Digital microscope camera

The basement matrix Geltrex™ (Invitrogen, Eugene, OR) was added (30 μl/well) to coat a 96-well plate and allowed to polymerize for 40 min at 37°C. HUVEC cells (5 × 10³) suspended in RPMI medium supplemented with 0.2% of fetal calf serum were plated alone (control) or mixed with 5 μM of Rb44L1 peptide. The cells were incubated at 37°C for 6 h and images were captured with a microscope digital camera (Olympus, Tokyo, Japan). The numbers of pro-angiogenic structures (typically closed compartments or rings formed after endothelial cell sprouting) were counted from 3 different wells.

SA-β-gal assay was performed using a β-galactosidase staining kit (Beyotime, Shanghai, China), cultured cells were fixed at room temperature for 5 min in 2% formaldehyde and 0.2% glutaraldehyde, and then stained with SA-β-gal staining solution at 37°C overnight, and images were captured using a microscope digital camera (Olympus). The percentage SA-β-gal-positive cells were calculated with ImageJ.

Next, 1 cm long samples of the jejunum, ileum, and colon were fixed in 10% formaldehyde solution, dehydrated, and embedded in paraffin wax. The preserved samples were prepared after cutting, installing, and staining with haematoxylin and eosin and/or periodic acid-Schiff and Alcian blue. In total, 10 well-orientated sections of villus-crypt units in the jejunum and ileum were randomly selected, and the villus height and crypt depth were measured by using a light microscope (Olympus, Tokyo, Japan) and a digital microscope camera (Olympus Optical Company, Guangzhou, China). In addition, the number of goblet cells in the ileum and colon was counted using a previously described method [24 (link)].