Cat. # | Size | Qty. | Price |
---|---|---|---|
77986S | 100 µl |
|
REACTIVITY | H |
SENSITIVITY | Endogenous |
MW (kDa) | 70 |
Source/Isotype | Rabbit IgG |
Product Information
Application | Dilution |
---|---|
Western Blotting | 1:1000 |
Immunoprecipitation | 1:100 |
Immunofluorescence (Immunocytochemistry) | 1:400 - 1:1600 |
For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.
NOTE: Please refer to primary antibody product webpage for recommended antibody dilution.
From sample preparation to detection, the reagents you need for your Western Blot are now in one convenient kit: #12957 Western Blotting Application Solutions Kit
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
Load 20 µl onto SDS-PAGE gel (10 cm x 10 cm).
NOTE: Loading of prestained molecular weight markers (#59329, 10 µl/lane) to verify electrotransfer and biotinylated protein ladder (#7727, 10 µl/lane) to determine molecular weights are recommended.
NOTE: Volumes are for 10 cm x 10 cm (100 cm2) of membrane; for different sized membranes, adjust volumes accordingly.
* Avoid repeated exposure to skin.
posted June 2005
revised June 2020
Protocol Id: 10
This protocol is intended for immunoprecipitation of native proteins utilizing Protein A agarose beads for analysis by western immunoblot or kinase activity.
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
10X Cell Lysis Buffer: (#9803) To prepare 10 ml of 1X cell lysis buffer, add 1 ml cell lysis buffer to 9 ml dH2O, mix.
NOTE: Add 1 mM PMSF (#8553) immediately prior to use.
IMPORTANT: Appropriate isotype controls are highly recommended in order to show specific binding in your primary antibody immunoprecipitation. Use Normal Rabbit IgG #2729 for rabbit polyclonal primary antibodies, Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 for rabbit monoclonal primary antibodies, Mouse (G3A1) mAb IgG1 Isotype Control #5415 for mouse monoclonal IgG1 primary antibodies, Mouse (E5Y6Q) mAb IgG2a Isotype Control #61656 for mouse monoclonal IgG2a primary antibodies, Mouse (E7Q5L) mAb IgG2b Isotype Control #53484 for mouse monoclonal IgG2b primary antibodies, and Mouse (E1D5H) mAb IgG3 Isotype Control #37988 for mouse monoclonal IgG3 primary antibodies. Isotype controls should be concentration matched and run alongside the primary antibody samples.
Proceed to one of the following specific set of steps.
NOTE: When using primary antibodies produced in rabbit to detect proteins with a molecular weight in the range of 50 kDa, we recommend using Mouse Anti-Rabbit IgG (Light-Chain Specific) (D4W3E) mAb (#45262) or Mouse Anti-Rabbit IgG (Conformation Specific) (L27A9) mAb (#3678) (or HRP conjugate #5127) as a secondary antibody to minimize interference produced by denatured rabbit heavy chain. For proteins with a molecular weight in the range of 25 kDa, Mouse Anti-Rabbit IgG (Conformation Specific) (L27A9) mAb (#3678) (or HRP conjugate #5127) is recommended to minimize interference produced by denatured mouse light chain.
When using primary antibodies produced in mouse to detect proteins with a molecular weight in the range of 50 kDa, we recommend using Rabbit Anti-Mouse IgG (Light Chain Specific) (D3V2A) mAb (HRP Conjugate) (#58802) as a secondary antibody to minimize interference produced by denatured mouse heavy chain.
posted December 2008
revised October 2021
Protocol Id: 409
Achieve higher quality immunofluorescent images using the efficient and cost-effective, pre-made reagents in our #12727 Immunofluorescence Application Solutions Kit
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
Recommended Fluorochrome-conjugated Anti-Rabbit secondary antibodies:
NOTE: Cells should be grown, treated, fixed and stained directly in multi-well plates, chamber slides or on coverslips.
Aspirate liquid, then cover cells to a depth of 2–3 mm with 4% formaldehyde diluted in 1X PBS.
NOTE: Formaldehyde is toxic, use only in a fume hood.
NOTE: All subsequent incubations should be carried out at room temperature unless otherwise noted in a humid light-tight box or covered dish/plate to prevent drying and fluorochrome fading.
posted November 2006
revised November 2013
Protocol Id: 24
Human
Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Leu290 of human MEF2D protein.
Myocyte enhancer factor 2D (MEF2D) is a member of the MEF2 family of transcription factors. In mammals, there are four MEF2C-related genes (MEF2A, MEF2B, MEF2C, and MEF2D) that encode proteins that exhibit significant amino acid sequence similarity within their DNA-binding domains and, to a lesser extent, throughout the rest of the proteins (1). MEF2 proteins contain a highly conserved N-terminal MADS-box domain, an MEF2 domain, and a more highly variable C-terminal transactivation domain (2). The MEF2 family members were originally described as muscle-specific DNA-binding proteins that recognize MEF2 motifs found within the promoters of many muscle-specific genes (3,4); however, more recently they have been found to play critical roles in other physiological processes, such as heart formation and nervous system development (5,6). As such, alterations in MEF2 protein levels can result in developmental and neurological disorders, as well as other diseases such as liver fibrosis and many types of cancer (7). Specifically, MEF2D expression in hepatocellular carcinoma (HCC) is associated with higher levels of proliferation and poor prognosis (8). MEF2D is also overexpressed in clinical colorectal cancer tissues, where its high expression correlates with metastatic process. Functional investigations show that MEF2D promotes cancer cell invasion and epithelial-mesenchymal transition (EMT) and that it is essential for certain microenvironment signals to induce EMT and metastasis in vivo (9). Alternatively, MEF2D may function as a tumor suppressor in lipo- and leiomyosarcoma, as decreased MEF2D activity results in increased cell proliferation and anchorage-independent growth (10). MEF2D may also act as a tumor suppressor in rhabdomyosarcoma, as loss of MEF2D expression results in inhibition of differentiation, increased cell proliferation, and increased anchorage-independent growth (11).
Explore pathways related to this product.
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