I first learned electron microscopy from Allen W. Clark (Anatomy, University of Wisconsin - Madison), and we published a research note (Drosophila Information Service vol 50, pp. 105-106, 1973). Starting in 1981, I had a collaboration with Stanley D. Carlson (Entomology, University of Wisconsin - Madison) which allowed me to establish my own program of ultrastructure research.

In the 1980's, I had a grant-funded technician, Randy Sapp, working in my lab. He won a staff recognition award for his high class electron microscopy. Eventually, he went to UMSL Optometry school and became a practicing optometrist in the Kansas City area..

Personal reflections are in my memoirs

Figure - Put the heads in the uncured resin in the right orientation then cure to get the blocks

Figure - Trim the blocks

*Figure - Here is one of about 8 trays of vials of the blocks I made during the EM part of my career

Figure - Put the block in the chuck of the ultramicrotome and first cut one micron sections for light microscopy

Figure - Put circles on one side of a cover slip, and locate sections within circles on the other side of the cover slip. Then the desired plane of section can be found in a phase contrast microscope before the slide is finally made using mounting medium.

Figure - Here's what an unstained section looks like in the phase contrast microscope

Figure - Then cut thin sections on a diamond knife and pick up the ribon of sections on a grid.

Figure - Here are grids ready to use

Figure - A coated slot grid (viewed in the SEM, top, and viewed in a light microscope, bottom) shows what a ribon of sections of Drosophila heads (with compound eyes and first order synaptic area) looks like

*Figure - Grids are stored in grid boxes, and this drawer cotains the grids I made during the EM part of my career

Figure - Put the grid in the scope to view and photograph

Figure - Photographic negatives are obtained

*Figure - This drawer plus another like it has most of the negatives I made during the EM part of my career

Figure - Contact print allows inventory (grid box, grid number, and negative number), and a grease pencil mark tells the dark room technician selected fields to enlarge.

*Figure - These notebooks contain the most of the contact prints I made during the EM part of my career

Figure - Prints were made with an enlarger in a dark room

Figure - Photographs are selected with ideal, matching contrast and brightness. They are trimmed, mounted and lettered to make the plate for journal submission.

Freeze fracture

Figure - Although I did not produce this work, I was coauthor of this important contribution (Harris et al., Nature 1977, 266, 648-650) showing the importance of the freeze fracture technique, in this case showing how vitamin A deprivation decreases membrane opsin concentration

Figure - The thermos is filled with liquid nitrogen which freezes the liquid freon put into that little cup at the top of the thermos

Figure - Rapid freezing was necessary, and, although liquid nitrogen in the thermos is very cold, it turns to air around the sample and does not freeze it quickly. The freon in the little cup is solid, and you melt a puddle in it with forceps and that does quickly freeze the sample

Figure - I made freeze fracture replicas with this apparatus. Specimen is prepared, frozen to liquid nitrogen temperature, put inside a vacuum, smashed with a razor (membranes break down the middle between the fatty acid tails), blasted from an angle with a platinum gun (to shadow protein with electron dense heavy metal), blasted from above with a carbon gun (to hold replica together), then the tissue is dissolved away.

Figure - A piece of filter paper well placed in the apparatus can show the quality of the platinum, carbon, and platinum plus carbon coatings

*I cannot part with these formidable vestiges of the EM component of my career (yet, summer, 2016, age 69, not far from retirement. However, eventually, these are of no further use to anybody, including myself

this page was last updated on Nov. 29, 2016

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