An amount of 1 × 104 CPCs were seeded on dECM-NH or dECM-PH mounted on 96-well plates and cultured for 7 days under standard culture conditions in the same cell culture medium used for hydration. As a control, 1 × 104 cells were seeded on fibronectin-coated wells. Cells were checked daily by an Olympus CKX41 inverted microscope equipped with a Colorview IIIu digital camera (Olympus Corporation, Tokyo, Japan). Beginning at 48 h after seeding, and then every day for one week, cell death rate was assessed using trypan blue exclusion assay adapting a previously described protocol [58 (link),61 (link)]. Specifically, every day cells were detached from a subset of wells in the multi-well plates by incubation with 0.25% trypsin-EDTA solution (Sigma-Aldrich, St. Louis, MO, USA) for 10 min. Detached cells were then stained with trypan blue stain (0.4% in PBS) (Lonza, Walkersville, MD, USA) for 2 min at room temperature and counted using a hemocytometer. The percentage of dead cells and of alive cells over total cells for each time point was expressed as the mean percentages.
Colorview iiiu digital camera
Manufactured by Olympus
Sourced in Japan
The Colorview IIIu is a digital camera designed for laboratory applications. It captures high-quality images with a resolution of up to 5.0 megapixels. The camera features a CMOS image sensor and can provide real-time image previewing.
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Lab products found in correlation
2 protocols using colorview iiiu digital camera
1
Cell Viability Assay on Decellularized Extracellular Matrices
2
Astrocytic Marker Immunohistochemistry in ARC
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To inhibit the endogenous peroxidase activity, brain slices containing ARC were treated with 0.4% H 2 O 2 for 30 min at room temperature (rt). In order to eliminate background staining the slides were incubated in 10% goat serum (Sigma-Aldrich) for 20 min. Immunohistochemical detection of astrocytic markers GFAP and S100β was carried out using a peroxidase-antiperoxidase method (PAP). A set of antibodies (Sigma-Aldrich) and reagents diluted in 0.5M Tris buffer (TBS, pH-7.6) were used. The monoclonal rabbit anti-GFAP antibody and mouse anti-S100β antibody were applied on the slides to reveal astrocytic proteins. The material was incubated in primary antibodies for 24 h at 4 o C. Subsequently, the secondary, species appropriate, monoclonal anti-IgG-peroxidase antibody was used for 1h at rt. The dilutions of antibodies were prepared according to the producer's recommendations. Diaminobenzidine (DAB) was used as a chromogen. Additionally, after immunostaining, the slides were counterstained with Mayer's haematoxylin. A reaction specificity control was performed omitting the primary antibodies and replacing them with normal goat serum. GFAP-IR and S100β-IR in ARC were analysed and photographed under a light microscope BX51 (Tokyo, Japan) using an Olympus Color View IIIu digital camera. On the basis of GFAP-IR and S100β-IR the morphology of the astrocytes was analysed.
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