During our recent Porto AFM Training Workshop, we held a little competition, asking the students to produce and process the best images they could.


This year, the winner of the competition was Magdalena Scheibe, who produced this beautiful image of human hair. Well done, Magdalena!

 

Magdalena's winning image
Magdalena's winning image

However, we must also acknowledge two very inventive submissions, firstly this Game of Thrones-inspired one from Izabela (scanned with Anita), which actually shows a "wall" of nanoparticles, and nicely processed in SPIP.

 

Game of Thrones Nanofans
Game of Thrones Nanofans - Winter Is Coming

And finally, this "Moonlanding" on another hair surface from Sandra and Rita. Very nice!

Ana Rita and SandraExploring Hair at the Nanoscale with AFM


Thanks to everyone who entered the competition!

 

All images Copyright AFMHelp.com and their respective creators.

 

The Porto AFM Workshop 2019 has been announced. This is the sixth edition of the course!

 

The course will run from the 15th to 18th April. This is a training workshop, aimed at any researcher or scientist, who wants to learn about AFM, or increase their knowledge of the technique. Following the successful courses that have run since 2011, the course will includes several hours hands-on training in acquiring images with the atomic force microscope as well as AFM data processing.

 

NEWS:

  • The course is now full, and no more registrations can be taken. Contact me at This email address is being protected from spambots. You need JavaScript enabled to view it. if you want to me notified about the next course.

  • The timetable of the course is here. Note the room change, all lectures are to be given in room 0.36, in the library of the maths department.

 

Please click the image below to download the flyer with more details.

link to .pdf of flyer

 

  UPDATES:

  • More details of the course will be announced soon, meanwhile, as usual, pre-registration can be made by emailing me at This email address is being protected from spambots. You need JavaScript enabled to view it.Places on the course are very limited, so interested students should make enquiries or register as soon as possible.

  • There will be some guest scientists, talking about advanced applications in different areas. More details will be announced soon

 

 A blog with information and student feedback from the previous courses can be seen here:2017 course, 2014 course2013 course2011 course.


The course is supported by AFMWorkshop, The Faculty of Sciences of The University of Porto and my research institution, LAQV/Requimte. 

In this article, I’m going to talk about what not to do in AFM. I’ll list 4 mistakes that are common in AFM use, which if you avoid, will certainly improve your results!

 

  1. Using standard settings.This is possibly the worst mistake you can make. AFM imaging is a highly adaptable technique. It’s able to image very large samples of tens of microns with extremely rough topography, or make tiny images of extremely smooth samples with subnanometer features. There is no standard setting which will get good images of all samples. You need to be able to adapt the imaging settings based on the response from the instrument. Optimal AFM imaging is attained through an iterative process! If you don’t know how to optimise imaging of the AFM, I highly recommend revising chapter 4 of my book, “Atomic Force Microscopy”.

   

  1. Interpreting image artifacts as image features. It’s important to know which features in your image come from your sample, and which are image artifacts. Learning this can save you some major embarrassment, and a lot of time!

 

bad feedback imageExample of image showing imaging artifacts. being able to spot and correct these artifacts is a crucial skill for an AFM operator.
  1. Trying to image dirty or contaminated samples. Sample preparation is the first, and most important part of an AFM experiment. If your sample has a layer of contaminant covering the the part you are interested in imaging, it will make your job almost impossible. Prepare your sample so that it only contains things you want to image.

  

  1. “Optimising” your imaging for a nice-looking amplitude (or deflection) image. I was amazed to find people do this. Amplitude and deflection images are made up of the error signal in AFM. The less contrast here is in the error signal, the more accurate your height image is. So, if you optimise imaging to produce a nice-looking high-contrast image in amplitude, you are decreasing the accuracy of your height image!

 

Avoid these pitfalls and your AFM work should be hassle-free! For help avoiding them I recommend reading my book, “Atomic Force Microscopy”, soon to be released in paperback!

 


All text and images copyright 2018 Peter Eaton, AFMHelp.com

In my job as a researcher using AFM, I often act as a scientific reviewer for papers describing AFM experiments. I also collaborate with many groups and spend a lot of time revising and correcting articles which contain AFM data. In this two part article, I’ll describe how you should go about preparing a scientific paper which contains AFM data.

NOTE: Some of this content is not really specific to AFM articles, but could be applied to any experimental scientific report. These parts should already already be known to you if you are writing an scientific article, but in the real world, many researchers are never taught how to correctly prepare a scientific article before they start writing them. For more general guides on preparing scientific articles, check here.

 

What are the overall requirements for an AFM paper?

  • Experimental conditions should be well described

  • Data should be presented in a useful format

  • Reproducibility should be addressed

  • Conclusions should be justified

 

These requirements may seem like “common sense”, but you might be surprised by the number of articles which are published but do not meet some, or even all of these criteria!

 

  • Experimental conditions:


    What should this include:

Imaging mode used: This is rather important, since the results can be different from mode to mode. It also helps to make your data reproducible. Note that not all instruments use the same terms to refer to specific modes :This is particularly the case for tapping mode. It can be useful to use both the manufacturer’s name, and the most commonly used name for example: Imaging was carried out in AC-AFM (Tapping) mode.”

Instrument used. You should name the instrument mode and the manufacturer. You can also include the version of the instrument model, if applicable. Some journals still request the location of the company, but this is becoming optional e.g., you might write “Bruker Multimode 8 (Bruker, Newhaven, CA)”.

Probe used. The type of probe used can determine the type of information available, as well as the interaction force. Since there are many probes available on the market, it’s also useful to say the (approximate) frequency of the probe, which gives some idea of the type of probe, without he use looking up the details. E.g. you might say “AppNano ACT probes with around 300 kHz frequency were used”.

Software used. All AFM images require image processing, or analysis before presentation. It is a good idea to describe the software used for processing of the AFM data. You might also describe the processes used for analysis and /or data processing. E.g. you might say “Gwyddion 2.49 software was used for processing the images, and Roughness was calculated with the Gwyddion 'Statistical Parameters' tool".


Amount of data collected, and statistical significance. This issue is addressed below. However, it's worth remembering, that the microscopy images you collect should be representative of the sample(s). A part of the “materials and methods” section should be explaining this. For example, you might write “At least six areas per sample were imaged, and representative images are shown here.” If you have a variety of different images, you could add additional images to a “supporting information” section.

  • Presenting data in a useful format

There are many different types of AFM images, and different ways to display them. Very often the instrument will produce several different image channels, such as height, phase, etc. The most commonly used channel is probably height, since this is the only one with three-dimensional data. However, sometimes, it’s hard to make out the sample details in height images. For this reason, other channels, such as deflection or phase are often used. I recommend that the height image, always come accompanied with a z scale (normally it’s a colour gradient), and they should have either an inset scale bar, distance markers on the outside, or the legend must state the scan size (x and y). Examples are shown below.

 

Different ways of showing scales. Left: a figure with distance marked on the figure frame. Middle: an image with an inset scale bar. Note how the length of the scale bar was chosen to highlight the repeat distance of the grid. Right: No lateral or vertical scale. in this case, the lateral scale should be described in the legend.

 

Does the colour I pick make any difference? No. You can choose which colours you like. For some reason, AFM images are “traditionally” shown in some shade of brown, but any colour scale is fine.

Should Amplitude/Deflection /Phase images have a z scale? No. The z scale on these channels is usually meaningless, as it might be in volts, or degrees, and in most conditions, is not needed. I usually remove it to avoid confusion.

Are the details you want to highlight easily visible? If not, the use of arrows to denote specific features in the image is recommended. The arrows should be referenced in the figure legend.

Talking of which, in part 2 of this article, I’ll talk about how a figure legend should be written, and how you should address reproducibility of the data, and draw appropriate conclusions from your results.


All images copyright peter Eaton 2019. Images produced with SPIP and Gwyddion software

This instrument is supervised by: Peter Eaton; Please contact Peter for any enquiries regarding access to the lab, or training or use of the instruments. Email: This email address is being protected from spambots. You need JavaScript enabled to view it.

TTAFM at porto

 

Instrument #1: Long Beach

This instrument is a modified TT-AFM from AFM Workshop.

It is equipped with two scanners, enabling large (low resolution) or small (high resolution) sample imaging. It has an standard liquid cell for in-situ measurements, and a custom-made liquid heater cell (ambient to 60 Celsius), with 0.1 degree accuracy. It has been upgraded with two new ADC cards, so it's now capable of recording 12 channels simultaneously, and displaying a life feed of height, scanner voltage, amplitude, etc (see left image above). It's installed in a custom-made vibration and acoustic isolation cabinet.

Here are more detailed specifications:

  • Instrument Configuration: Optical lever - based sample-scanning AFM
  • Sample Size: Maximum ca. 20x20x5 mm
  • Imaging Modes: vibrating (tapping), non-vibrating (contact), phase imaging, lateral force microscopy (friction force microscopy)
  • Imaging Environment: Air or Liquid.
  • Imaging temperature ambient to 60 ºC
  • Z-translation: Vertical direct drive (1 micron resolution)
  • XY Translation: Manual
  • Video Optical Microscope: Zoom to 400X, 3 micron resolution (3M pixel camera)
  • Scan Range: 50x50x17 microns or 20x20x7 microns
  • Linearisation: All axes (x, y and z) with strain guages, can be turned off for enhanced signal to noise ratio
  • Z noise level: less than 40 picometers
  • Vibration isolation: Passive vibration isolation, with acoustic shielding
  • Image aquisition: can collect up to 12 channels simultaneously

 

Instrument #2: San Diego

This is a Life Sciences AFM (model B) from AFM Workshop. It is a probe-scanning AFM with maximum 50 micron scan range. it is mounted on an inverted optical microscope, equipped for brightfield, phase, and fluorescence microscopy. We have both air and liquid scanning-capable probe holders. It is typically used for cell studies and other transparent samples. UPDATE: This instrument currently not available.

Instrument #3: Ontario

Recently installed, this instrument is very similar to the Longbeach instrument, it is a little newer and has the latest (and experimental) software installed.

  • Instrument Configuration: Optical lever - based sample-scanning AFM
  • Sample Size: Maximum ca. 20x20x5 mm
  • Imaging Modes: vibrating (tapping), non-vibrating (contact), phase imaging, lateral force microscopy (friction force microscopy)
  • Imaging Environment: Air only.
  • Imaging temperature: room temperature
  • Z-translation: Vertical direct drive (1 micron resolution)
  • XY Translation: Manual
  • Video Optical Microscope: Zoom to 400X, 3 micron resolution (3M pixel camera)
  • Scan Range: 50x50x17 microns. An optional smaller scanner can be fitted
  • Linearisation: All axes (x, y and z) with strain guages, can be turned off for enhanced signal to noise ratio
  • Z noise level: less than 50 picometers
  • Vibration isolation: Passive vibration isolation, with acoustic shielding



Booking Schedules and Calendars:

Click the link below to see the Longbeach Google calendar

TT-AFM - Longbeach

 

 Or click the link below to see the calendar for "Ontario"

TT-AFM - Ontario


Example Images
Here are just a few example images measured with the TT-AFM (Longbeach) instrument on a variety of samples.

More images can be found in the LS-AFM image gallery and the TT-AFM image gallery.

many bacteria amplitude e. coli
Grid spores in phase cells and spore
leishmania cell epithelial cell  



TT-AFM Protocols



Troubleshooting

There is a Google Docs file I have created which contains the solutions to some problems that might crop up using the TT-AFM. This could also apply to other TT-AFM instruments. You can reach it (for viewing) via this link: TT-AFM troubleshooting. If you have something to add to the document, and wish to contribute, just email me (This email address is being protected from spambots. You need JavaScript enabled to view it.) for access.

Back to Requimte Page

 

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