How to be an imaging master: Q&A

For the dynamic imaging system, the Sysmex FPIA, the typical measurement time is around five minutes. For the static imaging system, the Morphologi G3, the measurement time depends upon the application and can range from about 15 minutes to over half an hour.

Not every automated imaging technique controls particle orientation. As discussed, an uncontrolled orientation leads to discrepancies in measurements of length or shape of non-circular or asymmetric particles. In the Sysmex FPIA this orientation is controlled by a sheath flow system. For the static imaging technique, the Morphologi G3, particles are dispersed either via dry or wet methods but will assume the lowest energy position. You would not expect a pencil to stand up on its tip if dropped onto a table; it will fall onto its side presenting the largest dimension upwards. Dispersed particles will do the same and will present the largest dimension to the camera resulting in a consistent and repeatable measurement.

We will go into some detail about dispersion in future masterclasses. The static imaging system shown, the Morphologi G3, can be provided with an integrated dry powder dispersion unit. Dry powder dispersion is automated for the Morphologi G3. Particles can also be measured in suspension. For the dynamic imaging system, the Sysmex FPIA, particles are measured in suspension only, but can be measured in water or a number of solvents.

We have done some work with liquid droplets and the main difference is the difficulty of imaging the droplets in the matrix or medium and that of dispersion without affecting the droplets. If contrast is good enough that they can be resolved, then automated imaging can be used for the measurement of droplets.

The size range of the Morphologi G3 is 0.5 microns to a maximum of 1 mm. The size range of the sysmex FPIA is 0.8 to 300 microns

There are a number of ways of dispersing agglomerates. In a dry dispersion you can add more energy, when using a wet dispersion you can add surfactant and use sonication to disperse agglomerates. In a later masterclass we will discuss dispersion options is more detail.

When imaging soils in the past we have used what we call an evaporative dispersion technique where we suspend the soil in a solvent with a small amount of surfactant. We then disperse some of this suspension on a microscope slide and allow the solvent to evaporate, leaving the particles well dispersed on the slide. You may need to play with different solvents, ratios and perhaps sonication to achieve the best results.

This is a good question and the answer depends a lot on your background (mine is imaging), and what you want to know. To track changes in a product or process, a measurement like laser diffraction may work perfectly and will be quicker and easier than going to imaging. However, when dispersing your needles, do they break? Imaging allows you to assess this, and what size range you expect, which can then be compared and possibly be used to improve your laser diffraction methods. If length is critical then perhaps imaging would be the only way to go. So it does depend on what information you need. I see them as complimentary techniques; laser diffraction is great for routine analysis, but imaging allows you to verify your diffraction results and provides extra information which can be the difference in development or troubleshooting.