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I am having a bit f an issue selecting a probe for a particular type of same I need to image using QNM. The sample is relatively stiff, so in order to get any mechanical contrast using QNM, I need a stiff probe. The only probes I have gotten to work are Tap525 which have a stiffness of 200 N/m. However, the sample is biological and in order to believe the mechanical data, I need to be able to image and test in fluid. Tap525 probes will not work for fluid imaging. Does anyone have a suggestion for a probe the is stiff enough for the mechanical measurements, but will also image well in fluid?
In addition, because the probes are still, using the Thermal Tuning technique to get a spring constant is inaccurate. Does anyone know a relatively easy way to implement the Sader Method?
Thanks so much!
Joseph Wallace
Hello Joseph,
You are correct that you will need probes this stiff in order to measure very stiff samples, and you are correct in choosing TAP525 for this purpose. I don't see any reason why this shouldn't work in fluid. Could you elaborate more on why it does not work for you in fluid?
In general, there are some parameters that can be adjusted to improve stability in fluid. I would suggest trying imaging with ScanAsyst auto parameter optimization turned off, and engaging with scan size set to "0". Make sure you have a force curve on the surface before beginning to scan. If you are false engaging, you may need to increase peak force engage setpoint. You may be able to improve dynamics by reducing peak force amplitude, applying low pass filter, or by increasing peak force setpoint. If you describe the problem you are having, we can try to provide more specific advice on how to proceed.
As for Sader Method, there are a variety of resources available online to help with this, such as this:
http://www.ampc.ms.unimelb.edu.au/afm/theory.html
There is a nice online calculator to help here, (including link to the Sader Method iPhone App!)
http://www.ampc.ms.unimelb.edu.au/afm/calibration.html
Best regards,
Adam
Hi Joey,
To add to Adam's note, be aware that you can use the thermal tune function in the NanoScope software together with the Sader method calculators linked by Adam to implement the Sader method. You just follow the thermal tune process halfway, collecting the spectrum and then fitting the peak to the simple harmonic oscillator model. This yields the resonance frequency and Q of the cantilever, which are needed for the Sader method. This approach is described in greater detail on page 9 of this app note.
For that short and stiff of a cantilever, however, even the Sader method may not work well. I had trouble with the Sader method using similar length probes in this paper because the Length/width ratio was <3. The nominal values for the Tap525 cantilever are very close (~3.1), so it could be problematic. I don't think anyone knows the exact cutoff on what constitutes "reliable" pertaining to this limitation.
This is a key reason that we direct users to the "relative method" for PeakForce QNM calibration with the stiffer probes. By calibrating relative to a sample with known modulus we can reduce the uncertainty resulting from both spring constant and tip radius calibration.
Regards,
-Ben