QUARTERLY BEAT / DECEMBER 2025
DIAGNOSING AND IMMUNOPHENOTYPING CANINE LYMPHOMA
Lymphoma is one of the most common cancers diagnosed in pet dogs, with the most frequent presenting clinical sign being “lumps” in the neck (ultimately determined to be peripheral lymphadenopathy on exam). Most primary care veterinarians are proficient at performing fine needle aspiration to confirm the diagnosis via cytology, either by shipping samples to a reference lab or digital cytology, which is available through multiple platforms. We encourage our primary care vets to also take the next step, which is to collect samples for immunophenotyping, i.e., B-cell vs. T-cell testing. The immunophenotype of lymphoma has a significant impact on prognosis (i.e., B-cell is better, T-cell is tougher) and in most progressive oncology practices, it will help the oncologist tailor the chemotherapy protocol. When the immunophenotyping information is available at the time of initial oncology consultation, it allows the specialist and pet owners to have a more informed and directed conversation about treatment options and expected prognosis. There are several options for immunophenotyping available, all of which can be performed by the primary care veterinarian via fine needle aspirate. 1. Flow cytometry: This is almost always the test of choice for immunophenotyping because it can both confirm the diagnosis as well as provide prognostic information via subtyping, which allows for the differentiation of unique forms of lymphoma such as indolent T-zone lymphoma. Flow cytometry requires the cells to remain alive for analysis, so most labs either recommend placing the aspirate in a saline/serum solution in a non-additive tube or using a lab-provided tube pre-filled with cell media. Often this may necessitate having the owner bring the dog back to the clinic to perform this test, which can be done during the waiting period for referral. Typically, 2-3 aspirates will provide enough cells for analysis. A turbid solution usually indicates adequate cellularity for flow cytometry testing. Samples must be sent on cold packs (not ice) via overnight shipping to ensure cell survival. 2 2. Immunocytochemistry (ICC): ICC can be performed on cytology slides at some reference labs and does not require overnight shipping of cells in a fluid medium (as with flow cytometric analysis) or an incisional biopsy (as with IHC). The sample required for ICC submission is simply 4 adequately cellular, unstained or stained, air-dried glass slides prepared the same way as those for standard cytologic evaluation. While cell size and thus an impression of lymphoma grade/aggressiveness can be discerned through cytology with ICC, the determination is somewhat subjective and not as reliable as with flow cytometry. 3 3. PCR for antigen receptor rearrangement (PARR): PARR testing can be performed on both cytology and histopathology samples at most reference labs and universities. While choosing PARR saves the pet owner from having to bring the dog back for additional aspiration (as with flow cytometry), PARR will only provide a binary B-cell or T-cell designation and cannot differentiate the grade or subtype, so indolent forms may be missed. As such, PARR is most indicated when seeking to confirm lymphoma rather than to determine immunophenotype.
CADET® BRAF MUTATION DETECTION ASSAY FOR CANINE TRANSITIONAL CELL CARCINOMA/ UROTHELIAL CARCINOMA (TCC/UC) Transitional cell carcinoma (TCC), also known as urothelial carcinoma (UC), is the most common bladder cancer in dogs. Traditionally, diagnosis has required imaging, cytology, and often biopsy—each with limitations in sensitivity or risk of invasiveness. Recent research has identified a specific mutation in the BRAF gene that occurs in about 85% of canine TCC/UC cases. This mutation, known as BRAF V595E (analogous to the human V600E mutation), was independently discovered by two research groups using different sequencing approaches— one comparing DNA from tumors versus normal tissues, and another analyzing RNA from affected tissue. The consistent discovery across both methods confirmed its strong association with TCC/UC. Importantly, this BRAF mutation is not found in non-cancerous bladder tissues or in most other canine tumors, making it a reliable diagnostic biomarker. The CADET® BRAF test (developed by Sentinel Biomedical; available through Antech Diagnostics) 4 detects the presence of tumor cells carrying this mutation in free-catch urine samples. The assay is highly sensitive— able to identify as few as 10 cancer-bearing cells—and is not affected by blood or bacteria in the sample, unlike some older urine-based tests (e.g., VBTA). A second-generation assay has since been introduced to detect an additional 10% of cases that lack the original BRAF mutation, increasing overall detection sensitivity. The CADET® BRAF test can detect TCC weeks to months before clinical signs such as hematuria, dysuria, or stranguria become apparent. Therefore, it is a valuable non-invasive diagnostic and screening tool for at-risk breeds (e.g., Scottish terriers, Shetland sheepdogs, West Highland white terriers) or dogs showing chronic lower urinary tract signs. Clinical uses for this test include:
• Screening: Annual testing in high-risk breeds • Diagnosis: Helps confirm TCC/UC in dogs with persistent urinary signs • Monitoring: Tracks treatment response or tumor recurrence through repeat urine testing
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