Investigation of different wavelengths for scattering-based light sheet microscopy.

Scattering-based light sheet microscopy (sLSM) is a microscopy technique that can visualize cellular morphologic details based on the scattering signal. While sLSM was previously shown to image animal tissues ex vivo at a cellular resolution, the wavelength used was chosen based on other in vivo microscopy technologies rather than through a comparison of the sLSM imaging performance between different wavelengths. In this paper, we report the development of a multi-wavelength sLSM setup that facilitates the investigation of different wavelengths for sLSM imaging. Preliminary results of imaging human anal tissues ex vivo showed that the sLSM setup allowed for comparisons of the cellular imaging performance at the same tissue location between different wavelengths. Both the quantitative analysis of the image contrast and the visual assessment by a pathologist showed that the imaging depth increased with wavelength, and the imaging depth increase was most notable around 600 nm. The preliminary results showed that the multi-wavelength sLSM setup could be useful in identifying the optimal wavelength for the specific tissue type.

Low-cost, chromatic confocal endomicroscope for cellular imaging in vivo.

We have developed a low-cost, chromatic confocal endomicroscope (CCE) that can image a cross-section of the tissue at cellular resolution. In CCE, a custom miniature objective lens was used to focus different wavelengths into different tissue depths. Therefore, each tissue depth was encoded with the wavelength. A custom miniature spectrometer was used to spectrally-disperse light reflected from the tissue and generate cross-sectional confocal images. The CCE prototype had a diameter of 9.5 mm and a length of 68 mm. Measured resolution was high, 2 µm and 4 µm for lateral and axial directions, respectively. Effective field size was 468 µm. Preliminary results showed that CCE can visualize cellular details from cross-sections of the tissue in vivo down to the tissue depth of 100 µm.

Scattering-Based Light-Sheet Microscopy for Rapid Cellular Imaging of Fresh Tissue.

BACKGROUND AND OBJECTIVES: Light-sheet microscopy (LSM) is a novel imaging technology that has been used for imaging fluorescence contrast in basic life science research. In this paper, we have developed a scattering-based LSM (sLSM) for rapidly imaging the cellular morphology of fresh tissues without any exogenous fluorescent dyes. STUDY DESIGN/MATERIALS AND METHODS: In the sLSM device, a thin light sheet with the central wavelength of 834 nm was incident on the tissue obliquely, 45° relative to the tissue surface. The detection optics was configured to map the light sheet-illuminated area onto a two-dimensional imaging sensor. The illumination numerical aperture (NA) was set as 0.0625, and the detection NA 0.3. RESULTS: The sLSM device achieved a light sheet thickness of less than 6.7 µm over 284 µm along the illumination optical axis. The detection optics of the sLSM device had a resolution of 1.8 µm. The sLSM images of the swine kidney ex vivo visualized tubules with similar sizes and shapes to those observed in histopathologic images. The swine duodenum sLSM images revealed cell nuclei and villi architecture in superficial lesions and glands in deeper regions. CONCLUSIONS: The preliminary results suggest that sLSM may have the potential for rapidly examining the freshly-excised tissue ex vivo or intact tissue in vivo at microscopic resolution. Further optimization and performance evaluation of the sLSM technology will be needed in the future. Lasers Surg. Med. © 2020 Wiley Periodicals LLC.

Smartphone epifluorescence microscopy for cellular imaging of fresh tissue in low-resource settings.

Disease diagnosis in low-resource settings can be challenging due to the lack of equipment and trained personnel required for histologic analysis. In this paper, we have developed a smartphone-based epifluorescence microscope (SeFM) for imaging fresh tissues at sub-cellular resolution. SeFM provides similar resolution and field of view (FOV) as those used during histologic analysis. The SeFM device achieved the lateral resolution of 0.57 µm and provided microscopy images over a sample area larger than 500 µm. The material cost was low, approximately $3,000. Preliminary images of human pancreatic tumor specimens clearly visualized cellular details. Quantitative analysis showed that using an excess dose of a chemotherapy drug significantly reduced the tumor-specific fluorescence signal, confirming the specificity of the drug and the detection potential of SeFM.

Low-cost, high-speed near infrared reflectance confocal microscope.

We have developed a low-cost, near-infrared (NIR) reflectance confocal microscope (RCM) to overcome challenges in the imaging depth and speed found in our previously-reported smartphone confocal microscope. In the new NIR RCM device, we have used 840 nm superluminescent LED (sLED) to increase the tissue imaging depth and speed. A new confocal detection optics has been developed to maintain high lateral resolution even when a relatively large slit width was used. The material cost of the NIR RCM device was still low, ~$5,200. The lateral resolution was 1.1 µm and 1.3 µm along the vertical and horizontal directions, respectively. Axial resolution was measured as 11.2 µm. In vivo confocal images of human forearm skin obtained at the imaging speed of 203 frames/sec clearly visualized characteristic epidermal and dermal cellular features of the human skin.

Clinical Profile of Primary Sjogren's Syndrome with Hypokalemic Periodic Paralysis.

Introduction: Primary Sjogren's Syndrome (pSS) with Hypokalemic Periodic Paralysis(HPP) whether an association or a different clinical subset needs review. Methods: Cross-sectional retrospective study of subjects of Primary Sjogren's Syndrome with Hypokalemic Periodic Paralysis(HPP) identified from database maintained at Centre For Rheumatic Diseases, Pune since 1996 with records of over 50000 patients. The diagnosis was clinical. Clinical and investigations data was extracted pertaining to initial examination and follow up. Standard investigations & ELISA, immunoblot and nephelometry to assay autoantibodies (AAb) were done. Results: 16 patients of Primary Sjogren's Syndrome (pSS) with Hypokalemic Periodic Paralysis (HPP) were identified in the period 2000-2014. Presenting feature was HPP in 86% with Dry eye (4%) and Arthralgias (10%) in remaining. Distal Renal Tubular Acidosis was identified in all. All were females with average age of 26 years. Symptomatic ocular sicca noted in 60% & Oral sicca in 50% patients. Other features - Arthralgias (91%), arthritis (42%), mucositis (38%), Neuropathy (30%), skin rash (20%) cytopenias (19%), Erosive arthritis (10%), interstitial lung disease (10%) and Raynaud Phenomenon (10%). 100% were positive for ANA. SSA was positive in 100%, SSB in 50% of patients & Rheumatoid Factor in 70 %. Hypothyroidism was associated in 70% patients. Conclusion: We present a large series of Primary Sjogren's Syndrome with Hypokalemic Periodic Paralysis(HPP) from India. Prominent features of female dominance, younger age of onset and SSA positivity noted in this cohort of patients on Routine clinical and serology phenotype suggests existence of a distinct subset. HPP was presenting feature in majority.