Structured Illumination Microscopy and a Quantitative Image Analysis for the Detection of Positive Margins in a Pre-Clinical Genetically Engineered Mouse Model of Sarcoma.

Intraoperative assessment of surgical margins is critical to ensuring residual tumor does not remain in a patient. Previously, we developed a fluorescence structured illumination microscope (SIM) system with a single-shot field of view (FOV) of 2.1 × 1.6 mm (3.4 mm2) and sub-cellular resolution (4.4 μm). The goal of this study was to test the utility of this technology for the detection of residual disease in a genetically engineered mouse model of sarcoma. Primary soft tissue sarcomas were generated in the hindlimb and after the tumor was surgically removed, the relevant margin was stained with acridine orange (AO), a vital stain that brightly stains cell nuclei and fibrous tissues. The tissues were imaged with the SIM system with the primary goal of visualizing fluorescent features from tumor nuclei. Given the heterogeneity of the background tissue (presence of adipose tissue and muscle), an algorithm known as maximally stable extremal regions (MSER) was optimized and applied to the images to specifically segment nuclear features. A logistic regression model was used to classify a tissue site as positive or negative by calculating area fraction and shape of the segmented features that were present and the resulting receiver operator curve (ROC) was generated by varying the probability threshold. Based on the ROC curves, the model was able to classify tumor and normal tissue with 77% sensitivity and 81% specificity (Youden's index). For an unbiased measure of the model performance, it was applied to a separate validation dataset that resulted in 73% sensitivity and 80% specificity. When this approach was applied to representative whole margins, for a tumor probability threshold of 50%, only 1.2% of all regions from the negative margin exceeded this threshold, while over 14.8% of all regions from the positive margin exceeded this threshold.

Effect of lithotripter focal width on stone comminution in shock wave lithotripsy.

Using a reflector insert, the original HM-3 lithotripter field at 20 kV was altered significantly with the peak positive pressure (p(+)) in the focal plane increased from 49 to 87 MPa while the -6 dB focal width decreased concomitantly from 11 to 4 mm. Using the original reflector, p(+) of 33 MPa with a -6 dB focal width of 18 mm were measured in a pre-focal plane 15-mm proximal to the lithotripter focus. However, the acoustic pulse energy delivered to a 28-mm diameter area around the lithotripter axis was comparable ( approximately 120 mJ). For all three exposure conditions, similar stone comminution ( approximately 70%) was produced in a mesh holder of 15 mm after 250 shocks. In contrast, stone comminution produced by the modified reflector either in a 15-mm finger cot (45%) or in a 30-mm membrane holder (14%) was significantly reduced from the corresponding values (56% and 26%) produced by the original reflector (no statistically significant differences were observed between the focal and pre-focal planes). These observations suggest that a low-pressure/broad focal width lithotripter field will produce better stone comminution than its counterpart with high-pressure/narrow focal width under clinically relevant in vitro comminution conditions.

A low-cost, portable, and quantitative spectral imaging system for application to biological tissues.

The ability of diffuse reflectance spectroscopy to extract quantitative biological composition of tissues has been used to discern tissue types in both pre-clinical and clinical cancer studies. Typically, diffuse reflectance spectroscopy systems are designed for single-point measurements. Clinically, an imaging system would provide valuable spatial information on tissue composition. While it is feasible to build a multiplexed fiber-optic probe based spectral imaging system, these systems suffer from drawbacks with respect to cost and size. To address these we developed a compact and low cost system using a broadband light source with an 8-slot filter wheel for illumination and silicon photodiodes for detection. The spectral imaging system was tested on a set of tissue mimicking liquid phantoms which yielded an optical property extraction accuracy of 6.40 +/- 7.78% for the absorption coefficient (micro(a)) and 11.37 +/- 19.62% for the wavelength-averaged reduced scattering coefficient (micro(s)').