Introduction Tumors of circumanal glands are among most common in dogs. Mostly, these are benign tumors – adenomas. However there is also a considerable number of carcinomas and epitheliomas. Differences in biologic behaviour of epitheliomas and carcinomas, just as histopathologic caracterization, profiles of several interesting immunohistochemical stainings are not so far precisely defined or known. This investigation aims at revealing differences in features of benign and malignant types of circumanal gland tumors using several immunohistochemical stainings and determination of fractal dimension (FD). Materials and methods A total of 54 circumanal gland tumors were chosen from the archives of the Department of Veterinary Pathology, University of Zagreb. Precise number of samples was as follows: 15 adenomas, 11 epitheliomas, 21 well differentiated carcinomas, 7 poorly defferentiated carcinomas. A control of 10 samples of normal circumanal gland obtained from necropsies were also included. After precise histologic classification according to WHO, an immunohistochemical staining with seven different markers was performed. Used IHC markers were: vimentin, E-cadherin, N-cadherin, matrix metalloproteinase 9, Ki-67, estrogen receptor α and androgen receptor. The IHC reactions of mentioned markers were scored using conventional methods. Additionaly, vimentin IHC reactions were photographed at two different magnifications -100X and 400X and converted in black and white images thus enhancing the positive vimentin reactions. These images were used for determination of FD applying „box counting method“ and computer software „Fractalyse“. Independently, in order to gain clinical information regarding individual tumor behaviour (reoccurrence of tumors or metastasis, survival after initial diagnosis etc.), a telephone survey of dog owners that were included in the study was conducted. To determine significance of results, conventional statistics were used using Microsoft Excel and Statistica software. Results IHC staining showed differences considering the tumor type. Above all this was pronounced in Introduction Tumors of circumanal glands are among most common in dogs. Mostly, these are benign tumors – adenomas. However there is also a considerable number of carcinomas and epitheliomas. Differences in biologic behaviour of epitheliomas and carcinomas, just as histopathologic caracterization, profiles of several interesting immunohistochemical stainings are not so far precisely defined or known. This investigation aims at revealing differences in features of benign and malignant types of circumanal gland tumors using several immunohistochemical stainings and determination of fractal dimension (FD). Materials and methods A total of 54 circumanal gland tumors were chosen from the archives of the Department of Veterinary Pathology, University of Zagreb. Precise number of samples was as follows: 15 adenomas, 11 epitheliomas, 21 well differentiated carcinomas, 7 poorly defferentiated carcinomas. A control of 10 samples of normal circumanal gland obtained from necropsies were also included. After precise histologic classification according to WHO, an immunohistochemical staining with seven different markers was performed. Used IHC markers were: vimentin, E-cadherin, N-cadherin, matrix metalloproteinase 9, Ki-67, estrogen receptor α and androgen receptor. The IHC reactions of mentioned markers were scored using conventional methods. Additionaly, vimentin IHC reactions were photographed at two different magnifications -100X and 400X and converted in black and white images thus enhancing the positive vimentin reactions. These images were used for determination of FD applying „box counting method“ and computer software „Fractalyse“. Independently, in order to gain clinical information regarding individual tumor behaviour (reoccurrence of tumors or metastasis, survival after initial diagnosis etc.), a telephone survey of dog owners that were included in the study was conducted. To determine significance of results, conventional statistics were used using Microsoft Excel and Statistica software. Results IHC staining showed differences considering the tumor type. Above all this was pronounced in vimentin stain, which was significantly more expressed in malignant types of circumanal gland tumors. Ki-67 proliferative index showed also significant differences between benign and malignant types of circumanal gland tumors, namely the values for indivudual groups (diagnoses) were as follows: normal gland – 5,67% , adenoma – 4,89%, epithelioma – 14,44%, well differentiated carcinoma – 16,44%, poorly differentiated carcinoma – 20,76%. Staining of matrix metalloproteinase 9 and estogen receptor α were showed significant differences only when comparing group of normal circumanal glands and adenomas versus group consisted of epitheliomas, well and poorly differentiated carcinomas. E-cadherin showed very high staining level disregarding the tumor type (or normal circumanal gland). The same is true for N-cadherin, with important difference that this expression was at all time very low. Androgen receptor showed great variety of expression within each of investigated group, but failed to show any significant differences between any tumour types or normal glands. Values of determined FD varied between different circumanal tumors and normal glands. They were as follows: normal circumanal glands - 1,318 (1,372), adenomas - 1,384 (1,408), epitheliomas - 1,547 (1,597), well differentiated carcinomas - 1,569 (1,607), poorly differentiated carcinomas - 1,679 (1,723); [first value is FD determined from 100X magnification micrographs, second (in brackets) is one determined at 400X magnification micrographs]. After application od HSD Tukey test, significant differences (p< 0,05) were determined individually between normal glands or adenomas and epitheliomas, well differentiated carcinomas and poorly differentiated carcinomas. Simuntaneously, Pearson correlation between fractal dimensions and type of circumanal tumor (or normal gland) were between 0,65 and 0,70 for both dimensions (100X and 400X). Results of telephone survey failed to show any significant difference or corellation between tumor behaviour and tumor type, value of FD or staining of applied IHC markers. Discussion and conclusions Above results indicate vimentin IHC staining and determination of FD as a ancillary diagnostic methods of choice when discerning between benign and malignant tumors of circumanal glands. Additional development of method of FD determination may yield a possibility for this tool to discern within benign and malignant circumanal tumors. Ki-67 staining confirms recent reports which show similar results, and indicates significantly higher Ki-67 proliferation index in malignant circumanal tumors. We suggest for future studies involving FD determination in circumanal tumors a more prospective clinical monitoring of dogs with these tumors to avoid problems regarding retrospective surveys of the owners.