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Hi, can anyone please give some advice on the following problems of AFM Catalyst scanning and force measurement with the PFQNM mode?
1. How to remove the debris adhered to the tip in situ during the scanning of adhesive samples like cells, without changing the tip? Change of tips would take too much time, especially we have to calibrate the tip again.
2.How to enhance the tracking of live biological samples, especially if the sample is higher than the tip height? Increasing the setpoint alone will induce too much force which will damage the sample surface.
3. How to ensure that the tip ramps at the specified points during the point and shoot function?
(1) I used to do the point and shoot after the AFM image scan, but the tip didn’t return to the image center right after the image capture was done. From the optical window, it showed that the tip started from the last scanning point and did the point and shoot. So it could be seen that the ramp didn’t follow the exact point location.
(2) So I calibrated the tip position in the mirror canvas after engage on the sample. This time the optical view showed that the tip returned to the image center after the image capture was done, and the ramp followed the specified points. But on the mirror canvas window, the green cross visible on the image didn’t exactly match with the tip location. I guess this may be caused by the offset I input in the scanning page after engage.
So, do I need to calibrate the tip poison each time the tip is engaged, and ensure a zero-offset during the scanning to make sure the tip would follow the mark in the point and shoot function? Is there a convenient way to get around this?
Thank you very much for any help.
Hi Fenny,
Here are my suggestions:
1) The best way is probably to use a flow through, by using for instance the Perfusing Stage Incubator.
2) I remember your cell type. I think that in your case, the best is to use cantilevers having a tip height of 15 um instead of 7 or 10.
3) There might be a problem with your calibration/registration or maybe your scanner needs to have a fesh x,y calibration. Usually when you use MIRO and import and optical image, under the condition that the calibration/registration was properly done, the MIRO Point & Shoot is rather accurate.
Sorry, why are you talking about the Poisson's coefficient? This is not something that has to be calibrated. On your cells, you can always use 0.5 for instance.
Best,
Alex.
Hi, Alex,
Thank you very much for your reply. Here are my comments for your suggestions:
1) What type of flow should be employed in the PSI, the same culture medium used?
2) Actually, I've contacted the Team Nanotec for higher tips, but I am not sure if they are able to make a suitable probe as they claim that they have no experience of biological application.
3) I am afraid there was misspelling.I was wondering if I need to calibrate the tip position each time the tip is engaged, and ensure a zero-offset during the scanning to make sure the tip would follow the mark in the point and shoot function.
Best regards,
Fenny
I hope you are doing well.
1) Yes, you can very well used the culture medium (at about 1 ml/min). And don't forget the CO2 for the gas.
2) I am very surprised because I did it and it worked out. You should insist and give them the clear specs you expect. I agree that if you come up with a question like "can you make me probes so that I can image tissue and cells", they will not know what to do.
3) Sorry but I don't understand this one. Does that require any action from me?