Enhancer activity was assayed using an established mouse transgenic system26 (link),27 (link) using a vector containing a candidate enhancer, a minimal promoter, and the LacZ gene stably integrated into the mouse genome via pronuclear injection. Coordinates of enhancer elements given in Supplementary Table 2 . To visualize LacZ staining on histological sections, embryos were fixed in 4% paraformaldehyde at 4 °C, washed three times in phosphate-buffered saline, embedded in OCT-compound (Sakura Finetek), and cut on a cryostat (Leica, Deerfiled, IL) at 10 µM. Specimens were viewed and photographed using a Leica MZFLIII microscope and DFC300F camera. Images were viewed and stored using Openlab software (Improvision).
Dfc300f camera
Manufactured by Leica
The DFC300F is a Leica digital camera designed for use in laboratory and scientific applications. It features a high-resolution sensor and advanced imaging capabilities to capture detailed, high-quality images for scientific and research purposes.
Automatically generated - may contain errors
2 protocols using dfc300f camera
1
Transgenic Mouse Enhancer Assay
2
Immunofluorescence Imaging of Tight Junctions
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CFS cells were harvested from the monolayer culture using trypsin and seeded onto a 96-well plate at 1 × 10 4 cells/well and incubated at 20 • C until the cells formed a monolayer. The wells were then treated with anti-cTecrem mAb, anti-SCR1-2 pAb, and anti-SCR3-4 pAb at 25 • C for 48 h. The cells were washed with PBS, fixed with 200 µL of methanol. After washing twice with PBS, the wells were blocked by incubating with 1% BSA in 0.05% Tween 20-PBS for 1 h at room temperature. The cells were incubated with a 5 µg/mL concentration of rabbit anti-ZO1 polyclonal antibody (#61-7300, ThermoFisher Scientific, USA) for 1 h at room temperature, followed by 3x PBST and 2x PBS washes. After secondary antibody (anti-rabbit IgG-FITC) treatment and washings with PBST and PBS, the cells were counterstained with 1 µg/mL DAPI for 10 min and observed under the fluorescent microscope. The images were taken with a Leica DFC300F camera, and the fluorescent intensity of the cells were measured using ImageJ software.
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