Improving efficiency in stereology: a study applying the proportionator and the autodisector on virtual slides

Cell counting in stereology is time‐consuming. The proportionator is a new stereological sampling method combining automatic image analysis and non‐uniform sampling. The autodisector on virtual slides combines automatic generation of disector pairs with the use of digital images. The aim of the study was to investigate the time efficiency of the proportionator and the autodisector on virtual slides compared with traditional methods in a practical application, namely the estimation of osteoclast numbers in paws from mice with experimental arthritis and control mice. Tissue slides were scanned in a digital slide scanner and the autodisector was applied on the obtained virtual tissue slides. Every slide was partitioned into fields of view, and cells were counted in all of them. Based on the original exhaustive data set comprising 100% of fields of view and covering the total section area, a proportionator sampling and a systematic, uniform random sampling were simulated. We found that the proportionator was 50% to 90% more time efficient than systematic, uniform random sampling. The time efficiency of the autodisector on virtual slides was 60% to 100% better than the disector on tissue slides. We conclude that both the proportionator and the autodisector on virtual slides may improve efficiency of cell counting in stereology.

[1]  J R Nyengaard,et al.  The Conneulor: unbiased estimation of connectivity using physical disectors under projection. , 1993, Bone.

[2]  H J Gundersen,et al.  The efficiency of systematic sampling in stereology and its prediction * , 1987, Journal of microscopy.

[3]  D. Horvitz,et al.  A Generalization of Sampling Without Replacement from a Finite Universe , 1952 .

[4]  Clive R Taylor,et al.  From Microscopy to Whole Slide Digital Images: A Century and a Half of Image Analysis , 2011, Applied immunohistochemistry & molecular morphology : AIMM.

[5]  H J G Gundersen,et al.  The smooth fractionator , 2002, Journal of microscopy.

[6]  A. Jara-Lazaro,et al.  Digital pathology: exploring its applications in diagnostic surgical pathology practice , 2010, Pathology.

[7]  W. G. Madow On the Theory of Systematic Sampling, III. Comparison of Centered and Random Start Systematic Sampling , 1953 .

[8]  T. Nomura,et al.  Altered thymic T-cell selection due to a mutation of the ZAP-70 gene causes autoimmune arthritis in mice , 2003, Nature.

[9]  D. C. Sterio The unbiased estimation of number and sizes of arbitrary particles using the disector , 1984, Journal of microscopy.

[10]  J E Gardi,et al.  Automatic sampling for unbiased and efficient stereological estimation using the proportionator in biological studies , 2008, Journal of microscopy.

[11]  T. Suda,et al.  Cytokine production profile of splenocytes derived from zymosan A-treated SKG mice developing arthritis , 2006, Inflammation Research.

[12]  M. N. Murthy Sampling theory and methods. , 1967 .

[13]  T. Thornhill,et al.  Identification of cell types responsible for bone resorption in rheumatoid arthritis and juvenile rheumatoid arthritis. , 1998, The American journal of pathology.

[14]  T. Nomura,et al.  Complement drives Th17 cell differentiation and triggers autoimmune arthritis , 2010, The Journal of experimental medicine.

[15]  J. Nyengaard,et al.  Histological changes in chronic autoimmune SKG-arthritis evaluated by quantitative three-dimensional stereological estimators. , 2011, Clinical and experimental rheumatology.

[16]  W. G. Madow On the Theory of Systematic Sampling, II , 1944 .

[17]  J E Gardi,et al.  Using biased image analysis for improving unbiased stereological number estimation – a pilot simulation study of the smooth fractionator , 2006, Journal of microscopy.

[18]  J. Nyengaard,et al.  Bone Formation and Resorption Are Both Increased in Experimental Autoimmune Arthritis , 2012, PloS one.

[19]  H. Gundersen,et al.  The efficiency of systematic sampling in stereology — reconsidered , 1999, Journal of microscopy.

[20]  J. E. Gardi,et al.  The proportionator: Unbiased stereological estimation using biased automatic image analysis and non-uniform probability proportional to size sampling , 2008, Comput. Biol. Medicine.