Cell Signaling Technology

Product Pathways - Cytoskeletal Signaling

Intermediate Filaments Antibody Sampler Kit #4751

Kit Includes Quantity Applications Reactivity MW (kDa) Isotype
Pan-Keratin (C11) Mouse mAb #4545 40 µl W IHC-P IF-IC IF-P F H R Mk 46-62 Mouse IgG1
Plectin-1 Antibody #2863 40 µl W IF-IC H Mk 400-500 Rabbit
GFAP (GA5) Mouse mAb #3670 40 µl W IHC-P IF-F H M R 50 Mouse IgG1
Vimentin (D21H3) XP® Rabbit mAb #5741 40 µl W IHC-P IF-IC F H M R Mk 57 Rabbit IgG
Desmin (D93F5) XP® Rabbit mAb #5332 40 µl W IF-F IF-IC H M R (Mk) 53 Rabbit IgG
Anti-rabbit IgG, HRP-linked Antibody #7074 100 µl Goat
Anti-mouse IgG, HRP-linked Antibody #7076 100 µl Horse

Applications Key:  W=Western Blotting  IHC-P=Immunohistochemistry (Paraffin)  IF-F=Immunofluorescence (Frozen)  IF-IC=Immunofluorescence (Immunocytochemistry)  IF-P=Immunofluorescence (Paraffin)  F=Flow Cytometry
Reactivity Key:  H=Human  M=Mouse  R=Rat  Mk=Monkey
Species enclosed in parentheses are predicted to react based on 100% sequence homology.

Specificity / Sensitivity

Pan-Keratin (C11) Mouse mAb detects endogenous levels of total keratin 4, 5, 6, 8, 10, 13 and 18 and does not cross-react with other keratins. Plectin-1 Antibody detects endogenous levels of total plectin-1 protein and is expected to cross-react with all isoforms of plectin-1 based on the sequence of the immunogenic peptide. GFAP (GA5) Mouse mAb detects endogenous levels of total GFAP protein. Desmin Antibody detects endogenous levels of total desmin protein. Vimentin (R28) Antibody detects endogenous levels of total vimentin protein.

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell types using Plectin-1 Antibody #2863. *Molecular weights were determined using a pre-stained molecular weight marker (not shown).

Western Blotting

Western Blotting

Western blot analysis of extracts from mouse and rat brain, using GFAP (GA5) Mouse mAb #3670.

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell lines using Pan-Keratin (C11) Mouse mAb #4545.


Western Blotting

Western Blotting

Western blot analysis of extracts from C2C12 cells, rat heart and human heart using Desmin (D93F5) XP® Rabbit mAb #5332.

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell lines using Livin (D61D1) XP® Rabbit mAb #5471.

Description

The Intermediate Filaments Antibody Sampler Kit provides an economical means to evaluate the presence and status of intermediate filaments. The kit includes enough primary and secondary antibody to perform four Western blot experiments per antibody.

Source / Purification

Pan-Keratin (C11) monoclonal antibody is produced by immunizing animals with a cytoskeleton preparation from A431 cells. GFAP (GA5) monoclonal antibody is produced by immunizing animals with native GFAP purified from pig spinal cord. Desmin monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to carboxy terminal residues of human desmin protein. Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to the central sequence of human plectin-1 and residues surrounding Arg28 of human vimentin. Antibodies are purified by peptide affinity chromatography.

Background

The cytoskeleton consists of three types of cytosolic fibers: microfilaments (actin filaments), intermediate filaments and microtubules. Major types of intermediate filaments are distinguished and expressed in particular cell types: 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). 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).Desmin is a myogenic marker expressed in early development that forms a network of filaments that extends across the myofibril and surrounds Z discs. The desmin cytoskeleton provides a connection between myofibrils, organelles and the cytoskeleton (4). Desmin knockout mice develop cardiomyopathy as well as skeletal and smooth muscle defects (5). In humans, desmin related myopathies might be caused by mutations in the corresponding desmin gene or in proteins with which desmin interacts, including αB-crystallin and synemin. Disorganized desmin filaments and the accumulation of protein aggregates comprised predominantly of desmin characterize desmin-related myopathies (reviewed in 6,7).Keratins assemble into filaments, forming heterodimers of an acidic keratin (or type I keratin, keratins 9 to 23) and a basic keratin (or type II keratin, keratins 1 to 8) (8,9). Keratin isoforms demonstrate tissue- and differentiation-specific profiles, which make them useful as biomarkers (8). Mutations in keratin genes are associated with skin disorders, liver and pancreatic diseases, and inflammatory intestinal diseases (10-13).Plectin is a large, widely expressed protein that crosslinks the intermediate filament and actin cytoskeleton, mechanically stabilizing cells and tissues. Plectin also plays a role in the regulation of actin dynamics and acts as a scaffold for signaling molecules (14). It is important in the stabilization of hemidesmosomes, crosslinking them to the intermediate filament network. Plectin has been shown to be involved in several signaling cascades. It signals to PKC by binding to and sequestering RACK1, the receptor for activated C kinase 1 (15,16). Plectin is also involved in the regulation of cytokeratin architecture and cell stress response (16), signaling through the chemokine receptor CXCR4 (17), regulation of AMP-activated protein kinase (AMPK) activity and signaling in mouse myotubes (18).

  1. Eng, L.F. et al. (2000) Neurochem Res 25, 1439-51.
  2. Goebel, H.H. et al. (1987) Acta Histochem Suppl 34, 81-93.
  3. Leader, M. et al. (1987) Histopathology 11, 63-72.
  4. Capetanaki, Y. et al. (2007) Exp Cell Res 313, 2063-76.
  5. Li, Z. et al. (1996) Dev Biol 175, 362-6.
  6. Paulin, D. and Li, Z. (2004) Exp Cell Res 301, 1-7.
  7. Paulin, D. et al. (2004) J Pathol 204, 418-27.
  8. Moll, R. et al. (1982) Cell 31, 11-24.
  9. Chang, L. and Goldman, R.D. (2004) Nat Rev Mol Cell Biol 5, 601-13.
  10. Ramaekers, F.C. and Bosman, F.T. (2004) J Pathol 204, 351-4.
  11. Lane, E.B. and McLean, W.H. (2004) J Pathol 204, 355-66.
  12. Zatloukal, K. et al. (2004) J Pathol 204, 367-76.
  13. Owens, D.W. and Lane, E.B. (2004) J Pathol 204, 377-85.
  14. Wiche, G. (1998) J Cell Sci 111 ( Pt 17), 2477-86.
  15. Osmanagic-Myers, S. and Wiche, G. (2004) J Biol Chem 279, 18701-10.
  16. Osmanagic-Myers, S. et al. (2006) J Cell Biol 174, 557-68.
  17. Ding, Y. et al. (2008) Exp Cell Res 314, 590-602.
  18. Gregor, M. et al. (2006) J Cell Sci 119, 1864-75.

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This product is intended for research purposes only. The product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.

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