Primary antibody rabbit anti atp2b4

Uterine samples were collected at these time points ZT4, ZT8, ZT12, ZT16, ZT20, and ZT24, respectively, and were washed in sterile PBS thrice. Thereafter, they were fixed in 4% paraformaldehyde at room temperature (RT) for 20min, after which they were treated with hydrogen peroxide solution (3%) to deactivate the endogenous enzymes. Subsequently, the samples were washed in PBS solution for 5 min, and then a blocking reagent (goat serum) was added at RT for 20 min after which they were incubated with primary antibody rabbit anti-ATP2B4 (Abcam, Cambridge, UK) overnight at 4℃. After the incubation process, the samples were washed and incubated again with fluorescence-labeled secondary antibody at RT for 30 min. After the second incubation process, the samples were further washed in a PBS solution and incubated again for the third time with peroxidase (POD)-labeled streptavidin (DyLight 488) at RT for 30 min. A DAB kit (BBI, Canada) was used for color development at RT for 5 ~ 30 min, which was proceeded by observation, and then photomicrographs were obtained using a light microscope (Nikon Eclipse E100, Japan) equipped with an imaging system (Nikon DS-U3, Japan).
The images obtained were analyzed using Image-Pro Plus 6.0 software (Media Cybernetics, Silver Spring, USA).

Uterine samples were collected at these time points ZT4, ZT8, ZT12, ZT16, ZT20, and ZT24, respectively, and were washed in sterile PBS thrice. Thereafter, they were fixed in 4% paraformaldehyde at room temperature (RT) for 20min, after which they were treated with hydrogen peroxide solution (3%) to deactivate the endogenous enzymes. Subsequently, the samples were washed in PBS solution for 5 min, and then a blocking reagent (goat serum) was added at RT for 20 min after which they were incubated with primary antibody rabbit anti-ATP2B4 (Abcam, Cambridge, UK) overnight at 4℃. After the incubation process, the samples were washed and incubated again with fluorescence-labeled secondary antibody at RT for 30 min. After the second incubation process, the samples were further washed in a PBS solution and incubated again for the third time with peroxidase (POD)-labeled streptavidin (DyLight 488) at RT for 30 min. A DAB kit (BBI, Canada) was used for color development at RT for 5 ~ 30 min, which was proceeded by observation, and then photomicrographs were obtained using a light microscope (Nikon Eclipse E100, Japan) equipped with an imaging system (Nikon DS-U3, Japan).
The images obtained were analyzed using Image-Pro Plus 6.0 software (Media Cybernetics, Silver Spring, USA).