* Suggested working dilutions are given as a guide only. It is recommended that the user titrates the product for use in their own experiment using appropriate negative and positive controls.
1mg/ml BSA, 30% glycerol
0.05% sodium azide
Product Specific Information
MA1-104 detects Oct4/Pou5F1 in Human embryonic carcinomic samples, but not in adult samples (e.g. HeLa cells). This antibody did not show reactivity in a mouse embryonic cell line (i.e. F9 embryonic carcinoma cells).
MA1-104 has been successfully used in FACS, IF, and WB. By Western blot, this antibody detects a band at 40 kDa in human NCCIT and NTERA-2 embryonal carcinoma cells, but not in negative control cells.
Oct4 is a transcription factor which plays a key role in embryonic development, especially during early embryogenesis. Oct4 is one of the four Yamaka factors, necessary for embryonic stem cell pluripotency. A translocation of this gene with the Ewing's sarcoma gene, t(6;22)(p21;q12), has been linked to tumor formation.
Oct4/Pou5F1 Antibody (MA1-104) in WB Oct4_Antibody_MA1-104_Western-Blot.jpg
Western Blot with anti-Oct4/Pou5F1 Monoclonal Antibody [9B7] (MA1-104)
Western blot analysis of Oct4 was performed by loading 75 ug of the indicated whole cell lysates and 5ul of the Lane Marker Reducing Sample Buffer (Product # 39000) per well onto a 4-20% Tris-HCl polyacrylamide gel. Proteins were transferred to a PVDF membrane and blocked with 5% BSA/TBST (Product # 37525) for at least 1 hour. The membrane was probed with an Oct4 monoclonal antibody (Product # MA1-104) at a dilution of 1:1000 overnight at 4°C on a rocking platform, washed in TBS-0.1%Tween 20, and probed with a goat anti-mouse IgG-HRP secondary antibody (Product # 32430) at a dilution of 1:20,000 for at least 1 hour. Chemiluminescent detection was performed using SuperSignal West Dura (Product # 34075). **Note: the antibody is detecting Oct4 in the human embryonal carcinoma cells, but not in negative control HeLa cells. Unknown bands at 80kDa were also observed with this antibody.
Oct4/Pou5F1 Antibody (MA1-104) in IF Oct4_Antibody_MA1-104_Immunofluorescence_1.jpg
Immunofluorescence with anti-Oct4/Pou5F1 Monoclonal Antibody [9B7] (MA1-104)
Immunofluorescent analysis of Oct4 (green) in retinoic acid-treated NCCIT cells. Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 0.3% BSA/TBST (Product # 37525) for 15 minutes at room temperature. Cells were probed with an Oct4 monoclonal antibody (Product # MA1-104) at a dilution of 1:100 for at least 1 hour at room temperature, washed with PBS, and incubated with DyLight 488 goat anti-mouse IgG secondary antibody (Product # 35502) at a dilution of 1:400 for 30 minutes at room temperature. Nuclei (blue) were stained with Hoechst 33342 dye (Product # 62249). Images were taken on a Thermo Scientific ArrayScan at 20X magnification. **Note: the absence of Oct4 staining in a subset of the NCCIT cells suggests differentiation. In addition to the nuclear staining of Oct4, the antibody also shows some cytoplasmic staining.
Oct4/Pou5F1 Antibody (MA1-104) in FACS Oct4_Antibody_MA1-104_Flow-Cytometry.jpg
Flow Cytometry with anti-Oct4/Pou5F1 Monoclonal Antibody [9B7] (MA1-104)
Flow cytometry analysis of Oct4 (red histogram) on human NCCIT cells. Cells were harvested, fixed with 4% formaldehyde, washed with PBS, and incubated with an Oct4 monoclonal antibody (Product # MA1-104) at a 1:100 dilution or PBS alone (blue histogram) for 1 hour on ice. For flow analysis, a 30-minute incubation with DyLight 488 goat anti-mouse IgG secondary antibody (Product # 35502) was performed and 10,000 cells were acquired for each sample.
(This product is for In Vitro experimental use only. Not for resale without express authorization.)