Maximising cytological diagnostic outcomes.
Cytology is a very useful tool in evaluation of masses and diseases of solid organs. In comparison to histology, cytology is an inexpensive test, minimally invasive, provides rapid results, and with good site selection provides good correlation with histology.
However, more than any other testing modality, cytology relies very heavily on the clinician to provide a history, to select appropriate sites for collection and to make slides of adequate quality.
Setting expectations
When choosing cytology rather than histology for investigation of a mass or an organ, it is important to set realistic expectations both for yourself and the owner. Cytology is excellent at defining the problem but may not provide the precision that the evaluation of intact tissue structure in histology can achieve. For example, cytology provides good accuracy for differentiating neoplastic and inflammatory conditions, and in many cases benign or malignant neoplasia. However, grading and more specific classification of a malignancy will usually require histology +/- immunohistochemistry.
Some tissues are more amenable to cytological examination than others. This reflects the likely aetiologies, vascularity of the tissue, and the freedom with which the tissues will exfoliate. Many papers have been published on correlation between veterinary cytology and histology, and the table below provides a guide. Note that this data is usually sourced from secondary and tertiary referral clinics.
This does not mean that tissues with lower correlations should not be evaluated by cytology, but rather that cases should be individually evaluated based on the major differentials to determine if cytology is likely to be a suitable diagnostic step.
Choice of sampling method can also dramatically affect the diagnostic outcome. For example, prostatic cytology can be highly rewarding when direct FNA are collected. Prostatic washes and massages are often less rewarding regardless of the aetiology. Similarly, targeted aggressive sampling (using a cytology brush or urinary catheters) of identified lesions in the nose can be cytologically rewarding, while nasal swabs or flushes are commonly non diagnostic.
Importance of history
History is vitally important. For every request, please include:
The absence of an adequate history may prevent the pathologist from providing an accurate interpretation or comments regarding further testing, even when an adequate sample is provided.
Selection of collection site
Where to collect the sample from seems a straightforward idea but can dramatically alter the diagnostic outcome. When there is a choice of lesions, it is preferable to collect from mature lesions, whilst avoiding lesions with evidence of necrosis or ulceration. If only one mass is present and it is ulcerated, please include this information in the history to assist interpretation of the cytological findings.
If fluid is encountered when sampling, make slides from the fluid, but also collect aspirates from the surrounding tissue. Cytology of fluid collected from cysts is rarely diagnostic, but aspiration of the surrounding tissues may provide a cause for the cyst formation.
Aspiration of the sample
When collecting the sample, secure the lesion with the fingers of your non dominant hand. Introduce the needle into the tissue. If using a syringe, apply mild negative pressure. Redirect the needle in the tissue several times using a sewing machine needle like motion, without withdrawing the needle from the tissue. Release the negative pressure (if using) before withdrawing the needle. You should see a small amount of material in the needle hub. Splattering of material into the syringe usually reflects excessive negative pressure was used or the needles was withdrawn from the tissue before negative pressure was removed. Both can adversely affect cellular preservation.
Spreading the sample
Transferring the material to the slide seems very simple, but this step is often the cause of major artefacts.
To spread the slide, use a second slide held at a shallow angle, just as you would make a blood film. Place the slide in front of the droplet and draw it back to make contact with the droplet. Let the material spread along the bottom of the slide. Now slowly slide the spreader slide along the sample slide. No downward pressure is necessary. You should now have a smear that looks like a blood smear.
https://www.biorender.com/template/blood-smear-preparation
Dry the slides rapidly by waving them in the air, placing in front of a fan, or using a hair drier on a warm setting. There is no need to fix cytology slides prior to submission.
Spray slides are commonly submitted to Vetnostics. These are slides where the material has been sprayed onto the slide, often from a distance and with some force. The material is then usually not spread. Slides made in this manner are rarely diagnostic as the cells are commonly damaged by the spraying process.
The deleterious effects of spray (A) and squash (B) preparation of slides can be seen below in comparison to well-prepared slides (C). All samples are from the same tissue and all have good cellularity. However, only slide C is diagnostically useful.
Pleaseonly submit slides with 90-degree corners. Slideswith 45-degree (bevelled) corners cannot be digitally imaged and may result insignificant delay in releasing of cytology reports.
https://www.universalmedicalinc.com/
Please label slides with the animal’s name and the site of collection. Please use pencil to label all slides – the ink from most pens including permanent markers is removed during staining. Pencil writes well on all frosted slides.
Staining slides in house
It is often an advantage to fix and stain a slide in house and evaluate it for the adequacy of sample collection and cellular morphology. This can prevent submission of non-diagnostic samples and can allow immediate recollection if required, without having to have the animal revisit the clinic. We are happy to receive slides that you have stained in house. Where possible, please submit at least one unstained slide as well.
Submission of cytology samples with histology samples
We commonly receive cytology samples submitted at the same time as histology samples. If the cytology samples are exposed to formalin vapours from the fixed samples, this causes marked artefact in the samples and usually renders them non diagnostic. To avoid this, please submit cytology and histology samples in separate bags and with separate request forms.
If you have any questions about cytology sample collection, staining or submission, please feel free to call Vetnostics and discuss this with one of the pathologists.