Intermediate Filament Antibodies, MA3-745

Intermediate filaments are a diverse group of proteins that grant structure and function to the cytoskeleton. At about 10 nm in diameter, they are intermediate between the diameters of actin filaments and microtubules, the two other major components of the cytoskeleton. Similar to actin, intermediate filaments maintain cell shape by giving rigidity and bearing tension. Intermediate filaments assist in organizing the internal three-dimensional structure of the cell, anchoring organelles and the nucleus in place, and helping form some cell–cell and cell–matrix junctions. Intermediate filaments are not directly involved in cell movement, and because they are plentiful in cells subjected to mechanical stress, their primary role appears to be strengthening cells and tissues. Intermediate filaments are also structural parts of the nuclear lamina that lines the inside of the nuclear membrane and directs the shape of the nucleus. The six major types of intermediate filaments are distinguished and expressed in particular cell types.

Intermediate filament marker antibodies detect proteins specific to intermediate filaments and can aid in the study of their structure and function. Intermediate Filament marker antibodies can also help elucidate the role or roles a protein may play in a number of tasks that are centered in or influenced by intermediate filaments. Quality Invitrogen intermediate filament marker antibodies are available for a variety of research needs.

Types I and II intermediate filaments consist of two groups of keratins which are expressed in epithelial cells. A keratin filament forms when an acidic (type I) keratin and a basic (type II) keratin bind to each other to form a heterodimer which then associates with other heterodimers. Keratin mutations are linked to skin disorders, liver and pancreatic diseases, and inflammatory intestinal diseases.

Type III intermediate filaments include four proteins: desmin, glial fibrillary acidic protein (GFAP), vimentin, and peripherin. Desmin is expressed in early development and is specifically expressed in muscle cells, where it connects the Z discs of individual contractile elements. This desmin cytoskeleton supplies a connection between myofibrils, organelles and the cytoskeleton. GFAP filaments are found in astrocytes and other glial cells. Differentiated, mature brain astrocytes characteristically contain GFAP filaments, making GFAP a common marker for spinal and cranial tumors originating in astrocytes. Among all intermediate filaments, vimentin is the most widely distributed and can be found in variety of cells including leukocytes, fibroblasts, smooth muscle cells, and blood vessel endothelial cells. Some of its functions include supporting the cellular membranes, keeping some organelles in a fixed place within the cytoplasm, and transmitting membrane receptor signals to the nucleus. Vimentin filaments are also present at early developmental stages of astroglial cells. Peripherin intermediate filaments are expressed in peripheral neurons.

Type IV intermediate filaments consist of neurofilaments, alpha internexin, synemin, and syncoilin. The three neurofilaments (light, medium, and heavy) form the primary intermediate filaments for many kinds of mature neurons. They are especially plentiful in motor neuron axons where they are essential in supporting the long, thin structures. Alpha internexin is expressed at an earlier stage of neuron development. Synemin is found in several cell types and is particularly known to provide a mechanical link between contractile myofibrils of skeletal muscles and the extracellular matrix. Syncoilin is specific to muscle cells.

Nuclear lamins make up the type V intermediate filaments. Instead of functioning as part of the cytoskeleton, nuclear lamins are essential components of the nuclear envelope, helping give shape to the nucleus. Nuclear lamins also help to attach the chromosomes to the nuclear membrane and anchor nuclear pores. Nestin is the only type VI intermediate filament. It is expressed in stem cells of the central nervous system, early in the development of neurons.

Intermediate filament antibody targets

  • Desmin
  • Filamin
  • KRT5
  • KRT6C
  • KRT7
  • KRT8
  • KRT10
  • KRT14
  • KRT18
  • KRT19
  • LMNB1
  • Nestin
  • Vimentin

Featured product data

Immunohistochemistry detection of nestin on human tonsil tissue. 

Immunohistochemistry of nestin on human tonsil tissue. To expose target protein, antigen was retreived using 10 mM sodium citrate followed by microwave treatment for 8–15 minutes. Endogenous peroxidases were blocked in 3% H2O2-methanol for 15 minutes and tissues were blocked in 3% BSA-PBS for 30 minutes at room temperature. Cells were probed with a nestin rabbit polyclonal antibody (Cat. No. MA1-110) at a dilution of 1:100 overnight in a humidified chamber. Tissues were washed in PBST and detection was performed using a secondary antibody conjugated to HRP. DAB staining buffer was applied and tissues were counterstained with hematoxylin and prepped for mounting. Images were taken at 40x magnification.

Immunofluorescent analysis of vimentin shows staining in HEK293 cells.

Immunofluorescent analysis of vimentin shows staining in HEK293 cells. Vimentin staining (green), F-actin staining with phalloidin (red) and nuclei with DAPI (blue) is shown. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were probed without (control) or with or an antibody recognizing vimentin (Cat. No. MA3-745) at a dilution of 1:100-1:200 overnight at 4oC, washed with PBS and incubated with a DyLight 488-conjugated secondary antibody (Cat. No. 35552 for GAR, Cat. No. 35503 for GAM). Images were taken at 60x magnification.

Western blot analysis of nestin using a specific antibody

Western blot analysis of nestin. Analysis was performed by loading whole cell lysates (20 µg) of the indicated samples and 5 µL of a protein ladder per well onto a 3–8% Tris-acetate polyacrylamide gel. Human neural stem cell (derived from MSA iPSCs using Gibco PSC Neural Induction Medium [Cat. No. A164780], passage 4) and HepG2 lysates were prepared in IP lysis buffer (Cat. No. 87787). Rat brain lysate was prepared in reducing sample buffer. Proteins were transferred to a nitrocellulose membrane using a semi-dry blotter, and blocked with 5% milk in TBST for 1 hour at room temperature. Nestin was detected at approx. 225 kD using a nestin monoclonal antibody (Cat. No. MA1-110) at a dilution of 1:1,000 in 5% milk in TBST. Actin loading control was detected using an actin monoclonal antibody (Cat. No. MA1-744) at a dilution of 1:10,000 in 5% milk in TBST. Both membranes were probed overnight at 4oC on a rocking platform, followed by a HRP-conjugated goat anti-mouse IgG secondary antibody (Cat. No. 31430) at a dilution of 1:10,000 for at least 30 minutes. Chemiluminescent detection was performed using Thermo Scientific SuperSignal West Pico Chemiluminescent Substrate (Cat. No. 34080).

Annotated product references

Cat. No. MA5-11883 was used in immunocytochemistry to use synchronized HeLa cells to study the role of polyamines in the cell cycle G1/S and GM/2 phases. The international journal of biochemistry & cell biology (Jun 2013; 45: 1042) "Role of polyamines at the G1/S boundary and G2/M phase of the cell cycle." Yamashita T,Nishimura K,Saiki R,Okudaira H,Tome M,Higashi K,Nakamura M,Terui Y,Fujiwara K,Kashiwagi K,Igarashi K

Cat. No. MA3-745 was used in immunocytochemistry to investigate the role of COX2 in vascular endothelial growth factor production in retinal Muller cells. Experimental eye research (Jul 2010; 91: 34) "Genetic deletion of COX-2 diminishes VEGF production in mouse retinal Müller cells." Yanni SE,McCollum GW,Penn JS

Cat. No. MA5-11883 was used in immunohistochemistry to report on two cases of malignant transformation of adenomyoepithelioma of the breast by a monophasic population. APMIS : acta pathologica, microbiologica, et immunologica Scandinavica (Apr 2013; 121: 272) "Malignant transformation of adenomyoepithelioma of the breast by a monophasic population: a report of two cases and review of literature." Marian C,Boila A,Soanca D,Malau M,Podeanu DM,Resetkova E,Stolnicu S