Objective: To detect and reconstruct the 3D volume of permanently damaged tissue after thermal ablation.

Methods: TriphenylTetrazolium Chloride (TTC) staining of tissue sections has been used for viable tissue identification. Presented methodology employs TTC staining in 3D quantitative analysis of the extent of thermally ablated tissue in animal models. Ablated regions and approximately 10 mm of adjacent tissue (swine breast) were harvested and immediately embedded in HistOmer, an alginate cold polymer polymerizing at 26 ºC within 60 seconds. HistOmer-embedded blocks were sliced in 4-mm sections and TTC stained for viability for 1 hour. In viable tissue, the TTC is reduced by the mitochondrial enzyme succinate dehydrogenase to red formazan, which can be optically detected. Stained tissue sections were then immediately fixed in 10% neutral buffered formalin and photographed. Six millimeter punch biopsies of tissue specimens from the ablated and viable regions (per the TTC) were collected for histological observations.
A semi-automated image processing algorithm was developed in Matlab to segment the viable (red-tinted) tissue at image data. Data with normalized dynamical range were converted into hue-saturation-value [h,s,v] representation. Pixels with specific [h,s,v] were searched in radial directions from original ablation-probe-position to follow expected growth of ablated region. Optimal [h,s,v] thresholds were determined based on comparison of segmentation results with visual evaluation of image data. Following segmentation, a Matlab script registered individual slices of tissue to each other using fiducial markers (sliced plastic rods embedded in tissue). This procedure corrected for tissue shrinkage and spatial shifts due to the ablation and post-treatment tissue processing. A three-dimensional volume of the extent of ablation was then reconstructed and statistically analyzed.

Results: Optimal performance of the image segmentation algorithm was achieved when the [h,s,v] thresholds were set to h = (22º,40º), s = (0%,80%), and v = (0%,100%). Areas of ablated regions were delineated from viable tissue for each section. The histological examinations, within the ablated and viable regions, were in agreement with the TTC staining. Volume of the ablated tissue was calculated as function of the ablation parameters.

Conclusions: Volume of thermally ablated tissue can be reliably reconstructed from TTC stained tissue sections.