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(American Journal of Pathology. 2001;158:1997-2003.)
© 2001 American Society for Investigative Pathology

Technical Advance

A Precise and Efficient Stereological Method for Determining Murine Lung Metastasis Volumes

Boye Schnack Nielsen*, Leif Røge Lund*, Ib Jarle Christensen*, Morten Johnsen*{dagger}, Pernille Autzen Usher*, Linda Wulf-Andersen*, Thomas L. Frandsen*, Keld Danø* and Hans Jørgen G. Gundersen{ddagger}

From the Finsen Laboratory,*
Rigshospitalet Copenhagen University Hospital, Copenhagen; the Department of Molecular Cell Biology,{dagger}
University of Copenhagen, Copenhagen; and the Stereological Research Laboratory,{ddagger}
University of Åarhus, Åarhus, Denmark

We have developed a computer-assisted stereological method based on unbiased principles for estimating metastasis volumes in mouse lungs. We evaluated this method using the transplantable Lewis lung carcinoma. Twenty-one days after subcutaneous inoculation of 106 Lewis lung cells into C57BL/6J mice, the mice had primary tumors with an average volume of 2300 mm3. After perfusion fixation, the lungs were removed, embedded in OCT compound, snap-frozen, and processed for stereology. The metastasis volumes were estimated by application of the Cavalieri principle after evaluation of single sections from several evenly distributed tissue levels. The metastasis volume in a group of nine mice varied between 0.01 and 14.4 mm3, with an average of 6.1 mm3. The coefficient of variation was 0.9. The coefficient of error of the volume estimation was determined in five cases and varied from 0.08 to 0.23. Thus, the variation on the metastasis volumes that is achieved by this method contributes very little, 2.5%, to the total variance within the group of mice. In conclusion, we have developed an efficient and unbiased method to determine the metastasis burden in mouse lungs.




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