GFAP Overexpression Lysate

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.

Glial fibrillary acidic protein (GFAP) is a class III intermediate filament protein that is largely expressed in astrocytes in addition to non-myelinating Schwann cells and glial cells (1,2). Other members of the type III intermediate filament family include desmin, peripherin, and vimentin (2-4). GFAP was first identified in the brains of multiple sclerosis patients (2). Human GFAP protein is 432 amino acids in length with a theoretical molecular weight of ~50 kDa (1,3). GFAP has at least 10 known isoforms, with the most prevalent and common in the brain being GFAP-alpha which is made of a head domain, a rod domain with four coils (1A, 1B, 2A, 2B) joined by linker regions, and a tail domain (1). GFAP is a marker of mature astrocytes, but is also expressed throughout development in both fetal and adult neural stem cells (2). While the exact function of GFAP is still elusive, it has been shown to play a role in cellular processes such as migration, mitosis, structural integrity, and signaling (2).

An increase in GFAP levels is often associated with neuroinflammation which results in the activation and proliferation of astroglia cell population (1,2). GFAP expression is also observed in brains of patients with neurodegenerative diseases including Alzheimer's and Parkinson's, epilepsy disorders, and brain injuries (1-4). Lesion sites associated with neurodegeneration can exhibit an array of gliosis characteristics from glial scarring with reduced astrocyte proliferation to activated, GFAP-positive astrocytes surrounding amyloid plaques (2). Furthermore, the GFAP gene is a target of single nucleotide polymorphisms in the coding region, considered a gain-of-function mutation, characterized by astrocytic inclusions, termed Rosenthal fibers, resulting in Alexander Disease (1-4). GFAP is also a center of many post-translational modifications, such as phosphorylation, which can alter various aspects of filament assembly (1,4).

References

1. Yang, Z., & Wang, K. K. (2015). Glial fibrillary acidic protein: from intermediate filament assembly and gliosis to neurobiomarker. Trends in Neurosciences. https://doi.org/10.1016/j.tins.2015.04.003

2. Hol, E. M., & Capetanaki, Y. (2017). Type III Intermediate Filaments Desmin, Glial Fibrillary Acidic Protein (GFAP), Vimentin, and Peripherin. Cold Spring Harbor Perspectives in Biology. https://doi.org/10.1101/cshperspect.a021642

3. Potokar, M., Morita, M., Wiche, G., & Jorgacevski, J. (2020). The Diversity of Intermediate Filaments in Astrocytes. Cells. https://doi.org/10.3390/cells9071604

4. Viedma-Poyatos, a., Pajares, M. A., & Perez-Sala, D. (2020). Type III intermediate filaments as targets and effectors of electrophiles and oxidants. Redox Biology. https://doi.org/10.1016/j.redox.2020.101582

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.

Array NBL1-11043

• GFAP Antibodies
• GFAP ELISA Kits
• GFAP Lysate
• GFAP Proteins
• GFAP ELISA