The use of Beta Actin (AC-15) as a loading control across multiple species

Fri, 02/17/2017 - 14:46

Actin is a fundamental component of the cytoskeleton, where it has the ability to create and break down actin filament formation in response to various cell needs.  Actin has six highly conserved isoforms, however beta and gamma actin are the two isoforms that are highly and ubiquitously expressed in the cell.  For this reason, measuring beta-actin levels has served as a useful control in research experiments in order to have a baseline of protein expression to compare cell manipulations to.  However, beta actin has other implications in scientific research aside from acting as a housekeeping gene, and plays a role in cell motility, development, wound healing, cell division and more.  The following articles used the Beta Actin (AC-15) antibody as a loading control in chicken, human and rat samples in order to illustrate its conservation and reliability when used across species. 

beta-actin antibody

Western Blot: beta-Actin Antibody (AC-15) [NB600-501] - Whole cell extract of human fibroblasts was separated on SDS-PAGE and blotted with Monoclonal Anti-beta-Actin. The antibody was developed with Goat Anti-Mouse IgG, Peroxidase conjugate and AEC substrate. Lanes A: Antibody dilution 1:5,000 B: Negative control (only secondary antibody).

Juhasz et al used a Beta Actin (AC-15) antibody as a loading control in their research of the pituitary adenylate cyclase activating polypeptide (PACAP) pathway in chicken limb bud-derived chondrogenic cells. To begin, chondrifying micromass cell cultures (HDC) established from the cells of four-day-old chicken limb buds were found to have higher expression of cartilage after exposure to PACAP.  Next, these samples were tested in western blot for elevation of collage type X protein levels, where the Beta Actin (AC-15) antibody was used to show that there was no change in ubiquitous actin levels.  Additionally, the Beta Actin (AC-15) antibody was used to confirm their findings that the PAC1 receptor was also elevated in these samples. 

Furthermore, Cheng et al used a Beta Actin (AC-15) antibody as a loading control to confirm that the activation of an AMP-activated protein kinase can alleviate inflammation in human umbilical vein endothelial cells.  First, the consistent Beta Actin (AC-15) antibody results in western blot helped determine that Adenine inhibited the expression of a variety of adhesion molecules through the NF-kB pathway in TNF-alpha stimulated HUVEC cells.  Cheng also used total and phosphorylated versions of the AMPK and ACC proteins with AMPK and ACC total and phosphorylated antibodies as a control against expression change.

Finally, Yang et al used a Beta Actin (AC-15) antibody as a loading control to investigate the effect of programmed cell death 2 on tumor suppression in rat osteosarcoma (OS) development.  The main goal of their research was to explore the role of PDCD2 in the OS model in relation to CD4+/CD8+ T cells.  The Beta Actin (AC-15) antibody was used as a control in western blot in CD4+ and CD8+ cells in OS to confirm that PDCD2 was expressed after they were transplanted with UMR106 cells.  This experiment was the foundation of the remainder of their experiments, where they were able to conclude that PDCD2 is most likely involved in the development of OS. 

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  1. Bunnell TM, Burbach BJ, Shimizu Y, Ervasti JM. β-Actin specifically controls cell growth, migration, and the G-actin pool. [PMID: 21900491]
  2. Juhász T, Szentléleky E, Somogyi CS, Takács R, Dobrosi N, Engler M, Tamás A, Reglődi D, Zákány R. Pituitary Adenylate Cyclase Activating Polypeptide (PACAP) Pathway Is Induced by Mechanical Load and Reduces the Activity of Hedgehog Signaling in Chondrogenic Micromass Cell Cultures. [PMID: 26230691]
  3. Cheng YF, Young GH, Lin JT, Jang HH, Chen CC, Nong JY, Chen PK, Kuo CY, Kao SH, Liang YJ, Chen HM. Activation of AMP-Activated Protein Kinase by Adenine Alleviates TNF-Alpha-Induced Inflammation in Human Umbilical Vein Endothelial Cells. [PMID: 26544976]
  4. Yang Y, Jin Y, Du W. Programmed cell death 2 functions as a tumor suppressor in osteosarcoma. [PMID: 26617804]

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