Skin scrapings: Know your distribution patterns! Best lesions to scrape: Demodex - alopecia with hyperpigmentation; Sarcoptes - golden crusts or papules.

Trichoscopy (hair plucks): Growth phases of hair (anagen active, catagen finishing growth, telogen resting); integrity of hair (evidence of barbering, dermatophytes), melanin clumping - colour dilution alopecia, follicular casts - sebaceous adenitis, demodicosis, folliculitis, seborrhea, Vitamin A responsive dermatosis.

Wood’s lamp: Good screening tool (false negatives, false positives) *M. canis will fluoresce apple green from root up the hair shaft ~50% of time.

Fungal culture: Collect hair and scale (especially if fluorescent green); face, ears, tail, paws - sterile plucks from edges of lesions; MacKenzie tooth brush technique for asymptomatic carriers; hold up to 4 weeks; consider PCR testing - very sensitive, so not great for monitoring response to treatment.

Cytology: **with Diff-Quick staining, allow double time in purple stain to highlight yeast, mast cells and acantholytic keratinocytes

Direct smears: Collected using a dry cotton-tipped applicator use like a crayon and “colour” in the area of interest. Gently roll sample out in a line on a microscope slide. Heat fix and stain. Intact pustules can be lanced with a sterile 25-g needle to allow for collection.

Otic cytology: Ideally collected from the junction of the horizontal and vertical canals heat fix. It’s often helpful to massage the opposite ear while collecting samples. Impression smears: performed on moist, oily skin, or exudative lesions. Best for assessing presence of acantholytic keratinocytes. Crusts can be lifted with the edge of a microscope slide. Slide is then gently pressed to the moist lesion below, or removed crust as a stamp.

Superficial skin scraping with a dulled blade or spatula (no oil): Dry blade is used to collect surface scale or exudates. Debris is spread on slide and heat fixed prior to staining.

Acetate tape preps: Great for ectoparasites and cytology. Use clear acetate tape. Handle the tape carefully to allow yourself a “sacrificial finger print end” - Keep the tape stuck to your finger and use your thumb to press tape to lesion. Press to lesion repeatedly until the tape no longer sticks, then stain and examine under microscope.

There are many ways to stain tape; I prefer to stick the “sacrificial end” to clear edge of slide then dip into the eosin stain in a Diff-Quik stain set. Hold in stain and wobble back and forth for a count of 10, then allow excess to run off back into vial. Dip in purple stain and wobble back and forth for a count of 20, then allow excess to run off back into vial. Rinse thoroughly with a gentle stream of water on both sides of tape until water runs clear. Do not use the blue fixative as it will eat the glue on the tape, taking your sample with it! Lay the rinsed tape, sticky side down, on microscope slide with “sacrificial finger print end” towards frosted end of slide. Place slide inside folded paper towel and press tape down to remove air bubbles and excess water. Apply one drop of immersion oil on top of dried tape, then coverslip and examine under microscope.

Skin biopsy: Send to pathologist with a special interest in skin disease include history, clinical presentation, differential diagnosis. Detail the number of biopsies, type of biopsy, sites of biopsy. Include lesion diagram and photos if possible. Often, a specific diagnosis is not forthcoming this may be because the biopsy did not show the primary lesion or characteristic lesions of the disease, such as pustules or bullae, were not available at the time of biopsy. Even under these circumstances, skin biopsy can still be an effective diagnostic tool as it may allow you to eliminate potential differentials, allowing the investigation to focus in other areas. Proper biopsy technique and provision of detailed clinical findings allows you to optimize results. Take 3–5 skin biopsies from lesions in all stages of development. Ulcerations are avoided since the dermis is damaged and won’t provide information to the histopathologist. Depending on the stage of a disease process, biopsies may need to be repeated. Always inform your clients of this possibility.

Hypoallergenic diet trials: Several types of testing, including serum allergy testing exists, but we are still unable to correlate serum test results with actual patient clinical signs so please don’t waste your client’s money on serum testing for food allergies. (1,2,3,4,5,6,7,8,9)

The vast majority of adverse food reactions in animals are caused by the protein source in a diet. A hypoallergenic diet to rule out food allergy ideally contains a single, novel source of protein with generally a single source of novel carbohydrate. In theory any protein may be suitable, provided the patient has not been exposed to the protein significantly before.

Homemade diets are preferred to commercial diets where possible as they do not contain any additives (preservatives, binders, etc.). Commercial limited ingredient veterinary diets, and hydrolyzed diets are also options. In theory, hydrolyzed diets have the protein chopped up so finely, that the body does not recognize it and an allergic response is avoided. In my experience, I have seen many animals still react to these diets, so we usually reach for them when we have a patient with chronic GI symptoms. Having the protein hydrolyzed is very similar to it being partially digested, allowing for easier absorption in the gut. Another train of thought is to avoid an animal-based protein all together and go with a vegetarian diet. Response is very individual - there is no magic diet that will work for all cases.

A 2010 study (10) showed 100% of over the counter hypoallergenic diets were contaminated with protein sources not listed on ingredients. This is a result of cross contamination from other diets manufactured at the food plant. The same study showed veterinary prescription diets were not cross contaminated, so preference is given to prescription diets when doing a diet trial.

Diet trials must be strict, for a minimum 8 weeks. Warn clients that it could take several attempts before finding the diet that works for that particular individual. Intradermal Allergy Testing: Still regarded as the gold standard since we see what the skin reacts to.

Very specialised procedure - Only available through veterinarians with a special interest and training in skin disease. Intradermal skin tests can be significantly affected by medications such as corticosteroids, anti-histamines and fatty acid supplements.

Serum allergy testing: Tells us what antibodies are being produced in the bloodstream. This result gives an accurate assessment of how ready the immune system is to react to a given allergen, but not whether you will actually have a reaction. This is very useful if intradermal testing is not available or possible due to drug withdrawal periods. These tests may still be affected by medications, but usually less so. Results need to be interpreted by comparison with the environment and history of the pet. Currently our serum allergy test of choice at Yu of Guelph Veterinary Dermatology is VARL Liquid Gold.

This test is done at the Veterinary Allergy Reference Laboratory in California. Serum allergy testing is currently of no benefit when testing for food allergy.

References

1.  Mueller R, Olivry T. Critically appraised topic on adverse food reactions of companion animals: can we diagnose adverse food reactions in dogs and cats with in vivo or in vitro tests? BMC Veterinary Research BMC series - open, inclusive and trusted; 2017; 13:275.

2.  Johansen C, Mariani G, Mueller R, Evaluation of patch testing with proteins, carbohydrates and commercial foods for diagnosis of canine adverse food reactions. Proceedings from 26th Annual Congress of the ES-VD-ECVD/19–21 September 2013, Valencia Spain; 197.

3.  Mueller R, Tsohalis. Evaluation of serum allergen-specific IgE for the diagnosis of food adverse reactions in dogs. Veterinary Dermatology; 1998 July; 9(3):167–171.

4.  Hardy JI et al. Food specific IgE and IgG reactivity in dogs with and without skin disease: lack of correlation between laboratories. Veterinary Dermatology; 2014;25:447–455.

5.  Guilford et al. Development of gastroscopic food sensitivity testing in dogs. Journal of Veterinary Internal Medicine; 1994 Nov; 8(6):414–422.

6.  Allenspach et al. Chronic enteropathies in dogs: evaluation of risk factors for negative outcome. Journal of Veterinary Internal Medicine; 2007;21;700–708.

7.  Ishida et al. Lymphocyt blastogenic responses to inciting food allergens in dogs with food hypersensitivity. Journal of Veterinary Internal Medicine; 2004;18:25–30.

8.  Fujimura et al. Commensal microbe-derived butyrate induces the differentiation of colonic regulatory T cells. Nature International Journal of Science; 2013;504:446–450.

9.  Coyner K, Schick A. Inaccuracies of a hair and saliva test for allergies in dogs. Veterinary Dermatology; 2016;27:68

10.  Raditic DM, Remillard RL, Tater KC. ELISA testing for common food antigens in four dry dog foods used in dietary elimination trials*. Journal of Animal Physiology and Animal Nutrition 2011;95:90–97.