There are many books on histological technique. Most of them are expensive.
The 5th edition of John Kiernan's excellent HISTOLOGICAL AND HISTOCHEMICAL
METHODS will sell for $100 when it comes out in July. If that is too much for
your budget, there are older books that cover 95% of what you
I run an academic lab on a very tight budget. A paper towel used to dry washed
hands is used again. Outdated dye solutions are adsorbed onto a small pile of
old paper towels to save on waste disposal costs. (A quarter-pound of solid
waste costs less to dispose of than 2 liters of aqueous
Picric acid bound to collagen is not an explosion hazard. Even if it were,
the surrounding paraffin wax would cushion the picric acid to the point of
making it shockproof. Most of the picric acid in a fixative ends up in the
hazmat bottle rather than in the tissue. Thus even putting 50 or
Formalin does burn. If you soak a piece of paper in formalin, you can set it on
fire with a match.
Formalin is also slightly carcinogenic.
The biggest problem with formalin is that a few people are severely allergic to
it.
-Allen A. Smith, Ph.D.
Barry University School of Podiatric Medicine
Normally, I work alone. Sometimes a graduate student works with me. In a few
cases, where special hazards are involved (e.g. lithium aluminum hydride,
diazomethane), I will work only if I am alone in my lab so that I will be the
only one injured AND someone who could help is present in an
Alcian blue lasts until loss of dye onto the sections weakens staining. If one
runs many slides through it, it may not last 6 weeks. With light use, it may
last 6 years.
Allen A. Smith
Professor of Anatomy
Barry University School of Podiatric Medicine
-Original Message-
From:
Osmium tetroxide solutions will fix your skin on contact, but skin grows
back. Osmium tetroxide fumes will fix your corneas which are irreplaceable.
Osmium tetroxide is dangerous, but it is manageable. First, make sure your
fume hood draws really, really well. Put 100 ml of pH 7.2
We hire a licensed waste disposal company.
Allen A. Smith, Ph.D.
Professor of Anatomy
Barry University School of Podiatric Medicine
Miami Shores, Florida
-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu
[mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of
The time and trouble of making up a solution of aluminum mordanted nuclear fast
red repays you well. The staining solution contains to work well for an entire
year.
If you must have something else, brazalum, made by substituting brazilin for
hematoxylin in Mayer's hemalum, works very well.
For the last couple of years, I've thought this was true, but I didn't have the
guts to say so. Thank you, Richard for bringing the truth out and getting it
accepted.
Allen A. Smith, Ph.D.
Professor of Anatomy
Barry University School of Podiatric Medicine
Miami Shores, FL
-Original
I have used Vector's ImmPress and ABC secondary antibody systems. I like
ImmPress better: it has fewer steps, it gives me less background, and the
reagents last longer. Vector's Nova Red has given me better results than any
other stain that I have tried.
-Allen A. Smith
Barry University
Glass glassware, including glass pipets, has always been accurate enough for my
needs in making up histological stains. This is not (repeat NOT) true of
pipettors with disposable tips: Some of my pipettors deliver exactly what they
say they do, some deliver 50% more, and some deliver 3 times
My mentor, Nick Roman, told me that sections adhere to the slide better if they
go on shiny side down. Brenda Disbrey's HISTOLOGICAL LABORATORY METHODS says
that laying the sections on the water bath or water droplet shiny side down
makes it easier to remove creases. Benno Romeiss'
I am having difficulty localizing topoisomerase IIalpha with IHC. Mouse
primary/biotinylated horse secondary works for me only some of the time.
Rabbit primary/biotinylated goat secondary doesn't work at all for me. Does
anyone have any suggestions for improving the performance of either
Try coating the slides yourself with Vector Labs Vectabond. Neither I nor
any of my graduate students have ever lost a section from a slide coated with
Vectabond.
-Allen A. Smith, Ph.D.
Professor of Anatomy
Barry University School of Podiatric Medicine
Miami Shores, Florida
Formalin fixation kills most but not all bacteria. A significant percentage
(~10%) of the mycobacteria present in the tissue remain viable. Tuberculosis
bacteria have been cultured from ffpe blocks. Prions (Creutzfelt-Jacob
disease) are believed to survive formalin fixation.
Allen A. Smith,
The reference is K.F. Gerston et al: Viability of mycobacteria in
formalin-fixed tissues. International Journal of Tuberculosis and Lung
Diseases. 2:521-523 (1998).
-Allen A. Smith, Ph.D.
-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu
The best solution would be to give them to the Cooperative Human Tissue Network.
Allen A. Smith
Professor of Anatomy
Barry University School of Podiatric Medicine
Miami Shores, Florida
-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu
I like the Vector ABC horseradish peroxidase kits, especially with the Nova Red
chromogen. Mouse primary antibodies usually give me better results than rabbit
or goat primaries.
-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu
We bought materials for neutralizing formalin and started to use them. Two
years later, the local EPA authorities told us to stop because we lacked the
competence to neutralize hazardous materials. Our neutralizing solution
became a hazardous material. We now pay a waste hauler to take our
My mentor used forceps. Thus I have always used forceps. I think the paraffin
ribbon would melt onto fingers on a warm day.
Allen A. Smith
Professor of Anatomy
Barry University School of Podiatric Medicine
Miami Shores, Florida
-Original Message-
From:
We have separate storage cupboards for inorganic acids (many of which are
oxidizing agents) and organic acids (many of which are flammable). If we
didn't have an organic acids cupboard, we would store it with organic solvents.
The reaction of acetic acid with nitric acid might be violent, the
When I wrote my thesis on sweat glands, my experience was that fixation in
Helly's fluid (Zenker-formol) and paraffin embedding caused serious
shrinkage: about 10-15% in each dimension.
Allen A. Smith
Professor of Anatomy
Barry University School of Podiatric Medicine
-Original Message-
This is what comes of letting people with little or no training in chemistry
call themselves chemical waste mangers.
Yes, methyl methacrylate is highly reactive when mixed with 1% benzoyl
peroxide: it forms a hard, brittle polymer. Pour your left over methyl
methacrylate and benzoyl peroxide
Look up Waste Hauling in the Yellow Pages. Ask every company listed for a
quote.
Make sure the ammoniacal silver nitrate is wet; when it dries, it can rearrange
to silver azide, a powerful and touchy primary explosive.
-Original Message-
From:
I use Sheehan and Hrapchak's formula and stain 1/2 hour at 45 degrees C.
-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu
[mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Evans, Andria B
Sent: Monday, January 25, 2010 2:42 PM
To:
C.L. Sturkey (www.sturkey.com) in Lebanon, Pennsylvania, sharpens permanent
microtome knives. They also have two grades of disposable blades.
Allen A. Smith, Ph.D.
Professor of Anatomy
Barry University School of Podiatric Medicine
-Original Message-
From:
Cleaning slides strikes me as a very poor use of student time. If they are
paid even 3/4 minimum wage, it is uneconomical. If they are unpaid, it is
unfair to set them a task they can learn nothing from.
-Allen A. Smith, Ph.D.
Professor of Anatomy
Barry University School of Podiatric
The stuff found in the Judean caves is kal ilan, indigo derived from
Indigofera tinctoria. It is chemically identical to tekhelet, indigo derived
from Murex trunculus by photochemical debromination of Tyrian purple. Although
some second and third century Jews, knowingly or unknowingly, used
The problem is letting people with little or no knowledge of chemistry make
chemical decisions. Some chemicals are unstable on the shelf, some are not.
Methyl green decomposes in a year or two, tetracycline decomposes in 5 years,
indigo lasts for millennia. Indigo used by Bar Kochba's
Does he require his wife to wear chain mail gloves while paring potatoes?
-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu
[mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Jackie M
O'Connor
Sent: Friday, September 11, 2009 9:31 AM
To: Jean Warren
Cc:
Formaldehyde is flammable; formalin is not. Above 122 degrees Fahrenheit
enough formaldehyde evaporates from formalin to create a modest fire hazard in
the fumes just above the liquid. Try this: pour 3 ml of formalin (37%
formaldehyde) into a watch glass under a fume hood (fan off). Touch a
I can't imagine why any one would use dioxin, which used to be used as an
insulator and heat convector in electron microscope transformers, as a clearing
agent. I think you mean dioxane.
-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu
If you ripen it with air or sodium iodate, alum hematoxylin is quite safe.
FDC green #3 is food grade fast green FCF, an excellent stain for collagen.
FDC yellow #5 is tartrazine, a plasma stain.
-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu
I think a fume hood and a safety shower should be standard laboratory
fixtures.
I have a fume hood. I use it for manual coverslipping. I require my
students to do the same. OSHA says that exposure to xylene fumes can cause
liver damage. There is also a persistent rumor that repeated
I have been very happy with the Olympus C-7000. It will focus at 3 1/4 inches
(8 cm) in its macro mode.
-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu
[mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Reuel Cornelia
Sent: Wednesday, May 20, 2009 4:53 PM
We keep our DAB waste in a plastic container and give it to our waste disposal
company every 6 months.
No one has objected to this procedure.
Allen A. Smith, Ph.D.
Professor of Anatomy
Barry University School of Podiatric Medicine
Miami Shores, FL
-Original Message-
From:
We were told that, in Miami-Dade County, we would have to apply for a license
to do this. We gave up on the cumbersome licensing procedure.
Allen A. Smith, Ph.D.
-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu
[mailto:histonet-boun...@lists.utsouthwestern.edu] On
I can spot spam and o.t. posts and delete them in a half second each. The
questions on Histonet make me aware of the extent of use of special stains, and
the potential for new stains. Jjob postings are forwarded to the career service
office here. The safety warnings are often very useful.
-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu
[mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Jennifer
MacDonald
Sent: Friday, March 20, 2009 4:45 PM
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] Current books for Histotechnology
Our
Definitely get the 4th edition of Kiernan. Also get the latest edition of
Polak and van Noorden's INTRODUCTION TO IMMUNOCYTOCHEMISTRY. If you don't
already have them, used second or third editions of Lillie's HISTOPATHOLOGIC
TECHNIC AND PRACTICAL HISTOCHEMISTRY and Pierce's HISTOCHEMISTRY,
For an atlas, Ross and Pawlina's HISTOLOGY, Wheater's FUNCTIONAL HISTOLOGY, or
Gartner and Hiatt's COLOR ATLAS OF HISTOLOGY are all good. They are also more
accurate than DiFiore.
-Allen a. Smith,Ph.D.
-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu
Formaldehyde can mess up the bacteria that treat the sewage. We used to
neutralize our formaldehyde with Richard Allen's Vytac and send the results
down the drain. Two years ago we were told that we would have to get a license
and do quantitative analysis on each batch in order to continue
A lot depends on the stain chosen. I have used Vector's Nova Red for the last
3 years. I have seen no fading of my older Nova Red slides.
Prof. Allen A. Smith
Barry University School of Podiatric Medicine
Miami Shores, Florida
-Original Message-
From:
Almost any method will work in intestine. I have had good results with Holmes'
stain and more impressive results with Kiernan's method. I have also had good
results with Winkelmann's method, but it is terribly time consuming.
-Original Message-
From:
Some tap waters blue hematoxylin very well, some not so well. I avoid the
issue of differences in tap waters by blueing in 1.2% aqueous lithium carbonate
(almost a saturated solution).
- Allen A. Smith, Ph.D.
Professor of Anatomy
Barry University School of Podiatric Medicine
While I prefer to go by my own name, I recognize that some people rightly fear
to do so. An anonymous source may still have something worthwhile to say.
Experience does not always teach well. The wise and experienced Linus Pauling
and Erwin Chargaff were wrong about the structure of DNA; the
B5, Schaudinn's, SUSA, Helly's, and Zenker's fixatives contain mercury (II)
ions, which are VERY poisonous. Sewage bacteria convert mercury (II) to methyl
mercury, which is very, very poisonous. (For a shocking example of mercury
toxicity, see the photo essay MINAMATA by W. Eugene Smith.)
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