After Dinner microscopy

Sit through enough dinner meetings and you get an idea of what makes for a good after dinner speaker.

There can be not doubt the environment is important. The speaker needs a projector, laser pointer and any other props he or she utilizes. The audience needs to be comfortable, but in my opinion that not a limiting factor. I’ve sat in a kiddy chair with other adults in a dark basement of a museum and listened to world-class speakers. I was entranced by their presentation.

A well-known or famous speaker helps. We assume an Edison or Seabrook would have interesting things to say. Sadly that isn’t always the case. I heard Heisenberg speak on quantum mechanics. I was studying that aspect of chemistry at the time so I wanted to hear one of the founders, one of men who created QM. I understood the words, but the meaning of what he was attempting to expound escaped me. To be fair, I never saw Heisenberg, the lecture room was so crowded that I and a lot of other people had to stand in the hall. I suspect he didn’t use much in the way of illustrations.

Maybe the most important thing a speaker can do is to make the topic relevant to the audience. This is harder than it sounds. It requires a speaker instruct the audience without losing them while connecting to his or her topic. Not all speakers share this ability. Part of the audience’s duty in attending any lecture is to intellectually stretch and make connections between the topic and their interest.

If you want to make a presentation to a microscopy group there are some givens you must attend to. Here’s my top Five:

5 What am I looking at? Label the image.

4 What magnification, acceleration voltage, colored filtered, image enhancement was involved with that image? I want to know so maybe I could make one like it.

3 How did you fix, prep, thin section, stain, dehydrate, squash, dice, ash, grind, or polish the specimen? We really do want to know!

2 We want to see the microscopes and any other cool tools you used to collect the data. We are techies of sorts after all.

And finally,

1 Make us feel we are an active participant in studying the central purpose your lecture is about.

Glass

I was thinking of glass the other day. This is not uncommon for me. I admit to being amazed by it. It may be mankind’s oldest discovery, just after fire but before agriculture.

It’s not hard to imagine primitive man throwing different earths onto her (women were in charge of fire, too import for primitive man.) cooking and campfire. The different elements present in the wood and soil could produce interesting colors. Even now, the campfire is often the evening’s entertainment.

The following morning, the women would carefully probe the ashes looking for hot embers to fan into the morning fire. What do you want to bet they found primitive glass?


I was specifically thinking about glass fibers. I was at a rubber company and they wanted to tell the difference between E, K and U type glass fibers. It did not take much thought to put samples of each into the SEM/EDS and collect spectra.

The results were rather nice. E glass contained calcium, (in addition to silicon). Types K and U did not contain calcium, but different amounts of magnesium and aluminum. It was simple to work up a scheme to identify the glass:

Run SEM/EDS on glass, if Ca is present it is a type E, if not check the Al and Mg ratio. If this ratio is greater than two it is a U type. Less than two, it’s K type.

Of course, this is on untreated glass, no etching, coating or leaching.

I felt pretty good about the whole thing until I remembered not everyone has a SEM/EDS. Besides I was white light microscopist and needed to get back to my roots.

What could be simpler than isotropic material and refractive index. I didn’t even control temperature or use a sodium filter!

A drop of liguid, a few glass fibers, close the condenser down, increase the working distance by focusing up and watch the becke line move..

I soon knew the following:

Type E glass had a refractive index equal to 1.555,
Type K glass had a refractive index equal to 1.526
Type U glass had a refractive index equal to 1.520.

Could I select one refractive index to sort them all?

Sure, mount all the fibers in 1.524 and check the contract and becke line.

Type E will have high contrast and be higher than the mounting media.
Type K will have very low contrast and be higher than the mounting media.
And
Type U will have low contrast and will be lower than the mounting media.

So why, would you be interested in this. I don’t know. It’s just the other day I was discussing Dr. McCrone to some non microscopist and I realized it’s been almost six years since he has passed on. Other than a short note published in the Microscope about Microscopist’s Heaven, reportedly communicated by him, his tremendous output of literature and guidance has been missed.

One of the first experiments I did in his introduction to microscopy classes was to distinguishing three isotropic mineral fragments with one refractive index liquid.

It was nice to revisit those memories.

Scales and Distance

I always suspected counting and therefore mathematics started when Ugha the caveman asserted he had more rocks than cave neighbor, Effua. As long as the question remains how many rocks, homes or TEM grids one has, simple counting suffices. It’s only when we want to measure objects (my rocks are bigger than your rocks) do we run into problems.

The yard is a sub-division of the rod, which was based on the total length of the left feet of 16 God-fearing Englishmen. How do we know they were God-fearing men? Easy, they were the first 16 to run out of church one English Sunday.

I’d like to remind the metric crowd that there are a few meter stories out there too. The first attempt to define a meter was the length equal to one ten-millionth of the distance at sea level from the pole to the equator. Not the easiest to use and they scrapped out this definition. For a long time the meter was the distance between to scratches on a platium bar. TWO SCRATCHES?!? I’m measuring microscopic particles based on two scratches!?!

For a while the meter was defined as the length equal to 1650763.73 wavelengths of the light from the 2p10 and 5d5 electron orbital transition of the krypton 86 atom in a vacuum. This still wasn’t good enough, so the physics community is considering a meter in terms of the speed of light in a vacuum. The value for the speed of light, (299,792,458 meters per second) is the current ruler. We will not go into what is a second.

Ultimately we need to calibrate our measing devices, from stereomicroscopes to STEM if we wish to compare results. So what do microscopists use to calibrate their TEMs? It was pointed out to me that most TEM aren’t calibrated, but the imaging system should be. I remember a TEM being installed 30 years ago in which the magnification read out was calibrated to match the magnification on the film. Prints were made by setting the enlarger to a specific magnification as determined with a negative of a grating replicant. I’m concerned about the calibration of my imaging system because it allows me to do manual measurements.

So what do microscopists use? My informal survey is based on 24 responses:
63% (14 people) use a grating replica,
68% (15 people) use crystal spacing.

This gives me 131%, due to microscopists who use both gratings and lattice spacings at different magnifications. It seems a sensible approach.

Of the crystal spacing gang:
47% Silicon based crystal,
7% MoO3,
7% Graphite,
27% Catalase crystals,
7% Tobacco Mosaic Virus (possibly the most unique!),
7% unspecified.
Again I exceed 100% due to utilizing several different crystals as a function of magnification.

26% of the respondents indicated they use a commercial product called Mag*I*Cal. This is an ion milled silicon crystal orientated in the 111 direction. The spacings it produces are used to calibrate your scope. Only one person or 7% mentioned it or any other product was NIST traceable. It seem to be the only one on the market.

I agree that many standards are “God-traceable” and have well documented constant features in the scientific literature. Their validity in a court of law is another question. Still most of the people who use these type of standards can recognize a 111 from a 011 face. I can’t, so I use a grating replica.

No matter what, every calibration is still dependent on deciding where a specific feature starts and ends. Seems we’re still using two scratches as a standard.

Wrong Abbreviations

I’m not much of a list reader, especially abbreviations. Two-letter state abbreviations or a listing of area codes and geographic locations are not what I consider any kind of literature.

Recently I found myself reading ASTM volume 14.02. I was interested in the list of standards that might have impact on me as a microscopist. Information on sieve size and standard practices for reporting particle sizing are both interesting and useful on a daily basis.

More along the lines of fantasy baseball were the procedures on microchemical testing for illegal drugs or the determination of refractive index of glass. These are techniques I have an introduction to, and some skill with, but no current application.

At the end of each ASTM volume is a listing of symbols, references and conversion factors. This section serves as a mini-style guide for authors and standard users to help make reported results similar in description and style.

I discovered several interesting items:
“Gy” is not Goodyear but a “gray” which is an absorbed dose. A dose of what isn’t clear, but if you worked for a tire company you might have a pretty good idea. It’s clearly rubber fumes.

A “ha” is not a unit of laughter, but area of one hectare. I believe it is a farming term. I quess there’s not much laughter in farming today.

“Wb” is of course a weber. Who would have guessed it is a unit of magnetic flux and not an unit of outdoor grilling surface?

A teaspoon has the approximate conversion factor of 5 ml. Somehow “a teaspoon of sugar helps makes the medicine go down” sounds much nicer than 5 milliliters of sugar. In case you’re wondering a tablespoon is about 15 ml.

The strongest value of the metric system, excluding its almost universal acceptance, is the ability to slide up and down the magnitude scale. One centimeter is 10 millimeters or 10 thousand microns (yes, the units are wrong. I’m old and I like microns rather than micrometers. So sue me!)

On the high end of the continuum I noticed mega (ten to the 6th). I didn’t think this was a real scientific prefix but I was wrong. It’s not just a unit of advertising as in Mega-mart. Giga and tera, old hat for some of us, but peta has nothing to do with ethical treatment of animals but stands for ten to the 15th. At the end of the continuum yotta weighs in at ten to the 24th. Clearly that’s a lotta anything.

Sliding down the scale to the small end is femto (ten to the -15) and atto (ten to the -18). That’s small, really small, but not small enough! Still smaller at ten to the -21 is zepto. I assume it was named after Zeppo Marks, a man who could do a lot with very little. Not wanting to be limited by such a small number, we find yotco, which is the number of laughs this column generates or ten to the -24th.

Glass

Glass

Recently the Material society has announced the top ten Material Moments in
History. Rumor has it that they are moving to create the on-demand video
cast to rival David Letterman Show. Who knows?

Number One is the introduction of the periodic table by Mendeleev. Not a
very gutsy move on their part, but quite possible the best call they could
make.

Of interest to microscopists is Number Four, the Invention of Glass in Iran
in 2200 BC. Current politics as it is, if Iran isn't careful they could
quite possible be turned into glass. Radioactive glass, but still glass.

Number Five is Optical Microscopy developed by Anton van Leeuwenhoek. It
took about 3868 year to go from glass to microscopy.

It's difficult for me to explain the wonder I feel about glass lens. It's
the shape and a ray of light that seems so significant to me. Simply
changing the curves or the type of glass and you'll alter the focus and the
dispersion of light. Moving the lens or adding a second will bring sample
image in or out of focus, sharpen or distort it.

I'm not interested in these off-the-island survivor-type shows, but I saw
the opening show of one that amazed me. One team found a lens in their
first aid kit and was able to make build a fire the first night. It took
the other team two cold uncomfortable days to catch on. Clearly they were
an evolutionary dead end. All it took was a curved piece of glass. No
electricity, no computer to constantly force stability and add correction
factors, no temperamental systems needing a multiplex of people just to
operate. Just a lens.

Microscopy: just you, a little glass and some spare light.

The microscopist of the so-called golden age of microscopy used an
illumination system called Critical Illumination. The mirror and condenser
would be set to focus the illumination source in the plane of the specimen.
The microscopist would sit, preferable by a northern window, and use the
blue northern sky as the illumination source. Every so often a fluffy
white cloud would drift past their rotifers and starch grains surprising
the microscopist.

I have often wondered how many microscopist suddenly realized they were
drawing in a slow moving cloud with their camera lucida in to the
background of a diatom strew? A recent trip to the Cleveland Natural
History Museum gave several MSNO members a chance to use a camera lucida.
The camera allows each eye to focus on different objects and the Mark 1
super computer in our head overlays the images. The microscopist /artist
then "traces" the image seen on a blank piece of paper. It's quite
remarkable.

Why draw? Well the ability to draw an object and incorporate the
significant points while omitting the defects, the damage, the non-typical
structure make for a much more powerful image. The drawing becomes an
archetype which will allow the viewer to recognize it in other examples
which are not quite the same. John Delly, master photomicrographer, always
told me that to draw something is to educated the eye. Drawing crystals,
pollen or any other structure will teach you to recognize all or part of it
in another slide preparation.

The cavemen in southern France understood it, to draw something means to
understand it better.