Klapperich Lab:Guide to Asylum AFM

Overview

This protocol gives instructions on how to take a simple image with the MFP-3D Asylum AFM.

Materials

• MFP-3D
• Appropriate tip (e.g. AC240TS)

Procedure

1. Reserve the Asylum AFM at Yahoo Calendars
   • User Name: BU_Asylum
   • Password: Research
2. Press the POWER button on the front of the computer. Press the POWER button on each of the monitors.
3. Toggle the controller POWER switch to the ON position.
4. Verify that the laser safety lock is plugged in, and the light source key is in the horizontal (unlocked) position. This can always be left on.
5. Verify the following LED’s are illuminated/powered:
   • Monitor 1 front
   • Monitor 2 front
   • Computer
   • Controller
   • Light Source
   • Turn on the isolation table and after a minute press “E”
6. Double-click the MFP-3D software icon. Minimize the window and manually resize to fill both monitors.
7. Using the MASTER PANEL tabs, for IMAGING MODE choose AC mode.
8. Remove the head from the base and turn the head upside-down so that the spring clip is exposed.
9. Remove the spring clip assembly from the head by pressing the lever holder it in place, and seat it in the cantilever holder.
10. Select a new cantilever and check it with a magnifying glass to make sure it has a tip. (The tips are the responsibility of the user.)
11. Unscrew the back screw (not all the way).
12. Carefully (using tweezers) place a new cantilever under the lip with the tip facing up. Align the cantilever so that the upper two corners touch the edges of the metal surface.
13. Tighten the screw.
14. Center the sample on the stage and use the magnets to hold it in place.
15. Remove the spring clip assembly from the cantilever hold stand, and replace it on the head. Insert the back end first.
16. Roll up the wheels on the stage so that the cantilever doesn’t crash into the sample.
17. Place the head on the stage. Hold up the front end and place the back legs in first.

1. Adjust the legs so that the head tilts slightly upwards, that way you just need to adjust the height in the front.
2. Click the VIDEO button to open the Video window (this button looks like a TV and is in the lower left of the screen).
3. Click on the SOURCE menu to bring up the light.
4. Adjust the camera so that the cantilever is in view using the wheels on the head.
5. Move the laser so that it shines on the cantilever (beware of the ghost images). The laser is over the cantilever when the SUM indicator reads 6 or 7. The Sum is the total amount of light collected by the photodetector in volts. The laser must be on the tip though even if the maximum value of the SUM is off the tip.
6. Align the DEFLECTION. The DEFLECTION measures how well the reflected light is centered on the photodetector in the Z direction. DEFLECTION is measured in volts and is computed as the difference between the light on the top half and bottom half of the photodetector. Adjust the wheels on the head until the DEFLECTION is 0.
7. In the MASTER PANEL, click the Thermal tab to bring it to the front of the window.
8. In the MASTER PANEL, click DO THERMAL. This test collects deflection data and the results are plotted as the deflection amplitude in m/sqrt(Hz) vs. freq (Hz).
9. When the graph doesn’t change much click STOP THERMAL.
10. Find the peak and use the cursor to draw a box around the peak.
11. Click and choose HORIZONTAL EXPAND.
12. Estimate the frequency at which the curve peaks and insert this value in the ZOOM CENTER box on the MASTER PANEL.
13. Click the SHOW FIT box, click on FIT GUESS, and then TRY FIT. This fit will tell you the resonant frequency of the cantilever.
14. In the MASTER PANEL, click the TUNE tab.
15. Take the resonance frequency obtained from the Thermal PSD (Power Spectral Density), and enter 30 kHz higher than that value for the AUTO TUNE HIGH.
16. Take the resonance frequency and enter 30 kHz lower for the AUTO TUNE LOW.
17. Click AUTO TUNE.
18. The AMPLITUDE should go to one and the curve should look like:
19. In the SUM and DEFLECTION window, click SIMPLE ENGAGE.
20. Turn the head thumb wheel counter-clockwise to lower the cantilever to the surface. Continue until you hear an audible beep. The z-voltage should be slightly in the blue region at about 70 V.
21. The deflection may change to a nonzero value. If this is the case adjust the deflection back to zero and repeat the Tuning steps.
22. In the MASTER PANEL, click DO SCAN.
23. When the scan is complete, you may save the results to a desired file or directory. In the MASTER PANEL, click the SAVE IMAGES check box.

Notes

1. List troubleshooting tips here.
2. You can also link to FAQs/tips provided by other sources such as the manufacturer or other websites.
3. Anecdotal observations that might be of use to others can also be posted here.

Please sign your name to your note by adding ('''~~~~''') to the end of your tip.

References

Relevant papers and books

1. Goldbeter A and Koshland DE Jr. An amplified sensitivity arising from covalent modification in biological systems. Proc Natl Acad Sci U S A. 1981 Nov;78(11):6840-4. DOI:10.1073/pnas.78.11.6840 |
2. JACOB F and MONOD J. Genetic regulatory mechanisms in the synthesis of proteins. J Mol Biol. 1961 Jun;3:318-56. DOI:10.1016/s0022-2836(61)80072-7 |
3. ISBN:0879697164

Contact

• Who has experience with this protocol?