Immunofluorescent analysis of Connexin 43 (red) in cultured mouse embryonic fibroblasts (MEFs). MEFs were isolated and cultured using the Mouse Embryonic Fibroblast Isolation Kit (Product # 88279). Cells were seeded at 100,000 cells per well in a 24-well plate. At day 7, cells were fixed with 4% paraformaldehyde, permeablilized with 0.1% Triton X-100 in HBSS for 10 minutes at room temperature, and blocked with 3% BSA in PBS (Product # 37525) for 30 minutes at room temperature. Cells were probed with Connexcin 43 polyclonal antibody (Product # PA1-37497) antibody at a dilution of 1:300 and a beta-actin monoclonal antibody at dilution of 1:500 for 2 hours at room temperature or overnight at 4°C, washed with HBSS, and incubated with a DyLight 488-conjugated goat anti-mouse IgG secondary antibody (Product# 35502) and a DyLight 680 goat anti-rabbit IgG secondary antibody (Product # 35568) at dilution of 1:500 for 1 hour at room temperature. Images were taken at 40X magnification on a Carl Zeiss microscope (AxioVision Rel. 4.7).
|Tested species reactivity||Human, Mouse, Rat|
|Host / Isotype||Rabbit / IgG|
|Immunogen||Synthetic peptide derived from a segment of the 3 cytoplasmic domain of rat connexin 43|
|Purification||Antigen affinity chromatography|
|Storage buffer||PBS, pH 7.6, with 1% BSA|
|Contains||<0.1% sodium azide|
|Storage Conditions||4° C, do not freeze|
|Tested Applications||Dilution *|
|Immunohistochemistry (Paraffin) (IHC (P))||1:100|
|Western Blot (WB)||1:25|
* 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.
This antibody is predicted to react with bovine, canine, mouse, porcine and rabbit based on sequence homology.
Heat-mediated antigen retrieval is recommended prior to staining, using a 10mM citrate buffer, pH 6.0, for 10 minutes followed by cooling at room temperature for 20 min. Following antigen retrieval, incubate samples with primary antibody for 10 min at room temperature. A suggested positive control is heart tissue.
Gap junctional intercellular communication is thought to play a key role in development and may also be involved in epilepsy (Aronica et al., 2001). Connexin43 forms gap-junctional channels and regulates the permeability of these gap junctions to small organic molecules. Permeability of Connexin43 is known to be regulated by phosphorylation at Ser368 by protein kinase C (Yogo et al., 2002; Bao et al., 2004a). Phosphorylation of Ser368 by PKC induces a conformational change of Connexin43 that results in a decrease in gap junction permeability (Bao et al., 2004b).
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.