Edited by Thomas E. Phillips University of Missouri

Selected postings from the Microscopy Listserver from January 1, 2018 to February 28, 2018. Complete listings and subscription information can be obtained at . Postings may have been edited to conserve space or for clarity.

Specimen Preparation pigments for microtomy

I am wondering if anyone one has any insight on how to prepare pigments or powders to be embedded for microtomy. I have only worked with biological tissue and do not know if materials samples need to be infi ltrated the way tissue samples do. In addition, if so is there a diff erent technique or diff erent embedding resin needed for an infi ltration process? I cannot fi nd any information anywhere! Janine Hernandez Mon Jan 29

Powders and pigments likely do not require encapsulation. It is usually much easier to place them on a supporting grid directly for imaging. T is of course assumes that the size of the powder or pigment is small enough for electron transmission. Oſt en it is worth it just to check in case it is possible; which makes sample prep easy. T ere are a few methods that could be used to prepare these- one that I have had some success with fi ne powders in the past is as follows: place an amount of the powder into isopropanol, ultrasonicate to break up agglomerations, dip a holy-carbon grid into the liquid, pull it out, and allowing the grid to air dry. If there was a small enough amount, but enough to be fi nely dispersed in the alcohol, you will pick up some on the grid. T is grid, when dried, can then be imaged with the powders suspended on the holy-carbon. It may take a few trials to get enough from the alcohol suspension. Another method that has worked for me in the past (assuming the powder is well separated) is a very light dusting of the powder mid-air above the grid (allowing some of the powder to fall down onto the grid on the table). T is one is less likely to obtain separated powders that are small enough to image, but I have seen some success with it as well in the past. Use of a clean fi ne artist paint brush to fl ick the powder into the air above the grid may work here. Please be careful not to allow the alcohol with the powder or the powder in air to get on your skin or to be breathed in. Some solvents (acetone notably) allows chemicals to enter through the skin, and we are learning that nanopowders are not healthy for you to breathe in as well. I hope this is helpful. You will likely get a number of great answers from the list-serve and I’m looking forward to learning from their answers also! Wishing you a grand time exploring small worlds! Allen J. Hall Mon Jan 29

Specimen Preparation vacuum desiccator sample storage

We are looking for a compact storage solution to keep membrane boxes under vacuum and/or inert atmosphere. I am considering the containers off ered by SPI ( spidry-sample-preservers/ ) and I am also aware of Ted Pella’s bell jars, which are a bit too large for our purposes. Does anyone have recommen- dations for other containers we can consider? Steven Spurgeon steven. Mon Feb 12

I have been using this model for storage and transport of SEM samples but also whole TEM grid boxes: sample-stub-vacuum-desiccator.html T ey work really well, up to the point where they start questioning you at the airport security check


where you’d be going with that “crystal ashtray”. Guenter Resch lists@ Tue Feb 13

Specimen Preparation ancient grain

I have been tasked with imaging an ancient grain. It is 1000-year-old millet and I have one only! I have done SEM/TEM of grain before but not one so old. I am thinking SEM may be the way to go. T e investigators would like to see the internal structure of the grain (if any) and I have no idea whether it will be ‘normal’, caramelized or powder inside! It must be fi xed in order to be released from quarantine so my fi rst question is should I use an aqueous fi xative or alcohol? Any other advice would be gratefully accepted! Lisa O’Donovan Fri Feb 16 It is a grain, so largely starch, meaning formaldehyde and glut

will not fi x much anyway. If alcohol is good enough to release from quarantine, use 70 or 80% ethanol, then go through to 100% ethanol. Either dry from ethanol or go to tert-butyl alcohol and vacuum sublimate at 20°C. Aſt er the 2nd or 3rd 100% ethanol, you could put the grain in liquid nitrogen and hit it with a razor blade, maybe gently crush it. You’ll get a brittle fracture that will expose the starch grains. T is part will be particularly fun if your grain is tiny. Phil Oshel Fri Feb 16

Fixation in 95-100% methanol or ethanol may be better for your grain - there will be less tissue swelling. T e dry grain will contain only about 8-10% water, so going into 100% solvent will be fi ne. A little water (i.e. 95% solvent) may help penetration. Methanol will penetrate a little better, but it will take some time for any solvent to get deep into a dry grain. Another option aſt er drying would be high-resolution (± phase contrast) x-ray CT - it would quickly show you if the grain had any contents and give you an idea of what they are without breaking the seed. T ere are at least a couple of labs I know of in Australia that do this, one of them is here in Canberra at the ANU - https://ctlab. , and since your seed will be dead aſt er going through solvent, you don’t have to worry about the high kV x-rays killing the tissue. Millet seed is pretty small, so you’d get good resolution of the innards. Rosemary White Fri Feb 16 I like Rosemary’s suggestion — mine about liquid nitrogen needs decent sized seeds. I’ve used it for corn and barley, but if you do have millet … oof. Phil Oshel Mon Feb 19

Specimen Preparation evaporating tin

I have been asked if our vacuum evaporator will put down tin to 1-micron thickness. We have a Cressington unit so it is not the principle, it is the practice. Does anyone have experience with tin or a thickness of 1 micron or both! Chris Christopher J. Gilpin Mon Feb 19 As the evaporated fi lms get thicker, internal stress becomes signif- icant. I am not sure how bad tin fi lms are but I have had issues with the fi lms peeling when they got much over 0.1 µm thickness. Henk Colijn Mon Feb 19

doi: 10.1017/S1551929518000433 • 2018 May

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