U.C.
Davis
Mutant Mouse HistoPathology Laboratories
Robert D. Cardiff
Professor
Department of Pathology
(530) 752-2726
rdcardiff@ucdavis.edu
The
We have been offering technical and interpretive services to our
scientific colleagues for over nine years. At the last count, we have received
samples from over 200 investigators from 100 laboratories in 8 countries. This
has provided us with a wonderful and unique experience with the pathology of GEM animals. We are always eager
to expand our knowledge and to help other interested scientists.
These services are provided at cost of labor, supplies and shipping. To
initiate services, please provide your purchasing or
billing number. We have added a special section for those of you interested in
the fixation and processing of samples for IHC or ISH. Please note and follow
the instruction sheet whimsically entitled "Laf-A-Pant Bag". Since transgenic pathology is
frequently different, the interpretation is greatly enhanced by having
experimental details. Fill out the attached Pathology Report
form. You may copy this form for your use. If you are primarily interested
in Mammary Pathology, you might use our synoptic forms that are in PDF format.
We can routinely return your H&E stained sections within seven
working days. Special stains are available upon request. If you need faster
service, mark the sample as ASAP on the form. My written description and
diagnoses are generally available within 14 days. FAX reports are available
upon request [(530)752-7914]. Telephone consultations are always welcome. I am
also on Internet (rdcardiff@ucdavis.edu). I enjoy discussing the data with
fellow scientists.
The histology is done by the Mutant Mouse Pathology Lab personell which
will be glad to answer technical questions or help with your shipping
[(530)754-5524]. We can also offer publication quality photomicrography when
appropriate. Bob Munn is our electron microscopist
and photographic specialist [(530)752-5524].
If you have any questions, please
let us know.
Sincerely yours,
Robert D. Cardiff M.D., Ph.D.
Please forward question and comments to: RDCardiff@UCDavis.edu
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FIXATION AND PROCESSING OF MOUSE TISSUES
Our laboratory has been experimenting with the fixation and processing mouse tissue for Immunohistochemistry and In Situ Hybridization. The following recommendations come from this experience.
GENERAL:
1) No ONE fixative works for all antigens and all antibodies. Conditions must
be optimized for each antigen-antibody combination. When first starting, we
recommend that you place the tissue in a variety of fixatives to find the
optimum for your tissue, antigen and antibody.
2) Fixation should be rapid but thorough.
a) The optimal tissue size is no thicker than 1-2mm. If a tissue is thicker,
the investigator should fix for one hour, and then cut 1mm thick sections
("bread-loaf") so that all surfaces are exposed equally.
b) The Volume of fixative should be at least 10x the volume of the tissue. If
the tissue is "bloody", remove the bloody fixative after an hour and
add fresh fixative.
c) A tissue can be placed in one fix and then portions placed in a second
fixative for "mordanting". (Example: fix in
paraformaldehyde for ISH. Place a paraformaldehyde
sample in Bouin's Fix for IHC.)
3) Once the tissue is fixed, it should be held in 70% alcohol and processed
promptly.
4) Paraformaldehyde is the best fixative for ISH. Bouin's Fixative is the best all-around fixative for IHC.
(Do not use the Bouin's substitute without Picric
Acid. The Picric Acid is required for best results.) However, excellent results
have been obtained with Carnoy's, B5, and
formaldehyde. Zenker's and Telly's
fixatives have seldom provided satisfactory results.
PROTOCOL:
1) Remove tissue as rapidly as possible. Place in 10 volumes of fixative. Cold
fixative is better than warm or room temperature fixative.
2) After one hour, inspect sample. If the fixative is cloudy, replace with
fresh fixative. If the sample is thick, remove sample, place on a cutting
board, cut 1-2mm slices to expose all surfaces and return to 10 volumes of
fixative. (note: the short fixation will firm up
hard-to-slice tissues such as lymph nodes, spleen and thymus, making them
easier to slice).
3) Fix for six to 12 hours of over night BUT no longer. Remove and place in 70%
ETOH until shipping or processing.
SAMPLING:
1) If the tissue is of interest, make sure that at least 50% of the sample is
composed of normal adjacent tissue.
2) In the case of the mammary gland, the gland should be spread on a slide so
that the fat does not inhibit fixation. After an hour, the flattened mammary
gland can be removed from the slide and placed in more fixative.
3) If one organ is diseased, make sure that the contralateral
organ is sampled (if the organs are bilateral).
4) The best interpretation is provided when age matched control organs are
provided.
INFORMATION:
1) Make sure that the sample is submitted with the appropriate request forms
that includes demographic and experimental data about the animal.
LAF-A-PANT BAG
(COLLECTION, FIXATION AND SHIPPING INSTRUCTIONS)
- (Label and Form):
1) Place the label with date, animal and specimen number written with a #2 lead
pencil (no ink please) on white index card in specimen container with fixative
and sample.
2) Fill out Pathology Report Form (see form below). The gross description
should identify the tissue source and describe the tissue. The experimental
notes will frequently become critical in the interpretation of the
histopathology. Please give us the necessary information so that we know what
we are searching for.
- A (Alcohol): Fix
sample in 10 volumes or more of a formalin or alcohol-based fixative such
as Omnifix. For the best fixation, samples
should not exceed 1 to 2 mm in thickness. Although formalin based
fixatives are accepted, they cannot be shipped through the mail without a
permit. Samples initially placed in a non-alcoholic fixative should be
dehydrated in alcohol and shipped in either 70% alcohol or a suitable
buffer. Alcohol fixed samples are not considered biohazards and may be
sent via the
- PANT (Pathologic and Normal Tissues): Specimens should be sampled across the transition area between grossly disease-free and diseased tissue. This transition zone should be at or towards the center of the sample. If the disease-free tissue is not available, a sample from the same area from a comparable disease-free mouse should be provided. All samples will be embedded with the cut of flat surface down unless other orientations are requested (i.e. on edge).
- BAG (Bag and Ship your
Sample): Place the fixed sample with 5-20 cc of fixative or buffer and the
index card-label into Kapak/Scotchpak heavy duty
heat sealable pouches (4"X6"). Press flat to remove air bubbles
and seal with hot iron about 2/3 up the bag. Double seal by sealing at the
very top of the pouch. Attach the Pathology Report Form. Place in padded
envelope and ship to :
Mutant Mouse Pathology
Lab
C/O Robert D. Cardiff, M.D.,Ph.D.
Professor of
County Road 98 and Hutchison Drive
University of California, Davis
Davis, CA 95616
Phone: (530)754-5524
FAX: (530)752-7914
rdcardiff@ucdavis.edu
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Histology Rates
Effective November 2006
|
Description |
On-Campus Rate (UC) |
Off-Campus Rate |
|
H&E Slide |
$ 11.00 |
$ 16.00 |
|
Embedding (per block) |
6.00 |
8.00 |
|
Recut (each section from one block) |
3.00 |
4.00 |
|
Special Stain (per slide) |
12.00 |
17.00 |
|
Silver Stain (per slide) |
23.33 |
33.00 |
|
Stain w/Special Handling (per hour) |
228.00 |
329.00 |
|
Immunohisto Chemistry - Routine |
62.00 |
89.00 |
|
Immunohisto Chemistry – Basic Workup |
438.00 |
633.00 |
|
Electronic Handling Fee (per admission) |
6.00 |
9.00 |
|
Non-Electronic Handling Fee (per admission) |
11.00 |
16.00 |
|
Basic Special Procedures |
62.00 |
89.00 |
|
Standard Consultation (per hour) |
117.00 |
169.00 |
|
Advanced Consultation (per hour) |
212.00 |
307.00 |
|
Whole Slide Imaging |
50.00 |
72.00 |
|
Photo Microscopy |
57.00 |
83.00 |
|
Dye Sublimation (printing supplies) |
6.00 |
9.00 |
|
Basic In Situ Hybridization (ISH) (per slide) |
141.00 |
204.00 |
|
Advanced ISH (per slide) |
1025.00 |
1480.00 |
|
Non-Targeted Tissue Microarray (per hour) |
227.00 |
328.00 |
|
Targeted Tissue Microarray (per hour) |
131.00 |
189.00 |
|
Necropsy/Surgery Services (per hour) |
55.00 |
80.00 |
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