Anti-Actin Antibody, clone C4

Actins are ubiquitous eukaryotic proteins that serve as a multi-functional, basic building blocks of cytoskeletal microfilaments. They play critical roles in a wide range of cellular processes, including cell division, cell migration, chromatin remodeling, trascriptional regulation and vesicle trafficking. These funstions are attributed to their ability to form filaments, which can quickly assemble and disassemble depending upon the needs of the cell. At least six different actin types have been reported in mammals. Although actins show about 90% overall sequence homology, isoforms do not show spatial, temporal and tissue-specific expression patterns and only 50-60% homology is found in their 18 N-terminal residues. Beta and gamma-actins, also known as cytoplasmic actins, are highly conserved in higher animals and are predominantly expressed in non-muscle cells where they control cell structure. Exocytosis, and motility. They are nearly identical proteins and differ only in four amino acids at the N-terminal region. The other four actin isoforms are typically found in specific adult muscle tissue types. Alpha-cardiac and alpha-skeletal actins are expressed in striated cardiac and skeletal muscles, respectively. Alpha and gamma actins are primarily found in vascular smooth muscle and enteric smooth muscles, respectively. It has been shown that under calcium-bound conditions, beta-actin exhibits more dynamic behavior than gamma-actin with higher rates of polymerization and depolymerization. Also, beta- and gamma-actins can readily copolymerize, and the resulting filaments exhibit polymerization and depolymerization rates that vary depending on the ration of beta- to gamma-actin (Lessard, JL.,et al.(1988). Cell Motility Cytoskeleton 10(3); 349-362.

MAB1501R is a pan-actin antibody that binds to an epitope in a highly conserved region of actin; therefore, this antibody reacts with all six isoforms of vertabrate actin (Lessard, 1988). Reacts with both globular (G) and fillimentous (F) forms of actin and does not interfere with actin polymerization to form filaments, at a ratio as high as one antibody per two actin monomers. However, this antibody does increase the extent of polymerization when used at a lower ratio of antibody to actin. In addition to labeling myotubes, anti-actin stains myoblasts and fibroblasts. Although clone C4 is prepared as an antibody to chicken gizzard muscles actin, it reacts with actins from all vertebrates, as well as with Dictyostelium discoideum and Physarum polycephalum actins (Lessard, 1988).

To date, all animal species and cell types with an actin form react by indirect immunofluorescence or immunoblot, including plant actin.

Immunocytochemistry:
10 μg/mL dilution from a previous lot was used (methanol fixed mouse 3T3 cells).

Immunohistochemistry:
10μg/mL dilution from a previous lot was used for paraffin embedded, 4% formaldehyde, 3% glutaraldehyde, sodium cacodylate treated sections {see Luciano, L et al. 2003}.

ELISA:
A previous lot was shown to be strongly reactive with the cytoplasmic actin and shows a significant binding to gizzard, skeletal, arterial and cardiac actins. Also shows a significant binding to both Dictyostelium discoidum and Physarum polycephalum.

Western blot:
1-20 µg/ml. On muscle homogenates subject to SDS-PAGE, reacts relatively uniformly with a 43 kD protein present in skeletal, cardiac, gizzard and aorta tissues. Appears to react with all isoforms of actin found in these preparations and shows a strong reaction with the alpha-actin found in skeletal, cardiac, and arterial muscle (Otey, 1987).

Immunohistochemistry:
10µg/mL for paraffin embedded, 4% formaldehyde, 3% glutaraldehyde, sodium cacodylate treated sections {see Luciano, L et al. 2003}.

Optimal working dilutions must be determined by end user.

This Anti-Actin Antibody, clone C4 is validated for use in ELISA, IC, IH, IH(P), WB for the detection of Actin and has published in over 70 citations.

Routinely evaluated by Western Blot on HeLa lysate.

Western Blot Analysis:
1:1000 dilution of this lot detected actin on 10 μg of HeLa lysate.

43 kDa

Protein G Purified in 0.1M Tris-Glycine (pH 7.4) 150mM NaCl with 0.05% NaN3.

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