Published: 2016-12-27

Factors affecting the assessment of mitotic count in histopathological sections of tumors: a study of interobserver and intraobserver variability

Navya Narayanan Orchid, Sathi Puthanpurayil


Background: Mitotic count is the most commonly used method of assessing the proliferative activity of a tumor. It is usually done in routine Hematoxylin & Eosin stained sections and is used for classification, grading, prognostication of tumors and sometimes as a decision factor for treatment. There are numerous variables that can influence the mitotic count like delay in fixation, thickness of the section, size of the high power field of the microscopes and so on. This study is designed to assess the relative importance of Intra-observer variability and Inter-observer variability on mitotic counting by keeping all different procedural factors constant.

Methods: Sections from forty cases of breast cancer and twenty cases of high grade lymphoma formed the study material. Mitotic counts done by the principal investigator under standard conditions were compared with counts done by another experienced observer under standard conditions to evaluate the inter-observer variability. The principal investigator will make all counts twice at intervals of one month to assess intra-observer variability. Paired t test and linear regression were the statistical tests used in analysis. Ki-67 immunohistochemistry was also done in sections fixed at different intervals.

Results: Our study showed that inter-observer (p=0.261) and intra-observer variation (p=0.261) is not statistically significant. In case of inter-observer variability the correlation is weak and not significant.

Conclusions: Reproducibility in mitotic counting can be achieved by following a strict morphological criteria as well as a strict counting protocol.


Observer, Mitosis, Proliferation

Full Text:



Graem N, Helweg-Larsen K. Mitotic activity and delay in fixation of tumour tissue. The influence of delay in fixation on mitotic activity of a human osteogenic sarcoma grown in athymic nude mice. Acta Pathol Microbiol Immunol Scand. 1979;87:375-8.

Donhuijsen K, Schmidt U, Hirche H. Changes in mitotic rate and cell cycle fractions caused by delayed fixation. Hum Pathol. 1990;21:709-14.

Donhuijsen K. Mitosis counts: reproducibility and significance in grading of malignancy. Hum Pathol 1986;17:1122-5.

Montironi R, Collan Y, Scarpelli M. Reproducibility of mitotic counts and identification of mitotic figures in malignant glial tumors. Appl Pathol. 1988;6:258-65.

Woosley JT. Measuring cell proliferation. Arch Pathol Lab Med. 1991;115:555-7.

Van Diest PJ, Baak JPA, Matze-Cok P. Reproducibility of mitosis counting in 2469 breast cancer specimens: results from the multicenter morphometric mammary carcinoma project. Hum Pathol. 1992;23:603-7.

Jannink I, Risberg B, van Diest PJ. Heterogeneity of mitotic activity in breast cancer. Histopathology. 1996;29:421-8.

Navya Narayanan O, Jose C, Sathi PP, Maliekkal JI. Effect of delay of fixation on mitotic counts in histopathological sections of tumors. International Journal of Pharma Research and Health Sciences. 2014;2(2):131-7.

Gal R, Rath-Wolfson L, Rosenblatt Y, Halpern M, Schwartz A, Koren R. An improved technique for mitosis counting. Int J Surg Pathol. 2005;13:161-5.

Lester SC, Bose S, Chen YY, Connolly JL, de Baca ME, Fitzgibbons PL, et al. Protocol for the examination of specimens from patients with invasive carcinoma of the breast. Arch Pathol Lab Med. 2009;133:1515-38.

Meyer JS, Cosatto E, Graf HP. Mitotic index of invasive breast carcinoma. Achieving clinically meaningful precision and evaluating tertial cutoffs. Arch Pathol Lab Med. 2009;133:1826-33.

Meyer JS, Alvarez C, Milikowski C, Olson N, Russo I, Russo J. Breast carcinoma malignancy grading by Bloom-Richardson system vs proliferation index: reproducibility of grade and advantages of proliferation index. Mod Pathol. 2005;18(8):1067-78.

Thunnissen FB, Ambergen AW, Koss M, Travis WD, O'Leary TJ, Ellis IO. Mitotic counting in surgical pathology: sampling bias, heterogeneity and statistical uncertainty. Histopathology. 2001;39:1-8.

Frierson HF Jr, Wolber RA, Berean KW, Franquemont DW, Gaffey MJ, Wilbur DC. Interobserver reproducibility of the Nottingham modification of the Bloom and Richardson histologic grading scheme for infiltrating ductal carcinoma.Am J Clin Pathol. 1995;103(2):195-8.