Table 2: Clinical and histological subtypes of human salivary glands carcinomas ex-pleomorphic adenomas (M – male; F – female) Case Age (years) Gender Gland Table 4: Clinical characterization of malignant mixed tumours of canine mammary glands Case Age Gender BMC Cancer 2007, 7:218 http://www.biomedcentral.com/1471-2407/7/218 Localization Thoracic-cranial Thoracic-caudal Abdominal-cranial Inguinal Thoracic-caudal Inguinal Inguinal Thoracic-cranial Thoracic-cranial Abdominal-cranial 1 2 3 4 5 6 7 8 9 10 1 2 3 4 5 6 7 8 9 10 33 22 71 53 43 02 65 70 92 66 5 7 9 4 6 3 7 5 7 8 F F M F M M F M F M minor (palate) submandibular parotid minor (palate) Parotid parotid parotid minor (palate) submandibular parotid Histological Subtype adenocarcinoma NOS adenocarcinoma NOS undifferentiated carcinoma myoepithelial carcinoma myoepithelial carcinoma myoepithelial carcinoma adenoid cystic carcinoma (solid) adenoid cystic carcinoma (tubular) polymorphous low- grade adenocarcinoma mucoepidermoid carcinoma ised in xilol and dehydrated in decreasing alcohol concen- trations. Next, they were submitted to antigenic retrieval (Target Retrieval Solution, pH 6.0, DakoCytomation, Carpinteria, USA) and endogenous peroxidase blocking (3% hydrogen peroxide in methanol). After incubation with primary antibodies (Table 5) and amplification (Ultra Vision Large Volume Detection System, Lab Vision, Fremont, USA), the reaction was revealed with diami- nobenzidine as chromogen and Mayer haematoxylin as contrast. As positive controls, sections of normal human salivary and mammary glands with previously recognized positivity for the antigens studied were used. Substitution of primary antibody for normal human serum constituted the negative control. Finally, morphological analysis of staining was per- formed, and then a semiquantitative protocol was Table 3: Clinical characterization of benign mixed tumours of canine mammaryglands Case Age Gender F F F F F F F F F F Localization Inguinal Thoracic-cranial Abdominal-caudal Abdominal-caudal Thoracic-cranial Inguinal Abdominal-caudal Inguinal Inguinal Inguinal 1 2 3 4 5 6 7 8 9 10 7 13 5 8 9 8 9 9 6 4 F F F F F F F F F F employed to segregate the cases. For this latter purpose the entire available tumoural tissue in the sections was evalu- ated. Next, it was determined whether the relative number of positive neoplastic cells was superior ("positive cases") or inferior ("negative cases") to 5% (for the analysis of p63 and estrogen receptor) [37,38] or 10% (cytokeratins, vimentin, E-cadherin, β-catenin) [29,32] from all of the neoplastic cells in the histological sections evaluated. Statistical analysis Frequency of positive immunostaining between the four groups of lesions was evaluated by Fisher's exact test with values of p < 0.05 considered statistically significant. Probability of α-error inferior to 5% was confirmed to be significant. Results In both species it was observed that the benign tumours occur in the younger individuals' group, while the malig- nant tumours are more frequent in older individuals' group. A slight predominance of female patients (six cases) was observed for PA, while a homogeneous distri- bution was observed among those patients with Ca ex-PA. In dogs, all lesions affected females. Histomorphological comparative illustrations are exem- plified in Figure 1. In general, both benign tumours pre- sented formation of ductal structures and also cells with myoepithelial features, arranged in solid aggregations, Table 5: Primary antibodies, resources and dilutions used in immunohistochemical assays Antibody Anti-pan-cytokeratin Anti-vimentin Anti-p63 Anti-β-catenin Anti-E-cadherin Anti-estrogen receptor Clone Resource Dilution NCL-AE1/AE3 V9 4A4 E-5 4A2C7 CC4–5 Novocastra DAKO Santa Cruz Santa Cruz Zymed Novocastra 1:100 1:50 1:100 1:400 1:40 1:50 Page 3 of 9 (page number not for citation purposes)