Histopathology and Tissue Shared Resource

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Submission Guidelines

Formalin Fixed Tissues

Fixation:

  • Fix the tissues in minimally a 5 to 1 ratio of neutral buffered formalin to tissue volume. We recommend that if possible, fix in a 10 to 1 ratio to ensure thorough fixation, especially with particularly large tissues.
  • For adequate fluid exchange, it is preferred that tissues be less than 5 mm in thickness.
  • The larger your tissue is, the longer you should leave your tissue in fixative (leave for multiple days rather than just 24 hours). If your tissue is thick or if you have mammary gland tissues, we recommend cutting the tissue in half through its thickness or into multiple pieces of approximately 2-3 mm thick to ensure adequate penetration of the fixative to the center of the tissue.
  • Be sure the formalin has room to circulate in the container. You may have a 10:1 ratio of formalin, but if a large number of tissues are all stacked up in the bottom of the container, then the tissues in the middle and at the bottom will not be exposed to adequate formalin for proper fixation.
  • Tissues will harden in fixative so flatten tissues such as gut, skin, uterus, etc., prior to fixation. We can provide sponges and cassettes to facilitate this.
  • For tissues within the stomach and intestinal tract, please flush the tissues of their contents prior to submission. Stomach and intestinal contents do not fix well and create significant morphological artifacts and tears within your tissue sections. The final tissue sections will be much improved by removal of intestinal contents prior to fixation.
  • Once a tissue has been adequately fixed, remove the formalin and transfer tissues to 70% ethanol and bring to HTSR for processing and embedding.
    * *An exception to this rule is for neural tissues, leave them in formalin as the
    ethanol may eat holes in the tissues.

Trimming and Embedding:

  • We cannot section tissues with any form of metal such as medical staples or sutures. Please remove these from your tissues prior to submission into HTSR.
  • Please let us know any special instructions for your samples. We will trim based on the assumption that your sample is precious and every piece of tissue should be preserved and embedded. If this is not so or if you need us to trim in deeply, please let us know.
  • For proper infiltration, we will trim your tissues so that they are less than 0.5 mm in thickness. If you have particular areas within your tissue that you wish to have sectioned, you may want to trim your tissues prior to submission, or if you have thick samples, discuss with us where you need cuts made.
  • We are happy to provide cassettes, sponges, containers, etc. to help you prepare your tissues for submission into HTSR.
  • If you are submitting your samples in cassettes, please label your cassettes so that the label is facing up, as demonstrated below.

    Cassette labeling
  • Be sure you are using the correct anatomical terms for planes of section. See below for a detailed description of sectional planes (reference - Developmental Biology ONLINE!)Sectional planes

 

Terms for Direction
The following terms are adjectives that are used to describe direction in the bodies of embryos and quadrupeds. The terms may be used as adverbs by adding the suffix -ly or -ad to the root.

caudal

Refers to the tail or tail-like structures. The opposite of cephalic or cranial.

cephalic

Cranial; refers to the head. The opposite of caudal.

cranial

Refers to the cranium (head).

distal

Farthest from the centre, from the midline or from the point of attachment. The opposite of proximal.

dorsal

Refers to the back. The opposite of ventral.

lateral

Refers to the side of the body.

medial

Refers to a position near the middle or midline. Median refers to a structure or section in the middle or midline.

Median

Refers to a structure or section in the middle or midline.

proximal

Nearest to the centre, to the midline, or to the point of attachment. The opposite of distal.

ventral

Pertaining to the belly. The opposite of dorsal.

 

Note: In human anatomy, the terms anterior and posterior are used instead of ventral and dorsal, and the terms superior and inferior are used to replace cranial and caudal. If you have studied human anatomy and are confused by the change in terms, think of the human being as a quadruped (on hands and knees), and the use of the terms dorsal, ventral, cranial and caudal should become clearer. However, this does not hold true for the terms anterior and superior. For an explanation to sort this confusion out, see the diagrams below:

Incorrect labeling

Tadpole planes
Human planes incorrect

On the top, we see a tadpole correctly labeled to describe the various directions and planes associated with it. Thus we would expect that a human on all fours (same relative position as the tadpole) would be labeled similarly, as we see on the lower image. Whereas this stands to reason, be aware that the labels on this human diagram are incorrect! Somewhere along the line, the terms got swapped around between humans and other animals. To put everything straight, the diagrams below show the human correctly labeled along side the tadpole.

Correct Labeling

Tadpole planes
Human planes correct

 

Here we see our friend the tadpole again above the correctly labeled diagram of the human. Note the swapping of the terms anterior and posterior from describing the difference between the "belly" and the "back" areas of the human, to describing the "head" and "buttock" areas of the tadpole.

 

 

 

 

 

 

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Immunohistochemistry

  • For each submission, we request that you submit positive and negative control slides along with your experimental slides so that we can confirm the staining procedure. We are happy to help you select appropriate controls as needed.
  • For antibodies you have previously used, submit your positive and negative control slides, and your experimental slides. You can contact us prior to your submission and we will calculate for you the volume of antibody you should submit with your slides.
  • We have years of accumulated experience with many common antibodies used in cancer research, if you are unsure which antibody you want to use, please contact us and we can try to help you select an antibody based on our database of previously used antibodies.
  • If we do not have previous experience with your antibody of choice, we can work to develop a new protocol for your antibody.
  • If you are unsure which antibody to order for your protein of interest, we can also help you select an antibody, please see Deborah Berry for assistance.

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