Rapid assessment of prostate core biopsy pathology at the point-of-procedure could provide benefit in a variety of clinical situations. Even with advanced trans-rectal ultrasound guidance and saturation biopsy protocols, prostate cancer can be missed in up to half of all initial biopsy procedures. In addition, collection of tumor specimens for downstream histological, molecular, and genetic analysis is hindered by low tumor yield due to inability to identify prostate cancer grossly. However, current point-of-procedure pathology protocols such as frozen section analysis (FSA) are destructive, and too time- and labor-intensive to be practical or economical. Ex vivo microscopy of the excised specimens, stained with fast-acting fluorescent histology dyes, could be an attractive non-destructive alternative to FSA. In this work, we report the first demonstration of video-rate structured illumination microscopy (VR-SIM) for rapid high-resolution diagnostic imaging of prostate biopsies in realistic point-of-procedure timeframes. Large mosaic images of prostate biopsies stained with acridine orange are rendered in seconds, and contain excellent contrast and detail, exhibiting close correlation with corresponding H&E histology. A clinically-relevant review of VR-SIM images of 34 unfixed and uncut prostate core biopsies by two independent pathologists resulted in an area under the ROC curve (AUC) of 0.82–0.88, with a sensitivity ranging from 63–88% and a specificity ranging from 78–89%. When biopsies contained more than 5% tumor content, the sensitivity improved to 75–92%. The image quality, speed, minimal complexity, and ease of use of VR-SIM could prove to be features in favor of adoption as an alternative to destructive pathology at the point-of-procedure.
Abstract:We report the development of a structured illumination microscopy instrument specifically designed for the requirements for higharea-throughput, optically-sectioned imaging of large, fluorescently-stained tissue specimens. The system achieves optical sectioning frame-rates of up to 33 Hz (and pixel sampling rates of up to 138.4 MHz), by combining a fast, ferroelectric spatial light modulator for pattern generation with the latest large-format, high frame-rate scientific CMOS camera technology. Using a 10X 0.45 NA objective and a 7 mm/sec scan stage, we demonstrate 4.4 cm 2 /min area-throughput rates in bright tissue-simulating phantoms, and 2 cm 2 /min area-throughput rates in thick, highly-absorbing, fluorescentlystained muscle tissue, with 1.3 μm lateral resolution. We demonstrate highcontrast, high-resolution imaging of a fluorescently-stained 30.4 cm 2 bovine muscle specimen in 15 minutes comprising 7.55 gigapixels, demonstrating the feasibility of the approach for gigapixel imaging of large tissues in short timeframes, such as would be needed for intraoperative imaging of tumor resection specimens.
We report video-rate structured illumination microscopy (VR-SIM) as a practical method for high-resolution imaging of whole core-needle biopsies in seconds. Prostate cancer is rapidly and reliably identified in VR-SIM images, suggesting a use for in-procedure biopsy screening.
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