alpha-Smooth Muscle Actin Overexpression Lysate

Western Blot: alpha-Smooth Muscle Actin Overexpression Lysate [NBL1-07272] - Left-Empty vector transfected control cell lysate (HEK293 cell lysate); Right -Over-expression Lysate for Actin Alpha 2 Smooth Muscle. The theoretical molecular weight is shown here at ~42 kDa.

• Reactivity: Hu
• Applications: WB

alpha-Smooth Muscle Actin Overexpression Lysate Summary

• Description:

  alpha-Smooth Muscle Actin Transient Overexpression Lysate

  
  Expression Host: HEK293T
  
  Plasmid: RC210641
  
  Accession#: NM_001613
  
  Protein Tag: C-MYC/DDK
  
  You will receive 1 vial of lysate (100ug), 1 vial of empty vector negative control (100ug), and 1 vial of 2xSDS sample buffer (250ul). Each vial of cell lysate contains 100ug of total protein (at 1 mg/ml). The 2xSDS Sample Buffer consists of 4% SDS, 125mM Tris-HCl pH6.8, 10% Glycerol, 0.002% Bromophenol blue, 100mM DTT.
• Gene: ACTA2

Applications/Dilutions

• Dilutions:
  • Western Blot
• Application Notes: This product is intended for use as a positive control in Western Blot. Overexpression of the target protein was confirmed using an antibody to DDK (FLAG) epitope tag (NBP1-71705) present on the protein construct.
  
  Each vial of cell lysate contains 100ug of total protein which should be sufficient for 20-50 reactions. Depending on over-expression level, antibody affinity and detection system, some lysates can go as low as 0.1 ug per load. We recommend starting with 5ug of cell lysate. Add an equal amount of cell lysate and 2X SDS Sample buffer and boil the SDS samples for 10 minutes before loading.
• Theoretical MW: 42 kDa.
  Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.

Packaging, Storage & Formulations

• Storage: Store at -80C. Avoid freeze-thaw cycles.
• Buffer: RIPA buffer

Lysate Details for Array

• Type: Overexpression

Notes

HEK293T cells in 10-cm dishes were transiently transfected with a non-lipid polymer transfection reagent specially designed and manufactured for large volume DNA transfection. Transfected cells were cultured for 48hrs before collection. The cells were lysed in modified RIPA buffer (25mM Tris-HCl pH7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 1xProteinase inhibitor cocktail mix, 1mM PMSF and 1mM Na3VO4, and then centrifuged to clarify the lysate. Protein concentration was measured by BCA protein assay kit.

Alternate Names for alpha-Smooth Muscle Actin Overexpression Lysate

• AAT6
• ACTA2
• Actin alpha 2
• actin, alpha 2, smooth muscle, aorta
• actin, aortic smooth muscle
• ACTSA
• ACTVS
• alpha 2 actin
• alpha SMA
• alpha smooth muscle actin
• alpha-actin-2
• alpha-SMA
• alphaSmooth Muscle Actin
• alpha-Smooth Muscle Actin
• a-sma
• Cell growth-inhibiting gene 46 protein
• growth-inhibiting gene 46
• MYMY5
• SMA
• smooth muscle actin

Background

Actin proteins are highly conserved in eukaryotic cells and are involved in cell motility, structure and integrity. Actin and myosin are involved in a myriad of cellular processes including locomotion, secretion, cytoplasmic streaming, phagocytosis and cytokinesis. Actin filaments, along with microtubules, comprise a major component of the cytoskeleton and play essential roles in regulating cell shape and movement. Actin has been found to be expressed in at least six isoforms, each encoded by a separate gene and expressed in a developmentally regulated and tissue-specific manner. Alpha, beta and gamma actin isoforms have been identified. Alpha actins are a major constituent of the contractile apparatus, whereas beta and gamma actins are involved in the regulation of cell motility.

ACTA2 encodes alpha Smooth Muscle Actin and is also known as alpha-actin, alpha-actin-2, aortic smooth muscle or alpha smooth muscle actin (alpha-SMA, SMactin, alpha-SM-actin, ASMA). Alpha Smooth Muscle Actin is frequently used as a marker of smooth muscle differentiation and has a theoretical molecular weight of 42 kDa. Smooth muscle alpha actin is one of a few genes whose expression is predominantly expressed in vascular smooth muscle cells that is used to study myofibroblasts and fibrosis (1). Excessive accumulation of alpha smooth muscle actin-positive (ACTA2+) activated myofibroblasts is observed in Idiopathic pulmonary fibrosis (2). Expression of smooth muscle alpha actin is regulated by cell proliferation and is altered in pathological conditions including atherosclerosis and is common in metastatic cancers (1,3).

References

1. Chakraborty, R., Saddouk, F. Z., Carrao, A. C., Krause, D. S., Greif, D. M., & Martin, K. A. (2019). Promoters to Study Vascular Smooth Muscle. Arterioscler Thromb Vasc Biol, 39(4), 603-612. doi:10.1161/atvbaha.119.312449

2 .El Agha, E., Moiseenko, A., Kheirollahi, V., De Langhe, S., Crnkovic, S., Kwapiszewska, G., . . . Bellusci, S. (2017). Two-Way Conversion between Lipogenic and Myogenic Fibroblastic Phenotypes Marks the Progression and Resolution of Lung Fibrosis. Cell Stem Cell, 20(2), 261-273.e263. doi:10.1016/j.stem.2016.10.004

3. Chen, Y. C., Gonzalez, M. E., Burman, B., Zhao, X., Anwar, T., Tran, M., . . . Kleer, C. G. (2019). Mesenchymal Stem/Stromal Cell Engulfment Reveals Metastatic Advantage in Breast Cancer. Cell Rep, 27(13), 3916-3926.e3915. doi:10.1016/j.celrep.2019.05.084

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Lysates are guaranteed for 6 months from date of receipt.

FAQs for alpha-Smooth Muscle Actin Lysate (NBL1-07272). (Showing 1 - 1 of 1 FAQ).

1. I do not see horse listed as a predicted or confirmed species. Would you be able to confirm if any of your alpha-SMA antibodies will react with equine samples?
   • We have not directly tested equine samples nor do we have cited publications that have tested equine samples. Since alpha-Smooth Muscle Actin is a highly conserved sequence, these antibodies should react with most mammal species.

Bioinformatics

• Gene Symbol: ACTA2