In the atmospheric scanning electron microscope (ASEM), a 2- to 3-μm layer of the sample resting on a silicon nitride-film window in the base of an open sample dish is imaged, in liquid, at atmospheric pressure, from below by an inverted SEM. Thus, the time-consuming pretreatments generally required for biological samples to withstand the vacuum of a standard electron microscope are avoided. In the present study, various mouse tissues (brain, spinal cord, muscle, heart, lung, liver, kidney, spleen and stomach) were fixed, stained with heavy metals, and visualized in radical scavenger D-glucose solution using the ASEM. While some stains made the nuclei of cells very prominent (platinum-blue, phosphotungstic acid), others also emphasized cell organelles and membranous structures (uranium acetate or the NCMIR method). Notably, symbiotic bacteria were sometimes observed on stomach mucosa. Furthermore, kidney tissue could be stained and successfully imaged in <30 min. Lung and spinal cord tissue from normal mice and mice metastasized with breast cancer cells was also examined. Cancer cells present in lung alveoli and in parts of the spine tissue clearly had larger nuclei than normal cells. The results indicate that the ASEM has the potential to accelerate intraoperative cancer diagnosis, the diagnosis of kidney diseases and pathogen detection. Importantly, in the course of the present study it was possible to increase the observable tissue area by using a new multi-windowed ASEM sample dish and sliding the tissue across its eight windows.

The malformation and disordered remodeling of bones induce various diseases, including osteoporosis. We have developed atmospheric SEM (ASEM) to directly observe aldehyde-fixed bone tissue immersed in radical scavenger buffer without thin sectioning. The short procedure realized the observation of bone mineralization surrounded by many cells and matrices in natural aqueous buffer, decreasing the risk of changes. In osteoblast primary cultures, mineralization was visible without staining. Correlative energy dispersive X-ray spectrometry indicated the formation of calcium phosphate mineral. Fixed bone was sectioned, and the section surface was inspected by ASEM. Mineralized trabeculae of talus spongy bone were directly visible. Associated large and small cells were revealed by phosphotungstic acid staining, suggesting remodeling by bone-absorbing osteoclasts and bone-rebuilding osteoblasts. In tibia, cortical bone layer including dense grains, was bordered by many cells with protrusions. Tissue immuno-EM performed in solution for the first time and anti-cathepsin-K antibody, successfully identified osteoclasts in femur spongy bone. A microfluidics chamber fabricated on the silicon nitride film window of an ASEM dish allowed mineralization to be monitored in vitro ; calcium phosphate crystals as small as 50 nm were imaged. ASEM is expected to be widely applied to study bio-mineralization and bone-remodeling, and to help diagnose bone-related diseases.

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Background Ascariasis is one of the common intestinal infections in developing countries, including China. Migration of Ascaris lumbricoides into the gallbladder is rare, unlike ascariasis of the bile duct and when it does occur, treatment is generally by endoscopic or surgical extraction. Case presentation A 4-year-old Uyghur boy with a history of ascariasis developed intermittent upper abdominal pain for 7 days, was being treated by a local practitioner, and when the pain worsened with yellow sclera for 3 days, he was admitted to our hospital. On physical examination, found out the patient with yellowish skin tone, pale yellow fur on tongue, mild yellow staining of the sclera and tenderness in epigastrium. Laboratory data plus liver function test showed damage of liver function. Abdominal Ultrasonography (USG) and Magnetic resonance cholangiopancreatography (MRCP) showed a long, linear, echogenic structure in the gallbladder neck near to the common bile duct. Once the ascariasis diagnosis was established, he was given conservative treatment of magnesium sulfate with herbal medicine. In 4 days, the patient discharged Ascaris through the stool. Conclusions Conservative treatment of magnesium sulfate with Uyghur medicine treatment according to syndrome differentiation is proven to have curative effect.