Figure 2. The relationship between the protein concentration of lysates from HeLa cells, treated with Paclitaxel #9807 or hydroxyurea, and the absorbance at 450 nm using the PathScan® Total Vimentin Sandwich ELISA Kit is shown.Learn more about how we get our images
Figure 1. Treatment of HeLa cells with paclitaxel stimulates phosphorylation of vimentin at Ser56, while treatment with hydroxyurea reduces that phosphorylation, as detected by the PathScan® Phospho-Vimentin (Ser56) Sandwich ELISA Kit #7795. Neither treatment affects the levels of total vimentin as detected by the PathScan® Total Vimentin Sandwich ELISA Kit #7789. HeLa cells (80-90% confluent) were treated with 100 nM Paclitaxel #9807 or 4 mM hydroxyurea for 20 hr and lysed. The absorbance readings at 450 nm are shown in the top figure, while the corresponding western blots using Vimentin (D21H3) XP® Rabbit mAb #5741 (left panel) or Phospho-Vimentin (Ser56) (D5H2) Rabbit mAb #7391 (right panel) are shown in the bottom figure.Learn more about how we get our images
|Product Includes||Volume||Solution Color|
|Vimentin Mouse mAb Coated Microwells||96 tests|
|Vimentin Rabbit Detection mAb||1 ea||Green (Lyophilized)|
|Anti-rabbit IgG, HRP-linked Antibody (ELISA Formulated)||1 ea||Red (Lyophilized)|
|Detection Antibody Diluent||11 ml||Green|
|HRP Diluent||11 ml||Red|
|TMB Substrate 7004||11 ml|
|STOP Solution 7002||11 ml|
|Sealing Tape||2 ea|
|ELISA Sample Diluent||25 ml||Blue|
|ELISA Wash Buffer (20X)||25 ml|
|Cell Lysis Buffer (10X) 9803||15 ml|
NOTE: Prepare solutions with purified water.
*NOTE: Some PathScan® ELISA Kits may include HRP-Linked Streptavidin in place of HRP-Linked Antibody.
NOTE: Initial color of positive reaction is blue, which changes to yellow upon addition of STOP Solution.
posted November 2013
The PathScan® Total Vimentin Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of total vimentin protein. A Vimentin Mouse mAb has been coated onto the microwells. After incubation with cell lysates, both phospho- and nonphospho-vimentin protein is captured by the coated antibody. Following extensive washing, a Vimentin Rabbit Detection Antibody is added to detect the captured vimentin protein. Anti-rabbit IgG, HRP-linked antibody is then used to recognize the bound detection antibody. HRP substrate, TMB, is added to develop color. The magnitude of optical density for this developed color is proportional to the quantity of vimentin protein.
Antibodies in kit are custom formulations specific to kit.
The PathScan® Total Vimentin Sandwich ELISA Kit detects endogenous levels of vimentin protein as shown in Figure 1. Kit sensitivity is shown in Figure 2. This kit detects proteins from the indicated species, as determined through in-house testing, but may also detect homologous proteins from other species.
The cytoskeleton consists of three types of cytosolic fibers: microfilaments (actin filaments), intermediate filaments, and microtubules. Major types of intermediate filaments are distinguished by their cell-specific expression: cytokeratins (epithelial cells), glial fibrillary acidic protein (GFAP) (glial cells), desmin (skeletal, visceral, and certain vascular smooth muscle cells), vimentin (mesenchyme origin), and neurofilaments (neurons). GFAP and vimentin form intermediate filaments in astroglial cells and modulate their motility and shape (1). In particular, vimentin filaments are present at early developmental stages, while GFAP filaments are characteristic of differentiated and mature brain astrocytes. Thus, GFAP is commonly used as a marker for intracranial and intraspinal tumors arising from astrocytes (2). Research studies have shown that vimentin is present in sarcomas, but not carcinomas, and its expression is examined in conjunction with that of other markers to distinguish between the two (3). Vimentin's dynamic structural changes and spatial re-organization in response to extracellular stimuli help to coordinate various signaling pathways (4). Phosphorylation of vimentin at Ser56 in smooth muscle cells regulates the structural arrangement of vimentin filaments in response to serotonin (5,6). Remodeling of vimentin and other intermediate filaments is important during lymphocyte adhesion and migration through the endothelium (7).
During mitosis, CDK1 phosphorylates vimentin at Ser56. This phosphorylation provides a PLK binding site for vimentin-PLK interaction. PLK further phosphorylates vimentin at Ser82, which might serve as memory phosphorylation site and play a regulatory role in vimentin filament disassembly (8,9). Additionally, studies using various soft-tissue sarcoma cells have shown that phosphorylation of vimentin at Ser39 by Akt1 enhances cell migration and survival, suggesting that vimentin could be a potential target for soft-tissue sarcoma targeted therapy (10,11).
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc. PathScan is a trademark of Cell Signaling Technology, Inc.
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|7789C||1 Kit (96 assays)||$489.00.0|