The TT-AFM is a low cost AFM that can produce reasonably high resolution images. However, the "default settings" require some tweaking in order to get high resolution images.
SCANNING SETTINGS FOR HIGH RESOLUTION
The main issue that needs to be controlled, is the use of closed or open loop operation. Closed loop operation favours highly linear scanning and measurements that can be extremely accurate, assuming calibration is done correctly. However, the noise in the sensors is too high for high resolution scanning. So, you can revert to open-loop scanning when high resolution / low noise scanning is required. In this mode, the linearity is lower, but the noise is greatly reduced, allowing for higher resolution scans. Note that this kind of compromise is required on many AFM models, not just the TT-AFM. On the other hand, some modern instruments feature position sensors with very low noise. Essentially, I recommend two sets of settings to use, a "low resolution/high noise/large scan range/high linearity" set and a ""High resolution/low noise/small scan range/low linearity" set. The difference between these settings are numbers to set in the "System" tab.
The default settings are:
|Z HV Gain||15|
|XY HV Gain||15|
|X Gain %||100|
|Y Gain %||100|
(probably your instrument was originally configured using these settings)
To get a dramtic reduction in noise, and increase in z and xy resolution use:
|Z HV Gain||5|
|XY HV Gain||5|
|X Gain %||0|
|Y Gain %||0|
1- IMPORTANT! Be careful when adjusting these settings, double check that you do not enter some crazy values. Assume that these fields have no "error check" routine. If you use the wrong values you may damage your instrument!
2 – IMPORTANT! I am assuming you are using v2.03 of the software, if not, the procedure will probably be different. At the time of writing v2.03 seems to be the best version.
3- These values are, I believe, saved in the scanner.cfg file, so you could set up two files, with the two sets of values to be more convenient, and avoid any errors in typing the values.
4- Reducing XY HV Gain reduces the scan size, so a Range check is ABSOLUTELY necessary. Even if it does not look like it's necessary, in general I would recommend re-doing range check, after changing any of the XY parameters.
5- The difference compared to the images you sent, should be a reduction in XY jitter at the edges of your features (tubes). Essentially, the Gain(%) values control the feedback from the XY position sensors. The signal they give is very noisy compared to the precision of the piezos, so by setting this to 0, you turn off the sensors, and remove this noise.
6-Because of point 5, the "high resolution" mode is NOT LINEAR. This may mean some XY distortion in your images, and inaccurate zooming.
7- When you reduce XY HV Gain, you reduce the range in XY. So these values above gave you reduce range by 2/3, and you end up with a range of around 16.6 um. If you want to do large scans, you could turn off the sensors, but leave HV gains alone, so you would use:
|low noise/large range|
|Z HV Gain||15|
|XY HV Gain||15|
|X Gain %||0|
|Y Gain %||0|
8- As a side note, if you ever used another AFM, this may seem a bit fiddly, but in fact all of these settings are used in other AFM systems as well. The difference with the TT-AFM is it is manual, not done automatically. Yet. Of course, this is one reason why the instrument works out at such a low price!
TIP APPROACH FOR HIGH RESOLUTION
If you use the settings above for the first time, you may notice a drastic improvement in noise level. Then you may well start to see other issues become apparent. For one, you may be able to see whether your vibration/acoustic noise isolation system really works or not! Another issue is probe tip size.
In all AFMs, the achieved resolution is controlled by the size of the probe tip WHILE SCANNING. That means, not the size of the tip in the box, but on the sample. What happens between the probe being nice and sharp in your box, and scanning your sample? Tip Approach! Good tip approach is crucial for high resolution scanning. Below I include two tips to improve the chance of beginning your san with a sharp probe:
1. Use conservative tip approach parameters. By conservative, I mean a slow, well-controlled tip approach, which does not result in a low set point. By default, your system probably came with tip approach parameters that guarantee a successful approach every time, but also pretty much guarantee you will not have a sharp probe after approach. Again, I am assuming you are using V2.03 of the software. The tip approach routine is completely different in some other versions of the software. Look at the collection of controls in the System Tab, labelled “Tip approach parameters”, highlighted below.
Note that these parameters are considered “Advanced Controls” by the AFM Workshop Software. They may therefore be hidden. There is a grey button on the Pre-Scan tab (location highlighted below) that makes these control appear and disappear.
Incidentally, just above here, is a control that swaps between “Update Setpoint” and “Fixed Setpoint” Ensure this is set to “Update Setpoint” EVERY TIME you approach.
There are two things I recommend you can change in the tip approach parameters to enable a conservative approach. Firstly, and most importantly, you should adjust the value labelled “extend factor” to reflect the setpoint upon approach. This is a fraction of the free amplitude, which is used to detect approach. Lower values will lead to a lower setpoint (vibrating mode). So, lower values will be less prone to false feedback, but more prone to blunting your probe. I recommend you experiment with values between 0.91 and 0.94.
2. There is a bug in the software which might cause rather rapid translation when the scan region moves one place to another. Essentially, after range check, if you request a scan in the centre of the scan range, the probe will move very quickly to your requested spot, which could lead to probe damage. To avoid this (especially when you have a new probe, and want high resolution). Before any tip approach, set the scan center and scan range to your desired initial values, and begin a scan BEFORE approach. Allow the software to collect one scan line (it should be empty), then stop the scan. Now approach, and start scanning. In this way, translation to the initial scan position is minimised.
As always, please contact me with any comments on this article. All content copyright 2013 Peter Eaton.
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I recently received some praise for my book, and I thought I'd share some reviews with you. Alexander Kraft, from Wacker Chemie AG; said the book represents " A good possibility for beginners in AFM to understand the basics and to gain a deeper insight in this measurement method." Thanks, Alexander! Here are some more quotes:
"A super-clear, easy-to-read, informative, and intuitive introduction to AFM, the best I have found. Normally, I find that books like this can be a bit dense and/or skip over details of how things work, but this book builds everything up intuitively and with such clarity it'd probably be able to be understood by a freshman college student--but, without sacrificing the necessary detail."
- Reviewer at Amazon.com
"Atomic Force Microscopy provides the basic knowledge necessary for successful AFM operation while avoiding the trap of providing more detail than beginners can handle. It boasts seven chapters, each of them accessible and self-contained; readers can thus cherry-pick the topics of relevance for their specific problems. After a short introduction about the historical background and the contemporary context, the book covers practical issues such as understanding AFM design; working in operational modes; measuring, processing, and analyzing AFM images; and spotting and avoiding artifacts. For readers inclined to explore further uses, the book's last chapter discusses various applications that illustrate the multitude of measurement options available with AFMs...Atomic Force Microscopy is a great introduction to AFMs for beginners and, although light on theory, also serves as a good starting point for more serious users."
- Udo D. Schartz, in Physics Today
"I recommend this book to any reader who wants to enter the world of force microscopy. This book is easy to read, entertaining, with a practical approach that allows, after their reading, have a realistic idea and practice of this technique. This book touches on all the points and issues that are critical to understanding the proximity microscopy.
These include instrumentation, measurement modes, familiarization with the images, the routine procedures for image processing, one section devoted to artefacts and finally potential applications of the technique. From my point of view, is one of the books on microscopy of proximity, which is easier to read and with a high applicability in measuring routines."
- Carmen Serra, Nanotechnology and Surface Analysis Service, University of Vigo
Finally, a reminder: in April 2014, We will be giving another of our successful AFM training course. At the time of writing there is ONE places left on the course. I recommend anyone interested to sign up as soon as possible. Click the image below for more information.
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Up until now, this website has been funded by Google text adverts that appear on the right of the page. I don’t control what ads appear there, except that I can remove some categories of ads. Beginning soon, I expect to have actual ads from AFM companies appearing there.
The advertisers do not influence any of the other text I write here. Although I do work with some AFM manufacturer’s equipment more than others (and my co-author on the book, Paul West has been owner/CEO of various AFM companies), I do not really favour instruments of one manufacturer over others. I have used many (more than ten) different AFM instruments over the years and this has led me to think that -
ALL AFM instruments can produce great results.
What is necessary to get great results are a certain level of skill on the part of the operator, a good probe, careful sample preparation, patience, use of the right modes and settings, and sometimes, a dash of luck! While newer instruments certainly offer amazing new modes, and in some cases lower noise levels, increased ease of use, or faster scanning, in my experience 99% of AFM could actually be done on just about any instrument. In my teaching, I hope I explain things that are useful to users of all instruments. Furthermore, although I am happy to get new listings, and factual corrections for the “Where to buy instruments” and “Probes” and “Calibration artifacts” pages, I do not accept copy written by the companies for inclusion on those pages. Any inaccuracies, or opinions are mine alone.
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Our 2014 AFM course has had all the places reserved. I am pleased to see we have 16 students from all over the world, including the USA; Malaysia, Germay, Spain, the Czech Republic, Poland, and here in Portugal.
Meanwhile, I will be teaching on another upcoming course, in July 2014, at Kent State University, in Ohio. This course will be 5 days, with full 5 afternoons of instrument time. Places are very limited. More details can be found here: http://afmworkshop.com/atomic-force-microscope-workshop.php. The AFMWorkshop website also hosts a PDF flyer.
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This article contains a small extract from Chapter 7 of “Atomic Force Microscopy”. Chapter 7 contains descriptions of applications of AFM in materials science, chemistry and physics, biology and the life sciences, nanotechnology, and in industry. This short section describes some examples of applications of AFM in bacteriology. References lists, and the second figure can be found in the full book.
AFM is a highly suitable tool to examine bacteria, and has been widely applied to their study. Bacteria are commonly studied by optical microscopy, which can give an overall idea about gross cell morphology (via a two-dimensional projection), and is also useful for cell-counting studies. In comparison, AFM is slower, and thus is less useful for quantitative cell-counting, but allows measurement of a variety of other cellular properties, particularly by nanoindentation and force spectroscopy experiments . In addition, the greatly increased resolution of AFM allows for the imaging of finer details of cell morphology and sub-cellular features such as pili and fimbriae . The three dimensional information from AFM can also be useful in differentiating morphologies which would look the same in optical microscopy . Various other micro-organisms have been studied by AFM such as spores [178, 613–615], fungi [616, 617], including yeasts [171, 618], viruses [287, 619], and others  but here we concentrate on bacteria for the sake of brevity.
Fig. 7.20. Studies of bacterial morphology. Top left: Streptococcus, showing typical linear clusters. Top right: large clusters of Staphylococcus aureus. Bottom left: Salmonella biofilm showing pili-like fimbrial structures. Bottom right: E. coli. All these images were measured in air. Reproduced with permission from  (top left) and  (bottom left). Right hand images the author's own work.
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