Hand microtome -Take 2

Here you can discuss sample and specimen preparation issues.
Post Reply
Message
Author
TonyT
Posts: 184
Joined: Fri Sep 11, 2020 8:30 pm
Location: New Brunswick, CANADA

Hand microtome -Take 2

#1 Post by TonyT » Mon Mar 22, 2021 12:46 am

Cross section of a parsley leaf petiole using OLFA LBB blade and blade holder from MicroBob.
First attempt was using a petiole in wax, got a nice thin section. Stained and dehydrated and then cleared in Euparal Essence which severely shrunk and distorted the section rendering it useless.

This 2nd attempt was placing the petiole from alcohol in a foam support (essentially a foam carrot without the carrot). Section was a little thicker than the 1st section.
Mounted in very weak PVA/glycerine (non-permanent).
Width: 3.48 mm
Attachments
21iii21 parsley 768.jpg
21iii21 parsley 768.jpg (221.88 KiB) Viewed 2750 times
New Brunswick
Canada

Greg Howald
Posts: 1185
Joined: Tue Oct 20, 2020 6:44 am

Re: Hand microtome -Take 2

#2 Post by Greg Howald » Mon Mar 22, 2021 10:23 pm

Outstanding! Can you also include a picture of your accomplished microtome?
Greg

TonyT
Posts: 184
Joined: Fri Sep 11, 2020 8:30 pm
Location: New Brunswick, CANADA

Re: Hand microtome -Take 2

#3 Post by TonyT » Tue Mar 23, 2021 2:06 pm

Thanks Greg.
The microtome is a very basic piece from India
To make a bench-top mount I cut a 1" diameter hole in a piece of pine board, 1" is the diam. of the 'barrel'
Because the control knob is wider than 1" I cut the pine board across the diam. of the hole
Placed the barrel of the microtome in position and rejoined the 2 halves of the hole and locked them in place with side pieces.
Top image: clamped to bench, as used
Bottom: side view
Attachments
microtome.jpg
microtome.jpg (168.76 KiB) Viewed 2659 times
microtome-2.jpg
microtome-2.jpg (147.72 KiB) Viewed 2659 times
New Brunswick
Canada

Greg Howald
Posts: 1185
Joined: Tue Oct 20, 2020 6:44 am

Re: Hand microtome -Take 2

#4 Post by Greg Howald » Tue Mar 23, 2021 2:40 pm

Well done. Thanks. Wish you continued success.
Greg

MicroBob
Posts: 3154
Joined: Sun Dec 25, 2016 9:11 am
Location: Northern Germany

Re: Hand microtome -Take 2

#5 Post by MicroBob » Tue Mar 23, 2021 4:57 pm

Hi Tony,
nice section! The parsley leaf petiole really is a rewarding object with interesting structures. It is not unlikely that I have a paraffin block with one which would come handy for tests on a new stain. It also is nice to see that the blade holder is in use. I think that many of the blade holders I sold are not used on single time and on't like the thought.
In your first attempt - did you infiltrate the object in way or just fit it into the microtome with the wax?
For your microtome it might be useful to make a set of cylindrical spacers to fill the cylinder to a degree that the blocking in of the object is easier.

Bob

TonyT
Posts: 184
Joined: Fri Sep 11, 2020 8:30 pm
Location: New Brunswick, CANADA

Re: Hand microtome -Take 2

#6 Post by TonyT » Tue Mar 23, 2021 9:25 pm

for the very 1st one I made a wax block with the petiole inside. I trimmed the to be a little wider in diameter than the petiole.
Mounted it vertically in the cylinder of the microtome and filled the remaining space with liquid wax. I had already coated the inside of the cylinder with a thin layer of beeswax, figured it would stop the paraffin wax from sticking. Got a very nice thin slice using your blade holder. The Euparal mountant destroyed it.

For the 2nd sample the plug of foam was a poor choice as the foam did not cut smoothly but tended to drag. Did get a reasonably decent section - the one posted.

I had another section of parsley stem in wax. I held this in the microtome cylinder with the petiole sticking up above the foam and slightly above the top of the microtome. Used your plastic cutting accessory to anchor the petiole in one of the half-circles.
Gave some very nice thin sections. Dewaxing these thin sections was a pain. I will try again using a stem direct from alcohol. Using the cutting accessory seems to me to be the best way to proceed.
New Brunswick
Canada

hans
Posts: 986
Joined: Thu May 28, 2020 11:10 pm
Location: Southern California

Re: Hand microtome -Take 2

#7 Post by hans » Wed Mar 24, 2021 3:06 am

Nice setup and results.
TonyT wrote:
Tue Mar 23, 2021 9:25 pm
For the 2nd sample the plug of foam was a poor choice as the foam did not cut smoothly but tended to drag. Did get a reasonably decent section - the one posted.
What type of foam did you try?

MicroBob
Posts: 3154
Joined: Sun Dec 25, 2016 9:11 am
Location: Northern Germany

Re: Hand microtome -Take 2

#8 Post by MicroBob » Wed Mar 24, 2021 7:23 am

Foam dulls blades quicker than carrots do. The least damage comes from XPS foam, nor EPS foam.
Usually paraffin sections are moved to a water wath (about 45°C) to expand and picked up from there with a slide, often coated with a fine layer of egg white-glycerin mix. The slide is than dried for a day and heated to just below melting temperature to activate the sticky layer. The slide is then stored or processed immediately through 2x xylene, 2x isopropanol, alcohol ladder down, and then stained. I have made 20µ sections from PEG embedded material with cylinder microtome and blade holder, with paraffin it would work even better. For marking the slides a diamond pen is useful and inexpensive from china.

Post Reply