Immunofluorescence analysis of Oct4/Pou5F1 was done on 70% confluent log phase NTREA-2 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with Oct4/Pou5F1 (9B7) Mouse Monoclonal Antibody (MA1104) at 2ug/mL in 0.1% BSA and incubated for 3 hours at room temperature and then labeled with Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A28175) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (S36938). F-actin (Panel c: red) was stained with Alexa Fluor® 555 Rhodamine Phalloidin (Product # R415, 1:300). Panel d is a merged image showing nuclear localization. Panel e is a no primary antibody control. The images were captured at 60X magnification.
|Tested species reactivity||Human, Mouse|
|Host / Isotype||Mouse / IgG|
|Immunogen||Full length human recombinant protein of human POU5F1 produced in E.coli.|
|Storage buffer||PBS with 1mg/ml BSA, 30% glycerol|
|Contains||0.05% sodium azide|
|Tested Applications||Dilution *|
|Flow Cytometry (Flow)||3-5 µg/10^6 cells|
|Immunocytochemistry (ICC)||1-2 µg/ml|
|Immunofluorescence (IF)||1-2 µg/ml|
|Western Blot (WB)||1:500 - 1:5,000|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
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.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.