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How to prepare a sample for AFM-IR

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Introduction

Careful preparations are crucial for getting high-quality data from any sensitive measurement. PiF-IR nanoscale spectroscopy and PiFM nanoscale chemical mapping provide lots of important data. Maximize your results by using PiFM and PiF-IR after you have identified a region of interest, minimized contaminants, and properly fixtured the sample. This guide will explain and address the most common pitfalls that Molecular Vista sees in customer’s samples.

Macro-physical constraints

Because PiFM and PiF-IR are based off an AFM, the physical sample preparation requirements are the same as for any standard AFM. Therefore, the best way to ensure that your sample can be measured is to take some AFM images on the region of interest.

If taking your own AFM measurements is not possible, then there are some guidelines to follow.

  • The region of interest needs to be physically accessible by the probe. AFM cantilevers cannot reach down into holes or valleys. Recessed wells are a common culprit for this issue. At Molecular Vista, there needs to be at least 5 mm of access from one side to get the tip into position [Figure 1].
  • The sample needs to be fixtured securely. Because AFM’s scan the surface with a physical probe, any flex or movement of the sample will make imaging difficult or impossible. See the section of sample fixturing for details.
  • The sample must physically fit in the instrument. Molecular Vista can support samples up to 75 × 75 × 10 mm for normal samples. For samples that need to be imaged in a controlled environment (vacuum, inert gas, etc.) then the sample should be less than 50 × 50 × 5 mm.
AFM cantilever access
Figure 1. The cantilever does not have much clearance around it and so it cannot reach into wells or holes on the surface of a sample. Most importantly, the region of interest needs to have at least 5 mm of access from one side, and there should be a flat spot at least 10 mm2.

Micro-physical constraints

Because AFMs use mechanical detection, there is a limit to the surface roughness that is possible to image.

Molecular Vista has a Z-scanner range of 12 µm. However, some of that range is needed to account for Z-drift and the angle of the sample surface relative to the scanning axes. Therefore, in general samples with 4-6 µm of height variation should be possible to image depending on the specific topography. Samples with 7-9 µm of height variation may be possible, but very challenging.

Rough surface cross-section with AFM tip
Figure 2. If the surface of the sample is too rough, then the AFM tip will have trouble tracking the surface. Keep the peak-to-peak height variation less than 7 µm if possible.

If the sample is too rough, or large scans are desired, then you will have to make the sample flatter. For example, malleable materials can be pressed to make them denser and flatter. Additionally, some materials can be microtomed (or cryo-microtomed!) without smearing to smooth the surface.

Fixturing methods

Most samples are relatively easy to fixture. If the sample is not taller than it is wide, simply use an adhesive that does not outgas to attach it to a metal AFM disc. Five-minute epoxy is preferred, but double-stick adhesive tabs can work as well. Avoid double-stick tape from a roll since that can cause the sample to drift.

For cross sections, tall samples, flakes/powders, and other challenging surfaces, there are some special techniques discussed below.

Tall samples

Samples are most stable when they are not taller than they are wide. However, sometimes this is unavoidable. If the height is less than 5 times the width, it may still be possible to image. However, supporting a sample in a small vice, or cutting it down to improve the aspect ratio is preferred.

Samples cast in resin

Casting resin around a sample can be an effective way to fixture difficult materials or cross sections of thin films. However, the resin itself can cause contamination issues. Even a small layer of resin will make AFM-IR measurements impossible. Therefore, follow these steps for best results.

  1. Choose the right resin. Ask the company or lab you are working with if there is a particular resin that they prefer.
  2. Use a cryo-microtome to prepare the surface. Using a normal microtome can work well is some circumstances. However, in many cases the resin material is smeared across the surface. You will not be able to know if this has happened ahead of time.
  3. Use an ion mill to clean the microtombed surface. Cleaning the surface of a sample after the cutting process with a microtome is the best way to ensure that the sample will be free of any resin contamination. This is recommended regardless of whether the surface was microtomed or cryo-microtomed.

Samples deposited on resin

Powders, flakes, and other small amounts of material can be difficult to prepare. Often the best method is to deposit the flakes on top of some resin.

  1. Use a minimal amount of epoxy to smear a thin layer onto a magnetic AFM puck. Thinner is usually better.
  2. Dust the epoxy with the sample flakes or powder and leave to cure.
  3. Removed excess material with a burst of a clean and inert compressed gas such as N2 or Ar.

For imaging and spectroscopy, the AFM probe will have to land on the surface of the flakes or particles. Depending on the size of the individual pieces, these surfaces may be unstable and therefore require more rigid fixturing such as being cast in resin. Also, porous samples may wick some epoxy, and contaminate the surface if the sample is very thin. That is why it is important to smear the thinnest possible layer that will hold the material securely.

Reference sample materials

It is often helpful to analyze some pure reference materials for comparison to the primary sample. For example, comparing individual constituents to a manufactured composite material. Or, comparing some suspect materials to a defect to aid in identifying the process error.

When supplying Molecular Vista with reference materials, please provide enough for ATR-FTIR spectroscopy to complement the nano-IR measurements. For solids or powders make sure there are a few thousand cubic millimeters of material. For thin films, make sure the film is at least 500 nm thick for ATR-FTIR. Glass, silicon, mica, and sapphire are all good substrates that do not conduct too much.

Sample storage and transport

Contamination is a common issue many customers are unaware of. PiF-IR is sensitive enough to measure contaminants at the single-molecule-level. Therefore, samples that are not packed carefully may produce spurious results. Here are some tips to avoid contamination.

  • Avoid all gelpak containers. These materials outgas and will contaminate the surface of a sample in just a few hours. Instead, choose a sample container with little to no outgassing.
  • Avoid adhesives. Secure the sample mechanically whenever possible. If the sample is attached to a magnetic AFM puck, then use a container with a magnet to hold it. If needed, copper or carbon tape can be used provided the adhesive does not outgas.
  • Check all polymer-based materials used for packing and shipping. Even if a sample is packed in a safe container with inert materials, there could still be contamination during shipping from other stuff in the box. Make sure you seal the sample container well or avoid packing materials that outgas.
Outgassing symbol
Figure 3. Choose packing materials and adhesives carefully. Outgassing can easily deposit contaminants on the surface of your sample.

Conclusion

PiFM and PiF-IR chemical analysis can provide a wealth of information. To make sure you get the most value, it is important to minimize sample preparation issues. Most samples are easy to mount in a matter of minutes. However, paying attention to more difficult cases and taking a little extra time to prepare a sample for both analysis and shipping will go a long way to getting the best results in the first attempt.

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