Figure 2. The relationship between the protein concentration of lysates from HeLa and NTERA-2 cl.D1 cells and the absorbance at 450 nm is shown. Cells were lysed at 90% confluency.
Figure 1. The expression of Oct-4A can only be detected in embryonic cells, such as NTERA-2 cl.D1 cells, but not in non-embryonic cells, such as HeLa cells, as detected by the PathScan® Total Oct-4A Sandwich ELISA Kit. HeLa and NTERA-2 cl.D1 cells (90% confluent) were lysed. The absorbance readings at 450 nm are shown in the top figure, while the corresponding western blot using Oct-4A (C30A3) Rabbit mAb #2840 is shown in the bottom figure.
|Product Includes||Volume||Solution Color|
|Oct-4A Rabbit mAb Coated Microwells||96 tests|
|Oct-4A Mouse Detection mAb||1 ea||Green (Lyophilized)|
|Anti-mouse IgG, HRP-linked Antibody (ELISA Formulated)||1 ea||Red (Lyophilized)|
|Detection Antibody Diluent||11 ml||Green|
|HRP Diluent||11 ml||Red|
|TMB Substrate 7004||11 ml|
|STOP Solution 7002||11 ml|
|Sealing Tape||2 ea|
|ELISA Wash Buffer (20X) 9801||25 ml|
|ELISA Sample Diluent||25 ml||Blue|
|Cell Lysis Buffer (10X) 9803||15 ml|
The PathScan® Total Oct-4A Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of Oct-4A. An Oct-4A Rabbit Antibody has been coated onto the microwells. After incubation with cell lysates, Oct-4A protein is captured by the coated antibody. Following extensive washing, an Oct-4A Mouse Detection Antibody is added to detect the captured Oct-4A protein. Anti-mouse IgG, HRP-linked Antibody is then used to recognize the bound detection antibody. HRP substrate, TMB, is added to develop color. The magnitude of the absorbance for the developed color is proportional to the quantity of Oct-4A.
Antibodies in kit are custom formulations specific to kit.
NOTE: Prepare solutions with purified water.
*NOTE: Some PathScan® ELISA Kits may include HRP-Linked Streptavidin in place of HRP-Linked Antibody.
NOTE: Initial color of positive reaction is blue, which changes to yellow upon addition of STOP Solution.
posted November 2013
Protocol Id: 204
PathScan® Total Oct-4A Sandwich ELISA Kit detects endogenous levels of total Oct-4A protein. This kit detects proteins from the indicated species, as determined through in-house testing, but may also detect homologous proteins from other species.Species Reactivity:
Oct-4 (POU5F1) is a transcription factor highly expressed in undifferentiated embryonic stem cells and embryonic germ cells (1). A network of key factors that includes Oct-4, Nanog, and Sox2 is necessary for the maintenance of pluripotent potential, and downregulation of Oct-4 has been shown to trigger cell differentiation (2,3). Research studies have demonstrated that Oct-4 is a useful germ cell tumor marker (4). Oct-4 exists as two splice variants, Oct-4A and Oct-4B (5). Recent studies have suggested that the Oct-4A isoform has the ability to confer and sustain pluripotency, while Oct-4B may exist in some somatic, non-pluripotent cells (6,7).
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PathScan is a trademark of Cell Signaling Technology, Inc.