Comprehensive Surface Histology of Fresh Resection Margins With Rapid Open-Top Light-Sheet (OTLS) Microscopy
Gan Gao, Dominie Miyasato, Lindsey A. Barner, Robert Serafin, Kevin W. Bishop, Weisi Xie, Adam K. Glaser, Eben L. Rosenthal, Lawrence D. True, Jonathan Liu
Abstract
<italic xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">Objective:</i> For tumor resections, margin status typically correlates with patient survival but positive margin rates are generally high (up to 45% for head and neck cancer). Frozen section analysis (FSA) is often used to intraoperatively assess the margins of excised tissue, but suffers from severe under-sampling of the actual margin surface, inferior image quality, slow turnaround, and tissue destructiveness. <italic xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">Methods:</i> Here, we have developed an imaging workflow to generate <italic xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">en face</i> histologic images of freshly excised surgical margin surfaces based on open-top light-sheet (OTLS) microscopy. Key innovations include (1) the ability to generate false-colored H&E-mimicking images of tissue surfaces stained for < 1 min with a single fluorophore, (2) rapid OTLS surface imaging at a rate of 15 min/cm <sup xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">2</sup> followed by real-time post-processing of datasets within RAM at a rate of 5 min/cm <sup xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">2</sup> , and (3) rapid digital surface extraction to account for topological irregularities at the tissue surface. <italic xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">Results:</i> In addition to the performance metrics listed above, we show that the image quality generated by our rapid surface-histology method approaches that of gold-standard archival histology. <italic xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">Conclusion:</i> OTLS microscopy has the feasibility to provide intraoperative guidance of surgical oncology procedures. <italic xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">Significance:</i> The reported methods can potentially improve tumor-resection procedures, thereby improving patient outcomes and quality of life.