CD45 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.

CD45, also known as leukocyte common antigen (LCA), T200, or Ly5, is a member C of the class 1 receptor-like protein tyrosine phosphatase (PTPRC) family (1, 2). It is a transmembrane glycoprotein which, due to alternative splicing, has a multiple isoforms with a theoretical molecular weight ranging from 180 - 220 kDa (1, 3-5). Human CD45 is synthesized as a 1281 amino acid sequence consisting of an alternatively spliced extracellular receptor-like region, a cysteine-rich domain, fibronectin-like III repeats, a transmembrane segment, and a cytoplasmic region with tandem protein tyrosine phosphatase (PTP) domains: the membrane proximal domain (D1) and the membrane distal domain (D2) (3, 5). CD45 is expressed on all nucleated hematopoietic cells and their precursors, except mature red blood cells, and is one of the most abundantly-expressed cell-surface glycoproteins, comprising approximately 10% of surface proteins in lymphocytes (3). Functionally, CD45 is essential for development and activation of T-cells and B-cells (1-5). More specifically, CD45 positively regulates antigen receptor signaling and Src-family member kinase activity (1, 3). There are many ways to regulate CD45 phosphatase activity including ligand binding, dimerization, protein interactions, cellular localization, and covalent modifications (3, 6). Ligands for CD45 include pUL11, a transmembrane protein of the cytomegalovirus RL11 (CMV RL11) family, and placental protein 14 (PP14), both of which exclusively bind CD45, and various lectins including CD22, galectin-1, galectin-3, macrophage mannose receptor (MR), and macrophage galactose-type lectin (MGL) (6).

Given its role in immune cell development and activation, CD45 has also been linked to a variety of diseases. The importance of CD45 in immunity has been revealed in human and mouse studies where CD45-deficiency leads to a severe-combined immunodeficiency (SCID) phenotype (2, 3, 6). A CD45-knockout mice study revealed inhibited thymocyte production and poor B-cell response, whereas CD45 activation in mice causes lymphoproliferation and autoantibody production (3). CD45 variants have been associated with altered immune function and autoimmune disorders including multiple sclerosis, systemic lupus erythematosus (SLE), and rheumatoid arthritis (6). Furthermore, altered CD45 expression has been implicated in oncological conditions including chronic lymphatic leukemia, acute lymphatic leukemia, Hodgkin lymphoma, multiple myeloma, and diffuse large B-cell lymphoma (6). Considering its role in autoimmune disorders, immunodeficiency and cancer, CD45 is an ideal therapeutic target (3, 6). The main approaches to control CD45 function is through either selective inhibitors or anti-CD45 antibodies (3).

Alternative names for CD45 includes B220, CD antigen: CD45, CD45 antigen, CD45R, EC 3.1.3.48, GP180, LCA, Leukocyte common antigen, LY5, protein tyrosine phosphatase receptor type c polypeptide, PTPRC, receptor-type tyrosine-protein phosphatase C, T200 Glycoprotein, and T200.

References

1. Trowbridge, I. S., & Thomas, M. L. (1994). CD45: an emerging role as a protein tyrosine phosphatase required for lymphocyte activation and development. Annual review of immunology. https://doi.org/10.1146/annurev.iy.12.040194.000505

2. Andersen, J. N., Jansen, P. G., Echwald, S. M., Mortensen, O. H., Fukada, T., Del Vecchio, R., Tonks, N. K., & Moller, N. P. (2004). A genomic perspective on protein tyrosine phosphatases: gene structure, pseudogenes, and genetic disease linkage. FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

3. Hermiston, M. L., Xu, Z., & Weiss, A. (2003). CD45: a critical regulator of signaling thresholds in immune cells. Annual review of immunology. https://doi.org/10.1146/annurev.immunol.21.120601.140946

4. Tonks, N. K., Diltz, C. D., & Fischer, E. H. (1990). CD45, an integral membrane protein tyrosine phosphatase. Characterization of enzyme activity. The Journal of biological chemistry.

5. Nam, H. J., Poy, F., Saito, H., & Frederick, C. A. (2005). Structural basis for the function and regulation of the receptor protein tyrosine phosphatase CD45. The Journal of experimental medicine. https://doi.org/10.1084/jem.20041890

6. Rheinlander, A., Schraven, B., & Bommhardt, U. (2018). CD45 in human physiology and clinical medicine. Immunology letters. https://doi.org/10.1016/j.imlet.2018.01.009

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 NBL1-14981

• CD45 Antibodies
• CD45 Lysate
• CD45 Proteins
• CD45 Proteins and Enzymes
• CD45 ELISA