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Tissue Processing for Stereological Analyses of Lung Structure in Chronic Obstructive Pulmonary Disease
Mohamed Saad1,2, Saleela M Ruwanpura3,4
1Centre for Innate Immunity and Infectious Diseases, Hudson Institute of Medical Research, Clayton, VIC, Australia.
Methods in Molecular Biology (Clifton, N.J.)
|January 12, 2018
Summary
Accurate stereology requires minimizing lung tissue shrinkage. This study details precise fixation, sampling, and embedding methods for quantitative lung structure analysis in mice, crucial for disease research.
Area of Science:
- Pulmonary research
- Stereology
- Animal models
Background:
- Quantitative lung structure data is vital for assessing lung function in development and diseases like COPD.
- Stereology is the gold standard for quantifying 3D lung structures from 2D sections.
- Preventing tissue shrinkage during sample preparation is critical for stereological accuracy.
Purpose of the Study:
- To describe accurate methods for lung tissue fixation, sampling, and embedding.
- To ensure minimal shrinkage for unbiased stereological analysis of mouse lung structures.
- To provide a reproducible protocol for quantitative lung morphology studies.
Main Methods:
- Perfusion fixation using intratracheal instillation of glutaraldehyde/formaldehyde at 25 cm fluid column pressure.
- Systematic uniform random sampling of the fixed lung.
- Embedding of sampled sections in glycol methacrylate to prevent shrinkage.
Main Results:
- The described three-step method (fixation, sampling, embedding) effectively minimizes lung tissue shrinkage.
- This protocol enables theoretically unbiased stereological parameter estimation.
- Accurate quantification of lung structures in mice is achievable.
Conclusions:
- Precise tissue preparation is essential for reliable stereological analysis of lung structures.
- The presented methods ensure the integrity of lung dimensions for accurate quantitative assessment.
- This protocol supports robust research into lung development and disease in mouse models.
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