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Anti-Human CD28 (CD28)
Purified No Carrier Protein
(MGC138290, Tp44)
Monoclonal Antibody
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Prod. No.:C1400
Clone:37407
Isotype:Mouse IgG1
Pkg. Size:500 µg, 1 mg
Storage:-20°C to -70°C Detailed storage instructions below.
Description
Specificity:
Mouse Anti-Human CD28 (Clone 37407) recognizes an epitope on Human CD28. This monoclonal antibody was purified using multi-step affinity chromatography methods such as Protein A or G depending on the species and isotype. This antibody shows no cross-reactivity with rmCD28, rhCTLA4 or rmCTLA4.

Background:
CD28 (Cluster of Differentiation 28) is one of the molecules expressed on T cells that provide co-stimulatory signals, which are required for T cell activation. CD28 is composed of a single Ig V-like extracellular domain, a transmembrane domain and an intracellular domain. It is expressed on the cell surface as disulfide-linked homodimer or as monomer. CD28 is the receptor for B7.1 (CD80) and B7.2 (CD86). When activated by Toll-like receptor ligands, the B7.1 expression is upregulated in antigen presenting cells (APCs). The B7.2 expression on antigen presenting cells is constitutive. CD28 is the only B7 receptor constitutively expressed on naive T cells. Stimulation through CD28 in addition to the TCR can provide a potent co-stimulatory signal to T cells for the production of various interleukins (IL-2 and IL-6 in particular). CD28 possesses an intracellular domain with several residues that are critical for its effective signalling. CD28 also contains two proline-rich motifs that are able to bind SH3-containing proteins.

CD28 has also been found to stimulate eosinophil granulocytes where its ligation with anti-CD28 leads to the release of IL-2, IL4, IL-13 and IFN-γ.

Known Species Cross Reactivity: Hu+
Host Species
Mouse
Immunogen
Purified Recombinant Human CD28 (>98%)
Formulation
This monoclonal antibody has been 0.2 µm filtered and lyophilized from modified Dulbecco's phosphate buffered saline (1X PBS) pH 7.2 - 7.3 containing 5.0% w/v trehalose with no calcium, magnesium or preservatives present.
Reconstitution
This lyophilized antibody may be aseptically reconstituted with sterile PBS to a final volume of 1.0 ml. This will result in a final antibody concentration of 500 µg/ml.
Endotoxin
<0.1 EU/µg as determined by the LAL method
Storage and Stability
The lyophilized antibody can be stored desiccated at -20°C to -70°C for up to twelve months. The reconstituted antibody can be stored for at least four weeks at 2-8°C. For long-term storage of the reconstituted antibody, aseptically aliquot into working volumes and store at -20°C to -70°C in a manual defrost freezer. Avoid repeated freeze thaw cycles. No detectable loss of activity was observed after six months.
Reported Applications
Indirect ELISA, FC, WB, FA
Indirect ELISA: This antibody can be used in an indirect detection ELISA at 0.5 - 1.0 µg/ml with a suitable second step reagent such as Goat Anti-Mouse IgG (H&L)-Horseradish Peroxidase (Leinco Prod. No.: M114). The detection sensitivity of this indirect ELISA for Human CD28 is approximately 2.0 ng/well.
Flow Cytometry: It is recommended to use the indirect method for signal enhancement when enumerating cells expressing CD28. A suggested method would be to stain cells expressing CD28 with 1-2 µg per 1 x 106 cells in a total staining volume of 100 µl followed by Goat Anti-Mouse IgG (H&L)-R-phycoerythrin (Leinco Prod. No.: M1189).
Western Blotting: To detect Human CD28 this monoclonal antibody can be used at a concentration of 1-2 µg/ml. This monoclonal antibody should be used in conjunction with compatible second-step reagents such as Goat Anti-Mouse IgG (H&L)-Horseradish Peroxidase (Leinco Prod. No.: M114) and a chromogenic substrate such as TMB HRPO Membrane/IHC Substrate UltraSensitive™ One Component "Ready Use" (Leinco Prod. No.: T343). The detection limit for Human CD28 is 25 ng/lane under either reducing or non-reducing conditions. The sensitivity of detection may increase up to 50 fold when a chemiluminescent substrate is used. A suitable Western blotting control is Recombinant Human CD28 (Leinco Prod. No.: C1338).
Functional Assay: CD28 is a 44 kD homodimeric transmembrane glycoprotein expressed on most T cells and thymocytes. It has been identified as an important molecule involved in T cell activation. Ligation of CD28 with anti-CD28 antibody or the counter-receptors B7-1 or B7-2 delivers costimulatory signals for T cell activation. The biological activity of this anti-human CD28 was measured by its ability to enhance IL-2 production by Jurkat cells stimulated with PMA and calcium ionophore. The ED50 for this effect is typically 0.2 - 0.6 μg/mL under the assay conditions shown in the figure legend.
Each investigator should determine their own optimal working dilution for specific applications.


Country of Origin
USA
Related Products
Goat Anti-Mouse IgG (H&L)-Horseradish Peroxidase (Leinco Prod. No.: M114)
Goat Anti-Mouse IgG (H&L)-R-phycoerythrin (Leinco Prod. No.: M1189)
TMB HRPO Membrane/IHC Substrate UltraSensitive™ One Component "Ready Use" (Leinco Prod. No.: T343)
Recombinant Human CD28 (Leinco Prod. No.: C1338)
Anti-Mouse CD28 Purified Low Endotoxin Functional Formulation (Leinco Prod. No.: C379)
Anti-Human CD28-Biotin (Antigen Aff. Pur.) (Leinco Prod. No.: C1491)
Anti-Mouse CD28-Biotin (Antigen Aff. Pur.) (Leinco Prod. No.: C1503)
Anti-Human CD28-R-phycoerythrin (R-PE) (Leinco Prod. No.: C1580)
Anti-Mouse CD28 (Antigen Aff. Pur.) (Leinco Prod. No.: C1392)
Recombinant Mouse CD28 (Leinco Prod. No.: C1341)
Anti-Mouse CD28 Purified (Leinco Prod. No.: C243)
Anti-Mouse CD28-Biotin (Leinco Prod. No.: C244)
Anti-Mouse CD28-R-phycoerythrin (Leinco Prod. No.: C246)
PN:
Anti-Mouse CD28-Allophycocyanin (Leinco Prod. No.: C347)
Products are for research use only. Not for use in diagnostic or therapeutic procedures.
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