to do this!

@Ann:  Interesting to know! I think we might have one like this somewhere in our lab. I imagine that it must be a bit difficult to manipulate but is more reliable? :)

It is as reliable as they get in my hands!!

Dear Huong, 

Usual suspects are cable backlash (but you loosened those), controller electronic  drift (but you still see drift with the controller off), temperature-induced drift (but you have curtains and no heater). Air conditioning turning on and off or you breathing too close ? Voodoo sometimes works, try spreading rooster blood on the controller. Seriously, you mentioned drifting only in XY (not Z) so temperature is to be excluded but controller malfunction is possible. Can you swap cables so that the z-controller is connected to either X or Y? If the drift plane changes to ZY then I would suggest borrowing a controller from a colleague, for testing.

I hope this helps  

The proble is the micromanipulator

Thanks for the suggestion Thomas and Anaclet. 

@Thomas: LOL. Seems unlikely that it is either the breathing or air conditioner. I have to find some rooster then. J/k. Yes, I will try swapping the cables and see if this is what caused the problem. :) Will keep you updated! 

Dear Huong,

some time ago I had problems with pipette drifts because my table wasn't perfectly horizontal. Although you probably have a good steady air table, I would suggest you to maybe use the "old-school" spirit level and check if it's all perfect in x and y.

Keep us updated,

Best, Alenka

Thanks Alenka!

I've attached an old picture of an "electrode stabilizer" that I had machined (made of metal) that can be fastened onto the headstage and swung in to grip the electrode once its inserted into the electrode holder.

I had what seems like very similar problems when I was doing perforated patch recordings to measure synaptic plasticity. They got severe in the summer, when an air handler over the rig would periodically turn on to cool the room. [I think this lead to air temperature fluctuations that lead to non-uniform thermal expansion of the polyurethane electrode holder -- just enough to move the electrode tip 10s of microns. When I used this "electrode stabilizer" (pic attached), the problem went away immediately. 

Hope this is useful.

Hi Huong,

we solved this problem by making the length of the lever arm as small as possible. In other words, try to move the manipular as close as possible to the recording chamber and thus make the length of the lever arm (i.e the pipette +pipette holder+ manipular arm holder clamp), as short as possible. This solved our mechanical drift problems.

THanks Hendrique and Daniel for very interesting solutions. I will try to implement these ideas and keep you updated! 

I hope you can solve your problem with one or a combination of the nice suggetions above. But just in case (there is still Voodoo :)), have you noticed any dramatic visual change in the cells volume? a dramatic shrinking (or swelling) might probably affect it. I know it's far fetched...I am just trying to find an answer to the Vodoo issue :)

Hi Huong,

There are some small suggestions:

1. Since the drift still can be seen when the manipulator power off, please make sure the manipulator fixed well on the stage.

2. the tube connecting the holder to suction also could be one of source. To check it, just remove the tube.

3. Make sure the pressure of air table is in the normal range. You can find the number in the manual and check the meter of air table.

Good luck!

Hi Huong, I also just noticed you are at Stanford. Han-mi Lee in the Shatz lab made one of these stabilizers a few years ago. If she is still around (or if the machine shop already knows how to make one) this might make this solution easy to try. Best, Dan

Hi Huong,

I am a PhD student electrophysiologist and have experienced my own struggles pipette stability. I recently answered a nearly identical question somebody else asked here on ResearchGate. I will give very similar advice:

I recommend video-recording your pipette in the recording conditions and reviewing it at high speed. In my case, I wrote a script to take a screenshot every 2 seconds and convert it into a video file.

I used a manual micromanipulator and compared it with movement from two axon headstages with different pipette holders and realized the problem was in the axon headstage / holders. https://www.youtube.com/watch?v=JlAYViKUClw&list=UUs3-qUELPY-2AOHa7Q_db5w

I then compared long vs. short plastic pipette holders we had in the lab and realized the shorter ones dramatically outperformed the longer ones https://www.youtube.com/watch?v=_-4f0T2Hifo&list=UUs3-qUELPY-2AOHa7Q_db5w

I imagine the plastic (or the rubber holding the pipette) is changing its shape due to temperature fluctuations, and minimizing the size of the plastic pipette holder produced improved stability in my case.

I tried many things to maximize stability of my patch-clamp setup, including wrapping the faraday cage with trash bags to minimize air flow and temperature fluctuation through the cage. Although it looks ridiculous, there was a lot of instant improvement, so I continue to operate it this way. Some of the techniques I tried improved stability, some did not, but in every case video recording pipette drift was crucial to critically evaluating (A) how much of a problem there was and (B) which changes to my setup improved the situation.

Note finally that what I thought was mild "drift" was actually very serious random "wiggle", such that hour-long pipette location change was minimal, but minutes-long instability was resulting in poor patches for long durations in a way that I never would have noticed if I didn't video record the movement. I wonder if your probably may be similar... a total difference on final position of one cell body over two hours may be deceptively mild compared to the total movement actually experienced.

Good luck!

https://www.youtube.com/watch?v=JlAYViKUClw&list=UUs3-qUELPY-2AOHa7Q_db5w

https://www.youtube.com/watch?v=_-4f0T2Hifo&list=UUs3-qUELPY-2AOHa7Q_db5w

Thanks everyone for the suggestion! 

I am agreed with the above nice suggestion, mainly the length of the head stage and pipette holder with electrode from the manipulator holder should be lesser and electrode should be tight with 45 degree angle or lesser will reduce the drift.

I will be grateful to Danial  butt if you will send me a good picture and idea that without movement after whole cell how he is titetning the electrode holder at my email [email protected]