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Related Experiment Videos

Interferometric analysis of intrasection and intersection thickness variability associated with cryostat microtomy.

A Anthony, G J Colurso, T M Bocan

    The Histochemical Journal
    |January 1, 1984
    PubMed
    Summary

    Cryostat microtomy of adrenal sections shows thickness variability, with rotary microtomes offering greater consistency than rocking types. This variability is minimal for cytophotometric analyses when using replicate sections and sufficient measurements.

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    Area of Science:

    • Histology
    • Biomedical Engineering
    • Optical Metrology

    Background:

    • Cryostat microtomy is essential for preparing tissue sections for histological analysis.
    • Variability in section thickness can impact the accuracy of quantitative analyses, such as cytophotometry.
    • Understanding and quantifying section thickness variability is crucial for reliable experimental results.

    Purpose of the Study:

    • To assess inter- and intrasection thickness variability in adrenal sections produced by different cryostat microtomes.
    • To compare the performance of rocking and rotary cryostat microtomes regarding section thickness consistency.
    • To determine the impact of section thickness variability on cytophotometric analyses.

    Main Methods:

    • Mach-Zehnder interferometry was employed to measure section thickness.

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  • Adrenal sections were prepared using Bright's Cambridge rocking and Damon rotary cryostat microtomes.
  • Effective section thickness after flash drying was determined relative to microtome settings.
  • Inter- and intrasection variability were quantified for both microtome types.

    • Main Results:

    • A 90% reduction in effective section thickness was observed after flash drying.
    • Linear relationships were found between measured thickness and microtome settings for both instruments.
    • Rotary microtomes exhibited lower inter-section variability (5%) compared to rocking microtomes (11%).
    • Rotary microtomes also showed lower average intrasection variability (4%) than rocking microtomes (7%).

    Conclusions:

    • Rotary cryostat microtomes provide more consistent adrenal section thickness than rocking microtomes.
    • Section thickness variability, while present, is a negligible error source in cytophotometry with appropriate experimental design.
    • The use of replicate sections and adequate measurements per cell or tissue area mitigates the impact of thickness variability.