Nuclear volume estimation using different sampling, measurement and calculation methods.
OBJECTIVE
To study the reliability of volume parameter measured on tissue sections through different sampling, measurement and calculation methods.


METHODS
The largest nuclear profile image under a 100x, NA 1.30 oil immersion objective of primary spermatocytes and spherical spermatoblasts on 11-micron-thick seminiferous tubule sections and tissue images, under a 20x objective, on 4-micron sections were captured. Their volumes were measured and calculated by the five methods provided by the Technology for Image and Graphics Engineering Research cell image analysis system.


RESULTS
The nuclear volumes obtained by nucleator and area equivalent diameter on the largest nuclear profile image were almost the same, including binary images by automated and manual interactive nucleator and grey scale images only by the latter. Nuclear volumes, calculated by random Feret diameter and equivalent diameter of the perimeter, the minimal circumference of the largest nuclear profile binary image, were obviously larger than those of the nucleator and area equivalent diameter. Due to different-sized nuclear slices entrapped in the same section, those nuclear volumes from the seminiferous tubule tissue images were strikingly lower than that of the largest nuclei profile image. The shape factors of primary spermatocytes and spherical spermatoblast nuclei under 100x and 20x objectives were approximately the same.


CONCLUSIONS
The sample preparation, sampling methods and calculation formulas suitable to nuclear form are necessary to obtain reproducible volume parameters.
Xia
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Department of Histology and Embryology, Medical College, Jinan University, Guangzhou, People's Republic of China. lyxzp@jnu.edu.cn
Xia
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D000818:Animals; D002467:Cell Nucleus / Q000502:physiology*; D007091:Image Processing, Computer-Assisted / Q000379:methods*; D008297:Male; D051381:Rats; D017207:Rats, Sprague-Dawley; D015203:Reproducibility of Results; D013048:Specimen Handling; D013087:Spermatids / Q000166:cytology*; D013090:Spermatocytes / Q000166:cytology*