About InVivoMAb anti-rat CD28 The JJ316 monoclonal antibody reacts with rat CD28, a 45 kDa costimulatory receptor and a member of the Ig superfamily. CD28 is expressed by thymocytes, most peripheral T cells, and NK cells. CD28 is a receptor for CD80 (B7-1) and CD86 (B7-2). Signaling through CD28 augments IL-2 and IL-2 receptor expression as well as cytotoxicity of CD3-activated T cells. In contrast to JJ319 which is a “conventional” CD28-specific Mab that costimulates anti-TCR-triggered proliferation in vitro, JJ316 is a CD28 “superagonistic” antibody that induces polyclonal T cell proliferation in vitro and in vivo and does not require TCR ligation. InVivoMAb anti-rat CD28 Specifications IsotypeMouse IgG1, κ Recommended Isotype Control(s)InVivoMAb mouse IgG1 isotype control, unknown specificity Recommended Dilution BufferInVivoPure pH 7.0 Dilution Buffer ImmunogenRat CD28-transfected BALB/c mouse A2OJ cell line Reported Applicationsin vivo T cell stimulation/activation FormulationPBS, pH 7.0 Contains no stabilizers or preservatives Endotoxin<2EU/mg (<0.002EU/μg) Determined by LAL gel clotting assay Purity>95% Determined by SDS-PAGE Sterility0.2 μm filtered ProductionPurified from cell culture supernatant in an animal-free facility PurificationProtein G Molecular Weight150 kDa StorageThe antibody solution should be stored at the stock concentration at 4°C. Do not freeze. Application ReferencesInVivoMAb anti-rat CD28 (CLONE: JJ316)Takabatake Y, Li XK, Mizui M, Miyasato K, Matsui I, Kawada N, Imai E, Hünig T, Takahara S, Wada T, Furuichi K, Rakugi H, Isaka Y (2011). "A superagonistic monoclonal antibody for CD28 ameliorates crescentic glomerulonephritis in wistar-kyoto rats" Mol Med 17(7-8):686-96. PubMedRegulatory T (Treg) cells play an important role in the resolution of crescentic glomerulonephritis, where a T helper 1 (Th1)-predominant immune response promotes crescent formation. Therefore, agents that increase Treg cells appear to be ideal for suppressing T-cell-mediated renal pathology. We hypothesized that a superagonistic monoclonal antibody for CD28 (JJ316), which has been known to preferentially expand Treg cells in vivo, could prevent nephrotoxic serum-induced nephritis in Wistar-Kyoto rats, one of the experimental models of crescentic glomerulonephritis. Administration of JJ316 attenuated crescent formation, proteinuria and glomerular accumulation of macrophages and CD8(+) T cells. These changes were accompanied by increased infiltration of Treg cells. Among glomerular macrophages, the CD163(+) subset was significantly increased after treatment, suggesting that Treg cells may modulate the phenotype of macrophages leading to resolution of glomerulonephritis. In an adoptive transfer experiment, two T-cell subsets (CD4(+)CD25(+) and CD4(+)CD25(-) T cells) purified from spleens and lymph nodes of donor rats primed with JJ316 3 d before were inoculated into nephritic recipient rats, which recapitulated the beneficial effects of in vivo administration of JJ316. Furthermore, a single injection of JJ316 administered 3 d after disease induction completely protected nephritic rats from death for 2 months. In conclusion, we demonstrated that treatment with JJ316 has a dramatic therapeutic effect on an experimental crescentic glomerulonephritis, possibly due to expansion and activation of Treg cells.Wang S, Liu J, Wang M, Zhang J, Wang Z (2010). "Treatment and prevention of experimental autoimmune myocarditis with CD28 superagonists" Cardiology 115(2):107-13. PubMedExperimental autoimmune myocarditis (EAM), a rodent model of human dilated cardiomyopathy (DCM), is mediated by an autoimmune mechanism. We investigated whether a CD28 superagonistic antibody selectively targeting CD4+CD25+ regulatory T cells (T(regs)) provides effective therapy for EAM.Müller N, van den Brandt J, Odoardi F, Tischner D, Herath J, Flügel A, Reichardt HM (2008). "A CD28 superagonistic antibody elicits 2 functionally distinct waves of T cell activation in rats" J Clin Invest 118(4):1405-16. PubMedAdministration of the CD28 superagonistic antibody JJ316 is an efficient means to treat autoimmune diseases in rats, but the humanized antibody TGN1412 caused devastating side effects in healthy volunteers during a clinical trial. Here we show that JJ316 treatment of rats induced a dramatic redistribution of T lymphocytes from the periphery to the secondary lymphoid organs, resulting in severe T lymphopenia. Live imaging of secondary lymphoid organs revealed that JJ316 administration almost instantaneously (<2 minutes) arrested T cells in situ. This reduction in T cell motility was accompanied by profound cytoskeletal rearrangements and increased cell size. In addition, surface expression of lymphocyte function-associated antigen-1 was enhanced, endothelial differentiation sphingolipid G protein-coupled receptor 1 and L selectin levels were downregulated, and the cells lost their responsiveness to sphingosine 1-phosphate-directed migration. These proadhesive alterations were accompanied by signs of strong activation, including upregulation of CD25, CD69, CD134, and proinflammatory mediators. However, this did not lead to a cytokine storm similar to the clinical trial. While most of the early changes disappeared within 48 hours, we observed that CD4+CD25+FoxP3+ regulatory T cells experienced a second phase of activation, which resulted in massive cell enlargement, extensive polarization, and increased motility. These data suggest that CD28 superagonists elicit 2 qualitatively distinct waves of activation.Rodríguez-Palmero M, Franch A, Castell M, Pelegrí C, Pérez-Cano FJ, Kleinschnitz C, Stoll G, Hünig T, Castellote C (2006). "Effective treatment of adjuvant arthritis with a stimulatory CD28-specific monoclonal antibody" J Rheumatol 33(1):110-8. PubMedTo determine the immunomodulatory effects of the anti-rat CD28 monoclonal antibody (Mab) JJ316 on the onset of rat adjuvant arthritis (AA). JJ316 is a superagonistic Mab that induces polyclonal T cell proliferation in the absence of T cell receptor (TCR) ligation and promotes the expansion of regulatory T cells.
