- 701756 - Invitrogen Antibodies POU5F1 antibody

Submitted references Effects of DMSO on the Pluripotency of Cultured Mouse Embryonic Stem Cells (mESCs).

Sousa MI, Correia B, Branco AF, Rodrigues AS, Ramalho-Santos J
Stem cells international 2020;2020:8835353

Differentiation of Human Embryonic Stem Cells into Neuron, Cholinergic, and Glial Cells.

Hosseini K, Lekholm E, Ahemaiti A, Fredriksson R
Stem cells international 2020;2020:8827874

Four induced pluripotent stem cell lines (TRNDi021-C, TRNDi023-D, TRNDi024-D and TRNDi025-A) generated from fibroblasts of four healthy individuals.

Xu X, Pradhan M, Xu M, Cheng YS, Beers J, Linask KL, Lin Y, Zheng W, Zou J
Stem cell research 2020 Dec;49:102011

Dysfunction of iPSC-derived endothelial cells in human Hutchinson-Gilford progeria syndrome.

Matrone G, Thandavarayan RA, Walther BK, Meng S, Mojiri A, Cooke JP
Cell cycle (Georgetown, Tex.) 2019 Oct;18(19):2495-2508

Generation of two induced pluripotent stem cell lines (NHLBIi001-A and NHLBIi001-B) from a healthy Caucasian female volunteer with normal cardiac function.

Patterson K, Beers J, Linask KL, Lin Y, Hassanzadeh S, Sack MN, Zou J
Stem cell research 2019 Dec;41:101627

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis was performed on whole cell extracts (30 µg lysate) of NTERA-2 (Lane 1), NCCIT (Lane 2), F9 (Lane 3), HEK-293 (Lane 4), A431 (Lane 5) and HeLa (Lane 6). The blots were probed with Anti-OCT4 Recombinant Rabbit Monoclonal Antibody (Product # 701756, 1-2 µg/mL) and detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.4 µg/mL, 1:2500 dilution). A 38 kDa band corresponding to OCT4 was observed only in specific cell lines. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 4-12% Bis-Tris gel (Product # NP0321BOX), XCell SureLock™ Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody following blocking with 5% skimmed milk. Chemiluminescent detection was performed using Pierce™ ECL Western blotting Substrate (Product # 32106).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Knockdown of OCT4 was achieved by transfecting NTERA-2 cells with OCT4 specific siRNA (Silencer® select Product # s10871, Product # s10873). Western blot analysis (Fig a) was performed using Whole cell extract from the OCT4 knock down cells (lane 3), non-specific scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blots were probed with Anti-OCT4 Recombinant Rabbit Polyclonal Antibody (Product # 701756, 2 µg/mL) and Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.4 µg/mL, 1:5000 dilution). Densitometric analysis of this Western blot is shown in histogram (Fig b). Loss of signal upon siRNA mediated knock down confirms that antibody is specific to OCT4.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: For immunofluorescence analysis, NTERA-2 cells were fixed and permeabilized for detection of endogenous OCT4 using Anti- OCT4 Recombinant Rabbit Monoclonal Antibody (Product # 701756, 2 µg/mL) and labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034, 1:2000). Panel a) shows representative cells that were stained for detection and localization of OCT4 protein (green), Panel b) is stained for nuclei (blue) using SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). Panel c) represents cytoskeletal F-actin staining using Alexa Fluor® 555 Rhodamine Phalloidin (Product # R415, 1:300). Panel d) is a composite image of Panels a, b and c clearly demonstrating nuclear localization of OCT4. Panel e) represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Flow Cytometry analysis of endogenous OCT4 was performed on NTERA-2 cells labeled with ABfinity™ Anti-OCT4 Recombinant Rabbit Monoclonal Antibody (Product # 701756, 5 ug/1M cells) or with rabbit isotype control at 0.5 ug/ml and detected with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, (Alexa Fluor® 488 conjugate, Product # A27034, 0.4 ug/ml, 1:2500) as represented by the red and pink histograms respectively. The purple histogram represents unstained control cells and the green histogram represents no-primary-antibody control. A representative of 10,000 cells were acquired and analyzed for each sample using an Attune® Acoustic Focusing Cytometer (4468770).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Enrichment of endogenous OCT4 protein at specific gene loci using Anti-OCT4 Recombinant Rabbit Monoclonal Antibody: Chromatin Immunoprecipitation (ChIP) was performed using Anti-OCT4 Recombinant Rabbit Monoclonal Antibody (Product # 701756, 5 µg) on sheared chromatin from 2 million NTERA2 cells using the "MAGnify ChIP system" kit (Product # 49-2024). Normal Rabbit IgG (1 µg) was used as a negative IP control. The purified DNA was analyzed by 7500 Fast qPCR system (Product # 4351106) with optimized PCR primer pairs for the promoters of the active SOX4, FOXA2, TNC region used as positive control target genes, and the region of the inactive MYOD, SAT2 satellite repeat, used as negative control target gene. Data is presented as fold enrichment of the antibody signal versus the negative control IgG using the comparative CT method.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 3 Exposure to small percentages of DMSO for 48 hours does not negatively affect pluripotency marker expression or the differentiation potential of mESC. (a) Representative immunofluorescence images of plated colonies presenting the pluripotency marker Nanog (red) and counterstained with Hoechst 33342 (blue) (600x magnification) after a 48 h incubation with/without DMSO. (b) RT-PCR analysis for Oct4 , Nanog , Rexo1 , and Essrb mRNA gene expression normalized for endogenous beta-actin ( Actb ) at the 48 h time point. At least four independent experiments were performed, and the results are presented as means +- SEM. (c, d) Relative protein expression of the pluripotency factors Oct4 and Nanog evaluated by Western blot analysis and normalized by beta -actin expression at the 48 h time point. At least three independent experiments were performed, and the results are presented as means +- SEM. (e-g) Embryoid bodies (EBs) were generated from mESCs cultured for 48 h in the presence/absence of DMSO. After the differentiation protocol, every condition was able to generate fully differentiated cultures as shown in (e) by phase-contrast microscopy (100x magnification). (f, g) Western blot analysis and quantification of the protein levels of the three embryonic leaflet markers alpha -FP, SMA, and beta -3-TUB normalized by the expression of the loading control beta -actin. At least four independent experiments were performed, and the results are presented as means +- SEM. Statistic

701756

Antibody data