Human cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) is a member of the IgG superfamily. It encodes a 223 amino acid protein and possesses a single extracellular V domain, a transmembrane domain, and a cytoplasmic domain. CTLA-4 is ~30% homologous with CD28, and both bind to the same ligands (B7.1/CD80 and B7.2/CD86) although CTLA-4 binds with higher affinity than CD28. Parallel recognition of a specific MHC-peptide complex by the T-cell receptor and of CD80 or CD86 by the costimulatory receptor CD28 results in T-cell activation, cytokine production, proliferation, and differentiation. After the T cells' activation process and upregulation of CTLA-4, co-ligation of the TCR to MHC-peptide complex and CTLA-4 to its ligands results in cell cycle arrest and ends T cell activation. It has been suggested that the negative regulatory mechanism of CTLA-4 is mediated through trans-endocytosis of CD80 and CD86 (capture of ligands from opposing cells). The ligands are removed from the original cell and degraded inside of CTLA-4-expressing cells. As a result the action of CD28 is impaired. Alternative splice variants of CTLA-4 have been described in mouse and human that have transcripts that skip exon 2 (ligand-binding domain) or exon 3 (transmembrane domain). The ligand independent CTLA-4 is expressed in mice, but not in humans, and soluble CTLA-4 (sCTLA-4) is expressed in mice and humans. CTLA-4 is constitutively expressed by Tregs, and they are an important source of sCTLA-4. It has been suggested that sCTLA-4 potentiates regulatory T cell function. High levels of sCTLA-4 in Tregs have been associated with type 1 diabetes (T1D) and other autoimmune diseases, such as Grave's disease and myasthenia gravis. Also, CTLA-4 has been linked with proliferation and survival of chronic lymphocytic leukemia. CTLA-4 is a target for therapeutic intervention; in fact, a monoclonal antibody against CTLA-4 (Ipilimumab) has been approved for the treatment of melanoma.Product Details
- Human CTLA-4, amino acids (Ala37-Phe162) (Accession# BC070162), was expressed in CHO cells, with Human IgG-Fc at the carboxy terminus. The Fc portion for this protein is human IgG1 (Thr106-Lys330).
- Molecular Mass
- The 360 amino acid recombinant protein has a predicted molecular mass of approximately 39.9 kD. The DTT-reduced and non-reduced glycosylated protein migrate at approximately 56 kD and 105 kD respectively by SDS-PAGE. The N-terminal amino acid is Ala.
- >95%, as determined by Coomassie stained SDS-PAGE.
- 0.22 µm filtered protein solution is in PBS.
- Endotoxin Level
- Less than 0.01 ng per µg cytokine as determined by the LAL method.
- 10 and 25 µg sizes are bottled at 200 µg/mL. 100 µg size and larger sizes are lot-specific and bottled at the concentration indicated on the vial (please contact technical support for concentration, or use our Lookup tool if you have a lot number.) Please note, new lots of the 100 µg size will be lot-specific and may differ from previous lots that had a fixed concentration.
- Storage & Handling
- Unopened vial can be stored between 2°C and 8°C for up to 2 weeks, at -20°C for up to six months, or at -70°C or colder until the expiration date. For maximum results, quick spin vial prior to opening. The protein can be aliquoted and stored at -20°C or colder. Stock solutions can also be prepared at 50 - 100 µg/mL in appropriate sterile buffer, carrier protein such as 0.2 - 1% BSA or HSA can be added when preparing the stock solution. Aliquots can be stored between 2°C and 8°C for up to one week and stored at -20°C or colder for up to 3 months. Avoid repeated freeze/thaw cycles.
- ED50 = 75 - 375 ng/ml, corresponding to a specific activity of 0.2-1.3 x 104 units/mg, as determined by the dose dependent inhibition of IL-2 production induced by 0.5 µg/ml of CD80 on Jurkat cells.
- Application Notes
BioLegend carrier-free recombinant proteins provided in liquid format are shipped on blue-ice. Our comparison testing data indicates that when handled and stored as recommended, the liquid format has equal or better stability and shelf-life compared to commercially available lyophilized proteins after reconstitution. Our liquid proteins are validated in-house to maintain activity after shipping on blue ice and are backed by our 100% satisfaction guarantee. If you have any concerns, contact us at firstname.lastname@example.org.
Activated T cells, CD4, CD8, and Tregs
- Inhibits T cell activation by reducing IL-2 production and IL-2R expression, also arresting T cells in G1 phase of the cell cycle; protects against the development of autoimmunity
- Antigen presenting cells, B cells
- Biology Area
- Immunology, Inhibitory Molecules
- Molecular Family
- CD Molecules, Immune Checkpoint Receptors, Soluble Receptors
- Antigen References
1. Linsley PS, et al. 1991. J. Exp. Med. 174:561.
2. Pentcheva-Hoang T, et al. 2004. Immunity 2:401.
3. Qureshi OS, et al. 2011. Science 332:600.
4. Gerold KD, et al. 2011. Diabetes 60:1955.
5. Ryden A, et al. 2012. Diabetes Metab. Res. Rev. 28:84.
6. Mittal AK, et al. 2013. PloS One 8:e70352.
7. Ward FJ, et al. 2013. Eur. J. Immunol. 43:1274.
- Gene ID
- 1493 View all products for this Gene ID
- View information about CTLA-4 on UniProt.org
- Does specific activity of a recombinant protein vary between lots?
Specific activity will vary for each lot and for the type of experiment that is done to validate it, but all passed lots will have activity within the established ED50 range for the product and we guarantee that our products will have lot-to-lot consistency. Please conduct an experiment-specific validation to find the optimal ED50 for your system.
- Have your recombinants been tested for stability?
Our testing shows that the recombinant proteins are able to withstand room temperature for a week without losing activity. In addition the recombinant proteins were also found to withstand four cycles of freeze and thaw without losing activity.
- How do you convert activity as an ED50 in ng/ml to a specific activity in Units/mg?
- Use formula Specific activity (Units/mg) = 10e6/ ED50 (ng/mL)
- How does the activity of your recombinant proteins compare to competitors?
We quality control each and every lot of recombinant protein. Not only do we check its bioactivity, but we also compare it against other commercially available recombinant proteins. We make sure each recombinant protein’s activity is at least as good as or better than the competition’s. In order to provide you with the best possible product, we ensure that our testing process is rigorous and thorough. If you’re curious and eager to make the switch to BioLegend recombinants, contact your sales representative today!
- What is the specific activity or ED50 of my recombinant protein?
The specific activity range of the protein is indicated on the product datasheets. Because the exact activity values on a per unit basis can largely fluctuate depending on a number of factors, including the nature of the assay, cell density, age of cells/passage number, culture media used, and end user technique, the specific activity is best defined as a range and we guarantee the specific activity of all our lots will be within the range indicated on the datasheet. Please note this only applies to recombinants labeled for use in bioassays. ELISA standard recombinant proteins are not recommended for bioassay usage as they are not tested for these applications.