Cytology Catches Early Spread: Wright–Giemsa Beats H&E for LN Metastasis in Canine MCTs

Comparative Evaluation of Cytological and Histopathological Diagnostic Performance in Detecting Nodal Metastasis of Canine Mast Cell Tumours

Xavier Escoda Llorens; Ignacio Amarillo-Gómez; Isaac Carrasco-Rivero; Ignacio Rodríguez-Pizà; Antonio Meléndez-Lazo

Background

Regional/sentinel lymph node (LN) assessment is central to staging canine mast cell tumours (MCTs), but the diagnostic performance of cytology (Krick’s criteria) versus histopathology (Weishaar’s HN0–HN3 system) remains variably reported. Routine H&E histology can miss early metastasis (HN2) without mast cell–specific staining, while cytology using modified Wright–Giemsa stain (WGS) might detect early spread. This study compared paired cytology and histopathology to validate cytology’s accuracy and to test whether cytology outperforms H&E for early metastasis detection.

Methods

Multicentre retrospective study (May 2021–March 2025) of 65 LNs from 52 dogs with MCTs that underwent lymphadenectomy and had prior LN fine-needle aspiration classified by Krick’s criteria (normal/reactive; possible; probable; certain metastasis) using methanol-based WGS. Corresponding LNs were graded histologically per Weishaar (HN0–HN3); toluidine blue stain (TBS) was added when H&E did not show metastasis. Diagnostic metrics were calculated for three cytology “positivity” groupings: G1 = certain; G2 = certain+probable; G3 = certain+probable+possible. ROC and κ statistics assessed performance and agreement.

Results

Among 65 LNs, histopathology classified 22 HN0, 7 HN1, 16 HN2, and 20 HN3; TBS was used in 76.9%. Cytology distribution was 21 normal/reactive, 10 possible, 5 probable, 29 certain. Cytology detected 14/16 (87.5%) HN2 and 19/20 (95%) HN3 cases, whereas routine H&E identified 4/16 (25%) HN2 and 17/20 (85%) HN3; the remainder required TBS. Diagnostic performance:

G1 (certain positive): sensitivity 75%, specificity 93.1%, PPV 93.1%, NPV 75%, LR+ 10.9, accuracy 83.1%, κ = 0.665 (substantial).

G2 (certain+probable): sensitivity 80.6%, specificity 82.8%, PPV 85.3%, NPV 77.4%, accuracy 81.5%, κ = 0.629.

G3 (certain+probable+possible): sensitivity 91.7%, specificity 62.1%, PPV 75%, NPV 85.7%, LR− 0.13, accuracy 78.5%, κ = 0.55 (moderate).
Combined ROC AUC for cytology was 0.879, indicating good overall diagnostic performance.

Limitations

The retrospective design, modest sample size, and referral-center case mix introduce selection bias toward suspected metastatic nodes. Histology was read by multiple pathologists, and “possible/probable” cytology categories are inherently subjective. Long-term outcomes were not analyzed; the study focused on diagnostic performance rather than prognosis.

Conclusions

Cytology with modified WGS is a reliable LN staging tool for canine MCTs, showing good agreement with histopathology and superior detection of early metastasis compared with routine H&E alone. “Certain metastasis” on cytology is highly specific and strongly confirmatory; including “possible” and “probable” increases sensitivity but reduces specificity and should prompt histologic confirmation. Routine use of toluidine blue on LNs enhances histopathologic detection, and definitive staging should integrate cytology with TBS-augmented histology and appropriate nodal mapping to optimize sentinel LN identification.

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