Mouse anti gpx4 monoclonal antibody

The paraffin embedded brains were prepared on the third day after ICH with the method described above. For the primary cortical neurons, the cells were seeded into 24-well culture plate at 2 × 10⁵ cells/well for immunofluorescent staining. Tissue sections and cells were incubated overnight at 4 °C with rabbit anti-MAP2 monoclonal antibody (1:2000, Abcam, Cambridge, MA, USA), rabbit anti-NeuN monoclonal antibody (1:100, Abcam, Cambridge, MA, USA), rabbit anti-AIFM2 polyclonal antibody (1:100, Affinity Biosciences, OH, USA), mouse anti-GPX4 monoclonal antibody (1:50, Santa Cruz Biotechnology, CA, USA), or rabbit anti-FACL4 monoclonal antibody (1:100, Abcam, Cambridge, MA, USA). After PBS rinsing, the samples were incubated in the corresponding Alexa Fluor-conjugated secondary antibody (1:500, Abbkine, USA) for 1 h in the dark at room temperature. The anti-fluorescence quenching mounting medium containing DAPI (Solarbio, Beijing, China) was added to the sample area. Then all sections and cells were observed under fluorescence microscope and the positive cells or mean fluorescent intensity in 4 fields per section/wells were counted with Image Pro-Plus 6.0 software.

After various treatments, brains were harvested, paraffin-embedded and then cut into 5 μm thick sections. Sections from the hematoma center and 200 μm on both sides of the hematoma center were used for analysis. Briefly, sections were incubated under 4 °C overnight with primary antibodies against rabbit anti-AIFM2 polyclonal antibody (1:100, Affinity Biosciences, OH, USA), mouse anti-GPX4 monoclonal antibody (1:50, Santa Cruz Biotechnology, CA, USA), rabbit anti-GLP-1R monoclonal antibody (1:100, Abcam, Cambridge, MA, USA), or rabbit anti-FACL4 monoclonal antibody (1:100, Abcam, Cambridge, MA, USA). The sections were then incubated with horseradish peroxidase-combined secondary anti-rabbit or mouse immunoglobulin G antibodies (1:1000, Abcam, Cambridge, MA, USA) for 1 h at room temperature. All staining outcomes were observed using Olympus microscope (Olympus Co., Japan), and the positive cells or area in 4 fields per tissue section from the right brain hemisphere were analyzed with Image Pro-Plus 6.0 software. Analyses were conducted in a blinded manner.