125 - Buffered-Lugol, the Solution to diceCT Shrinkage?

Poster Board Number: 125
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Abstract Body : Lugol's iodine solution (I2KI) is used for diffusible iodine contrast-enhanced computed tomography (diceCT) imaging, which allows for 3D visualization of soft tissues. The limitations of I2KI include the substantial shrinkage factor that is speculated to occur due to the hypertonicity of Lugol's iodine. Based on previous results in the literature, osmotic imbalance and further acidification of the solution are indicated as the primary mechanism of drastic shrinkage. An acidic solution promotes the demineralization of bone, increasing its susceptibility to deformation. We used a buffered I2KI (B-Lugol) solution to stabilize acidity and mitigate specimens' tissue shrinkage. This study aimed to quantify the ability of B-Lugol’s solution to stabilize pH imbalances, and reduce tissue shrinkage in diceCT preparations of newborn primates.

We performed an experimental study of newborn Callithrix jacchus (n = 8) under four treatment conditions: 1% Lugol concentration (control), a 1.25% B-Lugol, 3.75% B-Lugol, and 1.25% B-Lugol with sucrose as a pretreatment. To mitigate specimen shrinkage, we preferentially chose low concentrations of Lugol's solutions and refreshed the solutions weekly. Using 3DSlicer, an open-source visualization software, we compared tissue shrinkage of brain, bone, and muscle tissue across treatment conditions. Imaging data from conventional CT before staining and periodic diceCT imaging allowed us to collect longitudinal data for the same tissues every two weeks and track changes in tissue over time.

Even maintaining low concentrations, there was evidence of brain tissue shrinkage in the 1% Lugol treatment group but not uniformly among specimens. After 66 days in 1% Lugol, one specimen had a 10.7% decrease in overall brain tissue. However, a specimen of comparable size exposed for the same period of time, in the 3.75% B-Lugol, only lost 0.81% in brain tissue volume, even when fully stained. Due to the pH stabilization provided by the buffered solution, we were able to use higher concentrations of Lugol, which provided a clear visualization of soft tissue after only 18 days in stain. The same clarity was not visible after 18 days in the 1% non-buffered Lugol treatment. Since maintaining the integrity of specimens by preventing tissue shrinkage is of paramount importance, the use of B-Lugol can be a highly effective tool. However, whereas some structures (e.g., the brain) and mucosal boundaries are optimally visualized using B-Lugol, there is poor discrimination of muscle fibers. Further analysis is necessary to improve muscle fiber visualization while minimizing tissue shrinkage. Additionally, we recommend increasing the volume of solution to specimen ratio to stabilize shrinkage, minimizing acidification, and allowing longer periods of time between refreshing the solution. Identifying the causes of shrinkage and differential effects across tissues is essential for further use of this technique in research.