Broom sections in different stains

[MicroBob]

Hi toghether,
here a couple of photos resulting from experiments to re-invent an old black stain for botanic sections.
The original was said to specifically stain unlignified cell walls. The new stain does this but more experimentation is needed to find good combination stains. One unexpected problem is that black or grey with a tint looks unclean where a colour would just change the tone a bit.

These sections are made with a cylinder microtome and a blade holder, 40µ thickness.

1. Safranine, differentiated, black stain
2. Only Safranine, differentiated
3. Wacker W3A simultaneous stain for comparison

Bob

[mazo4033]

Beautiful pictures and stain colors! Did you make all of these sections with the microtome you 3D printed?

[MicroBob]

The sections were cut with a nice japanese table microtome, like a hand microtome in a little stand. It is a bit more practical than my hand microtome with desk holder but largely the same. for sectioning I used the 3D-printed blade holder I designed. It offers a very steady support for the blade and makes it easy to do slicing cuts. And since the blade touches nothing but the specimen and the carrot support it stays sharp for a long time. With this blade I have cut a couple of hundered sections before and just pull it over a pice of plain leather a few times before every re-use. In some cases I use a 3D-printed cutting aid too. This can add support directly under the plane of the cut.

My next plant will be ivy as it is available for everyone and we are going to compare staining results with a couple of people.

Bob

[mrsonchus]

Lovely demonstration of technique Bob. Very interesting indeed to be able to see the tissue-types so easily. Nicely-done!

p.s. look out for the sun/shade forms of ivy leaf, may have some interesting comparative morphologies...

[MicroBob]

Hi John,
my biology knowledge is average at best and these differentiating stains give me at least a little guidance towards what is what. I made some test sections from ivy leaf stems now. The stems didn't cut very well, fresh a bit better than fixed. Problem with the test mixes is that the stained areas can turn brown from the iron content. So the way this two-pot stain is applied has to be optimized further.
BTW: On slides of which species would an englishman write "Sticky bud"? I got a box of original paraffin sections made by Robin Wacker, still in paraffin. He labeled them just for his own purpose and that is what he wrote on some of them. Might be chestnut?

Bob

[mrsonchus]

Hmmm - could be? Section images will enable them to be narrowed-down.
I like the above section a lot - very good contrast and again plenty of tissue-type detail!
Another cracker to my eyes. Keep 'em coming they're very intersting to see.

[MicroBob]

p.s. look out for the sun/shade forms of ivy leaf, may have some interesting comparative morphologies...

Than you for the hint, John!
I read up on this topic and this difference is really interesting.

Bob

[mazo4033]

Bob,

I like the microtome design you've made, I think it is very creative, very precise, and a great use of physics . What brand is the Japanese microtome? Also, do you (or John) ever identify the structures within the sections?

[MicroBob]

Hi,
the microtome is an old Sartorius hand/cylinder microtome of good quality. I only made the holder for it. What I designed was the 3D-printed blade holder that makes sectioning fairly easy and repeatable. Recently I have cut material embedded into PEG to nice even 20µ sections with this setup.
I'm no great biologist and to identify I would lean on previous work posted on the german microscopy forum. You can find information, sometimes with identification of the functional structures, on a lot of different plants there: http://www.mikroskopie-forum.de/index.php?topic=21220.0

I include a picture where I have labeled the structured using tihis nomenclature: http://www.mikroskopie-bonn.de/_downloa ... Gewebe.pdf

In case you are interested in botanic sectioning this website will probably be quite interesting for you.

Bob

[mazo4033]

Bob, this is a great and very helpful link! Thank you so much! And another great picture you've attached there - I like your sections and stains a lot!