Mouse Blood

[Cactusdave]

I thought you might like Klaus' website! If you navigate from his home page to 'price guide', at the top of that page you'll see the more modest examples I was thinking of, including the 8 form test plate. He also has stuff on Ebay from time to time, but if it's one of his more complex mounts they go for good prices.

As to what we get up to in the UK, I'll take the question literally. This is the website of the Quekett Microscopical club that several UK forum members belong to. http://www.quekett.org.uk/ We have regular meetings at the Natural History Museum in London as well as field meetings and longer field trips. It's a great pleasure to be able to meet fellow microscope enthusiasts in person and to have a good gossip.

David

[Mitch640]

Mitch: I thought you were mainly interested is testing optics rather than the art.
Moth scales have a lot of useful detail and are free. Heck if you can't find a moth wing I could let you have one from my 10,000+ collection of pinned moths (hence the moniker "Mothman").

I am interested in testing, but after looking at those slides, I had to check out some of his other stuff. This is what I do to pay for my hobby. I search for websites and list them in a directory that pays me to do it. Been doing it for a couple different directories for over 10 years now. I can't resist clicking links. In fact, I recently listed this very forum in the Directory.

http://botw.org/top/Science/Biology/Chats_and_Forums/

Dave, I have seen this website. Did they have a recent field trip that was posted? As far as I know, there is nothing like it here, or at least not near me. But I wasn't really serious.

[Cactusdave]

Unfortunately trip reports are in a 'members only' part of the Quekett website I don't exactly know why.

[rjlittlefield]

Thanks Rik, I stand corrected about the E plan X40, I didn't know it was part of Nikon's CF objective programme. As has been noted elsewhere on this site, the whole question of what Nikon objectives belong to what series, what tube length and what system of correction of CA if any they are designed to work with, is complicated and confusing. The designation on the objectives themselves is hardly helpful either.

I agree completely about the confusing designations. The E's and E plans that I'm aware of are illustrated on pages 11 and 12 of the "New CF Lenses" brochure HERE (thanks to Charlie Krebs!).

I believe CA can also arise if a compensating eyepiece is used with an objective such as a CF that does not require such an eyepiece and also if eyepieces from another manufacturer are used which provide greater compensation for CA (over compensation) or less compensation (under compensation) than the objective manufacturer's own matched eyepieces. Is that a correct understanding?

Correct. A good model is that the CA of each lens is just some signed number. The numbers for objective and eyepiece are supposed to add up to zero. The farther the sum is from zero, the worse the final CA. CF objectives are zero by themselves so they need a zero-correction eyepiece.

--Rik

[Cactusdave]

Thanks for that confirmation Rik.

[Mitch640]

The E's and E plans that I'm aware of are illustrated on pages 11 and 12 of the "New CF Lenses" brochure HERE (thanks to Charlie Krebs!).

And there is my E-40, right there on Page 11;

"E Achromat Objectives
E Achromat objectives provide rhe highest economic value of
the achromat objectives. Designed for exceptional quality, they
clearly outperform other objectives in their price range. An
excellent choice for use with student microscopes."

[discomorphella]

Hi Mitch--

I'll leave the optical theme for a minute. You'll likely get a lot more enjoyment out of your first histological or hematological slides if you stain them. "Live", or really, unprocessed, whole blood, is apt to be a bit, well, uninformative under regular brightfield. Staining is not, in the introductory phase, a difficult endeavor either. If you can (Ebay for example, has this kind of thing for hobbyists), obtain a small bottle of Wright's stain, in methanol. Its very easy to use, next mouse you get, just smear a small drop of the blood or make an imprint of a cut surface of an organ (say liver) onto a clean slide. Then drop the slide into a small jar of methanol (but higher alcohols like ethanol or isopropanol will be fine for your purposes too, you're not trying to render a diagnosis for this mouse, which is presumably beyond help at this point anyway...) for a few mins. Then remove the slide, lay it on a level surface, and gently drop approximately 10-15 drops of Wrights onto it. Leave it for 2 minutes, then add an approximately equal amount of distilled water, blow gently on the surface to mix the liquids and let it sit for 4 mins. Now wash the whole thing quickly in distilled water, let it dry and apply a coverslip (with mounting media, or even just a drop of immersion oil will be fine). You'll see all kinds of interesting things. There are many refinements to be tried later, but this simple technique basically works for all kinds of human and veterinary samples. Enjoy your new scope, we all have to work the bugs out of our setups, believe me, I've had to debug 500k$ new confocal systems (no brands mentioned of course...).
Happy Thanksgiving and good hunting. Actually, this reminds me, you could easily make an imprint slide from a small piece of turkey liver (just keep the Wright's out of the gravy... ).
--David

[Tom Jones]

Hi Mitch,

David's right. Blood is MUCH nicer stained.

If you don't happen to have any Wright's or Giemsa stain handy, you might try talking to the lab at your local hospital to see if they might be generous enough to run the blood smear the next time (usually just a few minutes) they stain their hematology slides. They'll probably look at you kinda funny for a moment, but they'll probably do it for you.

If you ask REALLY nice, they might be willing to give you a blood slide or two they're about to discard from a patient - after stripping anything resembling identification from it that is. Labs don't keep them forever - usually only a couple of weeks - so there are always some being discarded.

You also might ask if one of the techs would show you how to make a nice blood smear. The traditional "wedge" smear isn't hard but takes a bit of practice to get a nice feather edge to the smear. Another trick is to take a small drop of blood, place it more or less in the center of a slide, then place another slide on top of it, squeeze it together gently and then pull them quickly apart along the plane of contact. It's pretty easy to do and often gives a pretty smear. This method works really well with bone marrows since it flattens the spicules and spreads out the marrow cells nicely.

I've never had anyone ask, but I'd have been happy to stain a few slides for someone who was interested, or give out a few slides I'd removed ID from.

Tom

[Mitch640]

Thanks for the explanations David and Tom. I have saved them to a txt file so I don't forget.

As it just so happens, I live in a University town that specializes in the medical field. The biggest employer here is hospitals. My gears are already turning about where and who to hit up for some old slides. LOL It should be real easy, now that you have pointed me in the right direction.

Also, the suggestion for the turkey liver was great, and I bet a nice piece of bloody steak would also work, and ya know what, it's hunting season and most of my neighbors bring home all kinds of wildlife in the back of their pickups. It's just embarrassing that I didn't already think of it myself. LOL

I do think my first smear came out good though. Many years ago, I saw it done, using a slide, a drop of blood and a coverslip. That memory was still there and I was able to get a perfect feather edge out of it. And I can see where that is important to, as anything thicker than a single cell would be hard to see through or differentiate the cells at all. I do wish I had taken some more video of the whole thing though, cause watching those cells move was fascinating.

[discomorphella]

Hi Mitch--

Muscle tissue like steak is best viewed if you tease out a few fibers and squash them into some saline or better, some 0.05% Toluidine Blue O (you can add some glycerol to make a better temporary mount if you want). Softer organs you can just cut, wash in saline and then press against a very clean slide. In fact, while you are scrounging around the local histology labs, hopefully scoring some Wright's stain, see if you can get some Toluidine Blue O. If I had to name my "desert island" stain, it would be TBO. A very simple dilute aqueous solution (0.05 to 0.5%) will usefully stain almost any sample, from higher animal organ imprints to mushrooms. For some samples, you'll even see different structures highlighted in red or purple (metachromatic staining) in addition to the usual blue. Another good home-safe combination (meaning you can spill it on yourself and not worry about producing mutant offspring) is Methylene Blue and Phloxine B. The TBO can substitute nicely for the MB also. Let me know what dyes you have found, and I'll provide you with some simple techniques to use them. Your idea of helping everyone clean their prey and making slides is a good one, I get lots of my histological specimens from hunting and fishing. You know the old joke "biology is the art of killing with finesse". Or it was until biochemical and molecular biological methods improved to the point where you don't have to kill things anymore to figure out what's going on, you can just make them express GFP and look at them under the microscope. If you get enough specimens from your hunting buddies, eventually you may find yourself seated in front of a microtome in your spare time...Just don't blame us on the forum when you realize you have no spare time left as you work out your next camera adapter, and there's a pile of blocks to be sectioned and the yard still needs to be mowed!!

David

[Mitch640]

HAHA, spare time? What's that? I will keep in touch in this thread if I am able to find any stains. I have a doctors apointment in a couple weeks so I will start then. LOL

[NikonUser]

How does one make physiological saline?
I have a 100 mL graduated cylinder so can get a reasonably accurate volume of water, but how do I measure the small amount of NaCl needed without access to a decent balance?
A level teaspoon of salt would need how much water?

[discomorphella]

Hi NU--

Regular NaCl (as in reagent grade crystals) has a density of approximately 2.17 g/ml. You need 0.9% NaCl aq to get mammalian physiological saline (avian and amphibian and reptile are all a bit different). So if you had a decent chemically pure grade of salt, 5 ml (1 tsp) would need about 1.2 liters of DI water to get you to physiological concentration (2.17 g/ml * 5ml = 10.85g ; to make 0.009 g/ml, you would need 10.85g/0.009g/ml = 1205 ml). The units come out ok, unless my food-induced coma / hangover from last night has addled my brain. One thing to watch out for; many store-bought salt brands have some trace amounts of NaI or similar in them, and things like kosher salt may be less dense. I'd stick with the small, fine crystalline salt, if you can fine some that's non-iodized. One tsp in 1200 ml of water should be fine. If you can't find non-iodized salt don't worry; if you're trying to use this for general rinsing of samples and not tissue culture, its not going to matter.

David

[NikonUser]

Thanks David.
That's a lot of distilled water (no free/cheap supply), so:
If I want 0.75% saline (closer to invert. blood than 0.9%)
I would need 5mL salt in 1,447mL.
OR: 5mL salt in 95mL DI water, 7mL of this concentrate in 93mL DI water?
(It's been a while since I played with dilutions)

[Pau]

Because this solution has a lot of medical applications, it's sold in pharmacy stores. Here in Spain it's easyly available and inexpensive.