Tissue expansion of lung bronchi due to tissue processing for histology - A comparative analysis of paraffin versus frozen sections in a pig model.

Aim: Tissue shrinking due to fixation and processing is well known. However, the degree of shrinking varies significantly with the tissue type as well as the processing method and is not well studied in various tissues. In daily pathological routine workflow, histological specimens from frozen and paraffin sections are performed from the same tissue. In the present study we compared the thickness of bronchus walls obtained from paraffin and frozen sections.

Methods: Pig lungs were frozen in ventilated condition in liquid nitrogen and 36 bronchi were isolated after dissection. Frozen sections of 5 μm thickness were performed and the remaining tissue was fixed and embedded in paraffin after fixation in 4% formalin. Frozen and paraffin sections from the same cutting edge were analysed after haematoxylin and eosin staining by measuring the wall thickness of the bronchi using high power fields of 400-fold magnification. In each bronchus 40 measurements were implemented at different wall positions distributed over the entire wall area. Summed up, in each group 1440 wall measurements were performed in total. Statistical analysis was conducted using the Wilcoxon test and t-test as well as Pearson's correlation coefficient with a significance level at P < 0.05.

Results: The bronchial wall thickness was significantly (p < 0.001) smaller in frozen sections (median: 0.50 mm; min: 0.37 mm; max: 0.97 mm) compared to paraffin sections (median: 0.58 mm; min: 0.35 mm; max: 1.06 mm). The median difference between paraffin and frozen sections was 0.05 mm (min: -0.11 mm; max: 0.22 mm). The wall thickness ratio of both groups was as follows: frozen/paraffin section = 0.8609, thus yielding a difference between paraffin and frozen of 13.91%. High correlation was found between wall thickness measurements on paraffin and frozen sections (R = 0.87, p < 0.001).

Conclusions: The bronchus wall thickness in the frozen section was 14% reduced compared to the paraffin section. In routine pathology as well as in scientific studies these results are of relevance, as airway wall thickness represents a relevant marker for pathological interpretation, especially using CT image techniques.