Myosin antibody

Antigen: Myosin

Synonyms: DmMLC_C, MyoII, mlc-c, nmELC, DmelCG3201, CG3201, DFNA4, MHC16, MYH17, myosin, FP17425, FLJ13881, FLJ43092, KIAA2034, NMHC-II-C, DKFZp667A1311, MYH1, MYH3, ARABIDOPSIS THALIANA MYOSIN 1, ATM1, MYOSIN

Clonality: Polyclonal

Host: Rabbit

Reactivity: Human, Cow (Bovine), Mouse (Murine), Rat (Rattus)

Application: Immunohistochemistry (IHC), ELISA, Western Blotting (WB), Immunoprecipitation (IP)

Catalog no.: ABIN104491

Additional Information

Immunogen: Human Myosin Light Chain phospho peptide corresponding to a region of the human smooth/non-muscle form of myosin regulatory light chain conjugated to Keyhole Limpet Hemocyanin (KLH).

Format: Liquid (sterile filtered)

Description: Myosin is the major component of thick muscle filaments, and is a long asymmetric molecule containing a globular head and a long tail. The molecule consists of two heavy chains each ~200,000 daltons, and four light chains each ~16,000 - 21,000 daltons. Activation of smooth and cardiac muscle primarily involves pathways which increase calcium and myosin phosphorylation resulting in contraction. Myosin light chain phosphatase acts to regulate muscle contraction by dephosphorylating activated myosin light chain. This antibody is specific for the phosphorylated form of myosin light chain. The selected peptide sequence used to generate the polyclonal antibody is located near the amino terminal end of the polypeptide corresponding to the smooth/non-muscle form of myosin regulatory light chain found in cardiac myocytes in addition to smooth and non-muscle cells. This sequence differs from that of the sarcomeric/cardiac form of myosin regulatory light chain that has a different sequence around the phosphorylation site.

Specificity: This affinity purified antibody is directed against the regulatory light chain of smooth and non-muscle myosin. The antibody is phosphospecific and detect monophosphorylated and diphosphorylated forms of the protein. The product was affinity purified from monospecific antiserum by immunoaffinity purification. Antiserum was first purified against the phosphorylated form of the immunizing peptide. The resultant affinity purified antibody was then cross-adsorbed against the non-phosphorylated form of the immunizing peptide. This phospho specific polyclonal antibody is specific for phosphorylated pS19 of the protein. Reactivity with non-phosphorylated myosin light chain is less than 1% by ELISA. Cross reactivity with myosin light chain from other species has not been determined, however, the sequence of the immunogen is identical in mammalian and avian species. BLAST search analysis was used to determine that the smooth and non-muscle forms of myosin regulatory light chain have identical sequences. Cross reactivity is expected.

Application Details

Application Notes: This phospho specific polyclonal antibody was tested by ELISA, immunohistochemistry and immunoblotting. Immunoblotting was used to show reactivity with unstimulated and stimulated cardiac myocytes. The antibody was also reactive with the phosphorylated form of the immunizing peptide and minimally reactive with the non-phosphorylated form of the immunizing peptide. Although not tested, this antibody is likely functional by immunoprecipitation. Recommended Dilutions: ELISA:1:5,000 - 1:20,000 WB:1:1,000 - 1:5,000 IP:1:100 OTHER: User Optimized

Concentration: 1.0 mg/ml (by UV absorbance at 280 nm)

Purity: This affinity purified antibody is directed against the regulatory light chain of smooth and non-muscle myosin. The antibody is phosphospecific and detects monophosphorylated and diphosphorylated forms of the protein. The product was affinity purified from monospecific antiserum by immunoaffinity purification. Antiserum was first purified against the phosphorylated form of the immunizing peptide. The resultant affinity purified antibody was then cross-adsorbed against the nonphosphorylated form of the immunizing peptide. This phosphospecific polyclonal antibody is specific for the phosphorylated pS19/pS20 form of the protein, depending on the source origin of the protein. Reactivity with non-phosphorylated myosin light chain is less than 1% by ELISA. Cross reactivity is expected with myosin light chain from human and mouse. Reactivity with the protein from other species has not been determined. However, the sequence of the immunogen is nearly identical in mammalian and avian species. BLAST search analysis was used to determine that the smooth and non-muscle forms of myosin regulatory light chain have identical sequences. Cross reactivity is expected.

Buffer: Stabilizer: None. Preservative: 0.01% (w/v) Sodium Azide. Buffer: 0.02 M Potassium Phosphate, 0.15 M Sodium Chloride, pH 7.2.

Storage: Store vial at -20° C prior to opening. Centrifuge product if not completely clear after standing at room temperature. Dilute only prior to immediate use. For extended storage aliquot contents and freeze at -20° C or below. Avoid cycles of freezing and thawing. Expiration date is one (1) year from date of opening.

Research Area: Signaling, Cytoskeleton, Cell/Tissue Markers

Restrictions: For Research Use only

Images

Immunoblotting. Affinity Purified Phosphospecific antibody to Monophosphorylated Regulatory Light Chain of Smooth and Non-muscle Myosin at pS19 was used at a 1:5000 dilution to detect myosin light chain by Western blot. Either 13 or 20 ul of a mouse cardiac myocyte lysate was loaded on a 4-20% Criterion gel for SDS-PAGE. Samples were either mock-treated or CLA treated according as indicated. After washing, a 1:5,000 dilution of HRP conjugated Gt-a-Rabbit IgG (611- 103-122) preceded color development using Amersham's substrate system. Other detection methods will yield similar results.

Immunoblotting. Affinity purified phosphospecific antibody to monophosphorylated regulatory light chain of smooth and non-muscle Myosin at pS19 was used at a 1:1000 dilution to detect myosin light chain by Western blot. Approximately 12 ul of a mouse cardiac myocyte lysate was loaded on a 4-20% Criterion gel for SDS-PAGE. Samples were either mock treated or insulin or CLA treated according as indicated. After washing, a 1:5,000 dilution of HRP conjugated Gta- Rabbit IgG (ABIN102010) preceded color development using Amersham's substrate system. Other detection methods will yield similar results.