Histological Tests

The shipment of biological specimens must comply with the protocols established by the courier or transport service used. In some cases, air transport requires compliance with the International Air Transport Association (IATA) regulations for dangerous goods.

Tissues for microscopic examination collected by biopsy or during necropsy can be essential for obtaining a diagnosis. Immunohistochemistry (IHC) and other advanced tests that can be applied to formalin-fixed tissues further reinforce the usefulness of histological evaluation as a diagnostic technique.

Autolysed tissues are usually useless for histological evaluation; rapid necropsy and organ sampling are essential.

Tissues must not be frozen before fixation.

Apart from CNS tissues, samples collected for histological evaluation must never be more than 1 cm thick (preferably 5-7 mm) and must be placed immediately in ≥ 10 times their volume of phosphate-buffered 10% formalin to ensure adequate fixation.

Tissues must be representative of any lesion present and, in the case of punch skin biopsies and biopsies obtained by endoscopic collection, must be centred directly on the grossly visible lesions.
Wedge biopsies or tissue samples collected at necropsy must include part of the apparently normal surrounding tissue; the interface between normal and abnormal tissue can provide key information.

Excisional biopsies of small tumours (< 1.5 cm) may be cut in half. Larger tumours can be sliced like bread so that the formalin can penetrate to the face of each slice. Alternatively, several representative samples may be collected (7 mm wide, including the interface between normal and abnormal tissue).

Instead of sending large volumes of formalin, tissue samples may be fixed internally for at least 24 hours in an adequate volume and then transferred to a smaller volume of fresh formalin for shipment.

The submission of tissue stored in formalin is acceptable at any time, although sample quality will degrade over time. Prolonged fixation in formalin can adversely affect advanced tests; for example, immunohistochemical testing is not recommended on tissues that have been stored in formalin for more than 7 weeks, as the results may not be valid.

Samples must be sent in unbreakable containers approved by the laboratory with screw caps and packed so as to prevent spillage or freezing during shipment (see Packing samples for shipment).

Specific tissues collected at necropsy require additional attention:

As the gastrointestinal mucosa decomposes rapidly, short sections of intestine collected at necropsy must be opened longitudinally to allow adequate fixation.

If the spinal cord is submitted, a careful longitudinal incision must be made in the dura mater to allow faster penetration into the cord.

Fixation of the brain poses a special dilemma, especially if the neuroanatomical location of the lesions within the organ could not be determined antemortem.
- Ideally, a whole, intact, fixed brain is required for a complete histological evaluation.
- Immersion of the brain for many days in a large volume of formalin is necessary for adequate fixation; therefore, brains are normally transported in an only partially fixed state. If the specimen can be sent overnight, it may be acceptable to send a refrigerated, carefully packed, unfixed brain, which can then be processed at the diagnostic laboratory.
- Often, the brain is divided longitudinally in half and one half is sent unfixed (fresh), properly refrigerated, for microbiological testing, while the other half is partially fixed during transport. This method may prove unsatisfactory if it is a solitary unilateral lesion. Cutting the brain into suitable portions for rapid fixation introduces considerable fixation artefacts and should be avoided as far as possible; it is preferable to fix the brain intact (or divided in half) in a large volume of formalin for > 24 hours.