Image-based determination of chord lengths in air-dried lungs

An image-based method was developed in order to determine chord lengths in sections of dog and sheep lungs air-dried at 25 cm H2O transpulmonary pressure. To facilitate image processing, optical contrast in the sections was optimized with respect to section thickness, stain type, stain concentration, staining temperature, staining time and clearing method. Digital processing of images used standard procedures, e.g. thresholding, dilation and thinning, as well as algorithms written to subtract background, delete spots and measure chord lengths. Correlation of image-based vs. manual determination of mean chord length in 17 sections from a sheep lung, stained for optimal contrast yielded an R2 of 0.82 (P < 0.0001). For 95 sections from three dog lungs, stained with lower contrast, R2 was 0.65 (P < 0.0001). Weaker correlations were observed between image-based and manual determinations of the standard deviation, the geometric standard deviation, and the 95th percentile of chord lengths (P < 0.05). The results show that image-based stereology of inflated air-dried lungs can provide valid measures of mean chord length and other statistics of chord length distribution.