Half-Dozen Scary Details Concerning INK 128

After the cells were seeded in a monolayer, the sample was washed three times with D-Hank's solution, and digested with 0.25% trypsin for 5 min, passaged, and the passaged osteoblast suspension was collected. Detecting osteoblast proliferation by the MTT assay The passaged osteoblast suspension was centrifuged at 279.5 �� g for 10 min, the supernatant was discarded and the sample was washed twice with balanced salt solution. Following counting, the cell density was adjusted to 1��105/ml. The cells were seeded in a 96-well plate, diluted 1:40 in the drug-containing serum with culture medium, and 0.1 ml was added into each well, with 3 compound wells for each group. The Capmatinib concentration samples were placed in the 5% CO2 cell incubator at 37��C for 48 h, and the MTT solution was added prior to reading the colorimetric absorbance of each well, at wavelengths of 570 and 630 nm. Measuring the cycle changes of the osteoblasts with flow cytometry After 48 h, the 4 groups were digested with trypsin, and the cell concentration was adjusted to 5��106/ml, fixed with 70% ethanol for 1 h at 4��C, centrifuged at 279.5 �� g and washed with deionized water. Following this, 1 ml PI-containing DNA fluorescence staining solution was placed into each well, and the samples were dyed for 30 min INK128 at 4��C. Subsequent to filtering, the apoptotic cells in the G1 phase, S phase and the G2/M cell numbers were measured with flow cytometry. Measuring Bax and Bcl-2 expression levels After 2 h of spinal cord injury treatment, the spinal tissue was removed with 1 cm around the injury center, and the sample was immediately placed at ?80��C for extracting the protein. The spinal tissue was homogenized, centrifuged and the supernatant was decanted. The protein content was measured using the bicinchoninic acid assay method. A 15% gel was formulated, loaded and electrophoresis occurred prior to transferring to a membrane, which was closed with skimmed milk powder, and the samples were reacted with the primary Bcl-2 antibody, secondary antibody (goat anti-mouse) and chemiluminescence was performed. Statistical analysis All the experiments were repeated three times, and SPSS Quinapyramine 13.0 (SPSS, Inc., Chicago, IL, USA) was used for the Bonferroni test and one-way analysis of variance, and pairwise comparisons were made between multiple samples. P