REACTIVITY | SENSITIVITY | MW (kDa) | Isotype |
---|---|---|---|
H M R | Endogenous | 42 | Mouse IgG |
Western blot analysis of extracts from various tissues using α-Smooth Muscle Actin (1A4) Mouse mAb (IHC Formulated) (upper) and GAPDH (D16H11) XP® Rabbit mAb #5174 (lower). As expected, skeletal muscle samples are negative for α-smooth muscle actin.
Learn more about how we get our images.Immunohistochemical analysis of paraffin-embedded human ductal carcinoma of the breast using α-Smooth Muscle Actin (1A4) Mouse mAb (IHC Formulated).
Learn more about how we get our images.Immunohistochemical analysis of paraffin-embedded human colon carcinoma using α-Smooth Muscle Actin (1A4) Mouse mAb (IHC Formulated).
Learn more about how we get our images.Immunohistochemical analysis of paraffin-embedded normal human colon using α-Smooth Muscle Actin (1A4) Mouse mAb (IHC Formulated).
Learn more about how we get our images.Immunohistochemical analysis of paraffin-embedded human Non-Hodgkin's lymphoma using α-Smooth Muscle Actin (1A4) Mouse mAb (IHC Formulated).
Learn more about how we get our images.Immunohistochemical analysis of paraffin-embedded human prostate carcinoma using α-Smooth Muscle Actin (1A4) Mouse mAb (IHC Formulated).
Learn more about how we get our images.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 datasheet or product webpage for recommended antibody dilution.
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 (#13953, 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 November 2013
Reprobing of an existing membrane is a convenient means to immunoblot for multiple proteins independently when only a limited amount of sample is available. It should be noted that for the best possible results a fresh blot is always recommended. Reprobing can be a valuable method but with each reprobing of a blot there is potential for increased background signal. Additionally, it is recommended that you verify the removal of the first antibody complex prior to reprobing so that signal attributed to binding of the new antibody is not leftover signal from the first immunoblotting experiment. This can be done by re-exposing the blot to ECL reagents and making sure there is no signal prior to adding the next primary antibody.
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalently purified water.
posted June 2005
revised June 2016
Protocol Id: 262
This protocol is intended for immunoprecipitation of native proteins utilizing Protein G agarose beads for subsequent 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.
Proceed to one of the following specific set of steps.
NOTE: To minimize masking caused by denatured IgG heavy chains (~50 kDa), we recommend using Mouse Anti-Rabbit IgG (Light-Chain Specific) (L57A3) mAb (#3677) or Mouse Anti-Rabbit IgG (Conformation Specific) (L27A9) mAb (#3678) (or HRP conjugate #5127). To minimize masking caused by denatured IgG light chains (~25 kDa), we recommend using Mouse Anti-Rabbit IgG (Conformation Specific) (L27A9) mAb (#3678) (or HRP conjugate #5127).
posted October 2016
revised April 2018
Protocol Id: 1244
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
NOTE: Do not allow slides to dry at any time during this procedure.
For Citrate: Heat slides in a microwave submersed in 1X citrate unmasking solution until boiling is initiated; follow with 10 min at a sub-boiling temperature (95°-98°C). Cool slides on bench top for 30 min.
posted February 2010
revised March 2016
Protocol Id: 280
Application | Dilutions |
---|---|
Western Blotting | 1:1000 |
Immunoprecipitation | 1:100 |
Immunohistochemistry (Paraffin) | 1:250 |
Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.
α-Smooth Muscle Actin (1A4) Mouse mAb (IHC Formulated) recognizes endogenous levels of total α-smooth muscle actin protein.
Human, Mouse, Rat
Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues near the amino terminus of human α-smooth muscle protein.
Actin proteins are major components of the eukaryotic cytoskeleton. At least six vertebrate actin isoforms have been identified. The cytoplasmic β- and γ-actin proteins are referred to as “non-muscle” actin proteins as they are predominantly expressed in non-muscle cells where they control cell structure and motility (1). The α-cardiac and α-skeletal actin proteins are expressed in striated cardiac and skeletal muscles, respectively. The smooth muscle α-actin and γ-actin proteins are found primarily in vascular smooth muscle and enteric smooth muscle, respectively. The α-smooth muscle actin (ACTA2) is also known as aortic smooth muscle actin. These actin isoforms regulate the contractile potential of muscle cells (1).
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc. SignalStain is a trademark of Cell Signaling Technology, Inc. XP is a registered trademark of Cell Signaling Technology, Inc. Tween is a registered trademark of ICI Americas, Inc.
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Product # | Size | Price |
---|---|---|
56856S | 100 µl | N/A |
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