InVivoMAb anti-mouse CD3ε F(ab')2 fragment

CloneCatalog #Category
145-2C11 f(ab')2 Fragments BE0001-1FABInVivoMAb Antibodies
$925 - $3825 Login for Academic & Non-profit Pricing

About InVivoMAb anti-mouse CD3ε F(ab')2 fragment

The anti-mouse CD3 F(ab')2 fragment is the antigen binding F(ab')2 fragments of the 145-2C11 monoclonal antibody only. The majority of the Fc fragment has been removed via pepsin digestion. The 145-2C11 antibody F(ab')2 fragment reacts with mouse CD3ε, a 20 kDa transmembrane cell-surface protein that belongs to the immunoglobulin superfamily. CD3ε is one of five polypeptide chains that combine to form the TCR complex. CD3ε is expressed on T lymphocytes, NK-T cells, and to varying degrees on developing thymocytes. CD3 plays roles in TCR signaling, T lymphocyte activation, and antigen recognition. The complete 145-2C11 antibody is commonly used to stimulate T cells in vitro however, when used in vivo activation of resting T cells results in cytokine release and toxicity caused by Ab-mediated cross-linking of T cells and Fcγ receptor-bearing cells. To avoid these complications non-Fc receptor binding F(ab’)2 fragments of the 145-2C11 antibody are commonly used. This non-FcR-binding anti-CD3 induces apoptosis of Ag-activated T cells in vivo by allowing durable expression of the TCR and sustained signaling. Foxp3+ Tregs have been shown to be resistant to CD3 antibody-mediated depletion.

InVivoMAb anti-mouse CD3ε F(ab')2 fragment Specifications

Isotype Armenian Hamster IgG1
Immunogen Mouse BM10-37 cytotoxic T cells
Reported Applications

in vivo T cell depletion

  • PBS, pH 7.0
  • Contains no stabilizers or preservatives
  • <2EU/mg (<0.002EU/μg)
  • Determined by LAL gel clotting assay
  • >95%
  • Determined by SDS-PAGE
Sterility 0.2 μM filtered
Production Pepsin Digest
Purification Protein A
RRID AB_2687679
Storage The antibody solution should be stored at the stock concentration at 4°C. Do not freeze.

Application References

InVivoMAb anti-mouse CD3ε F(ab)2 fragment (Clone: 145-2C11 f(ab)2 Fragments)


Besancon, A., et al. (2017). “The Induction and Maintenance of Transplant Tolerance Engages Both Regulatory and Anergic CD4(+) T cells.” Front Immunol 8: 218. PubMed

Therapeutic tolerance to self-antigens or foreign antigens is thought to depend on constant vigilance by Foxp3(+) regulatory T cells (Tregs). Previous work using a pancreatic islet allograft model and a short pulse of CD3 antibody therapy has shown that CD8(+) T cells become anergic and use TGFbeta and coinhibitory signaling as their contribution to the tolerance process. Here, we examine the role of CD4(+) T cells in tolerization by CD3 antibodies. We show that both Foxp3(+) Tregs and CD4(+) T cell anergy play a role in the induction of tolerance and its maintenance. Foxp3(+) Tregs resisted CD3 antibody-mediated depletion, unlike intragraft Th1 CD4(+) lymphocytes coexpressing granzyme B and Tbx21, which were selectively eliminated. Tregs were mandatory for induction of tolerance as their depletion at the time of CD3 antibody therapy or for a short time thereafter, by an antibody to CD25 (PC61), led to graft rejection. Early treatment with CTLA-4 antibody gave the same outcome. In contrast, neither PC61 nor anti-CTLA-4 given late, at day 100 posttransplant, reversed tolerance once established. Ablation of Foxp3 T cells after diphtheria toxin injection in tolerant Foxp3(DTR) recipient mice provided the same outcome. Alloreactive T cells had been rendered intrinsically unresponsive as total CD4(+) or Treg-deprived CD4(+) T cells from tolerant recipients were unable to mount donor-specific IFN-gamma responses. In addition, intragraft Treg-deprived CD4(+) T cells lacked proliferative capacities, expressed high levels of the inhibitory receptor PD-1, and exhibited a CD73(hi)FR4(hi) phenotype, thus reflecting a state of T cell anergy. We conclude that Tregs play a substantive and critical role in guiding the immune system toward tolerance of the allograft, when induced by CD3 antibody, but are less important for maintenance of the tolerant state, where T cell anergy appears sufficient.


Chae, W. J., et al. (2017). “Membrane-bound Dickkopf-1 in Foxp3(+) regulatory T cells suppresses T-cell-mediated autoimmune colitis.” Immunology 152(2): 265-275. PubMed

Induction of tolerance is a key mechanism to maintain or to restore immunological homeostasis. Here we show that Foxp3(+) regulatory T (Treg) cells use Dickkopf-1 (DKK-1) to regulate T-cell-mediated tolerance in the T-cell-mediated autoimmune colitis model. Treg cells from DKK-1 hypomorphic doubleridge mice failed to control CD4(+) T-cell proliferation, resulting in CD4 T-cell-mediated autoimmune colitis. Thymus-derived Treg cells showed a robust expression of DKK-1 but not in naive or effector CD4 T cells. DKK-1 expression in Foxp3(+) Treg cells was further increased upon T-cell receptor stimulation in vitro and in vivo. Interestingly, Foxp3(+) Treg cells expressed DKK-1 in the cell membrane and the functional inhibition of DKK-1 using DKK-1 monoclonal antibody abrogated the suppressor function of Foxp3(+) Treg cells. DKK-1 expression was dependent on de novo protein synthesis and regulated by the mitogen-activated protein kinase pathway but not by the canonical Wnt pathway. Taken together, our results highlight membrane-bound DKK-1 as a novel Treg-derived mediator to maintain immunological tolerance in T-cell-mediated autoimmune colitis.


Sarikonda, G., et al. (2015). “The Hsp60 peptide p277 enhances anti-CD3 mediated diabetes remission in non-obese diabetic mice.” J Autoimmun 59: 61-66. PubMed

Type 1 diabetes (T1D) is characterized by the immune-mediated destruction of pancreatic beta cells leading to inadequate glycemic control. Trials with immunomodulatory monotherapies have shown that the disease course can in principle be altered. The observed preservation of endogenous insulin secretion however is typically transient and chronic treatment is often associated with significant side effects. Here we combined anti-CD3 with the Hsp60 peptide p277, two drugs that have been evaluated in Phase 3 trials, to test for enhanced efficacy. Female NOD mice with recent onset diabetes were given 5 mug anti-CD3 i.v., on three consecutive days in combination with 100 mug of p277 peptide in IFA s.c., once weekly for four weeks. Anti-CD3 alone restored normoglycemia in 44% of the mice while combination therapy with anti-CD3 and p277 induced stable remission in 83% of mice. The observed increase in protection occurred only in part through TLR2 signaling and was characterized by increased Treg numbers and decreased insulitis. These results have important implications for the design of combination therapies for the treatment of T1D.


Valle, A., et al. (2015). “Heterogeneous CD3 expression levels in differing T cell subsets correlate with the in vivo anti-CD3-mediated T cell modulation.” J Immunol 194(5): 2117-2127. PubMed

The tolerogenic anti-CD3epsilon monoclonal Abs (anti-CD3) are promising compounds for the treatment of type 1 diabetes. Anti-CD3 administration induces transient T cell depletion both in preclinical and in clinical studies. Notably, the said depletion mainly affects CD4(+) but not CD8(+) T cells. Moreover, type 1 diabetes reversal in preclinical models is accompanied by the selective expansion of CD4(+)Foxp3(+) T regulatory (Treg) cells, which are fundamental for the long-term maintenance of anti-CD3-mediated tolerance. The mechanisms that lead to this immune-shaping by affecting mainly CD4(+) T effector cells while sparing CD4(+)Foxp3(+) Treg cells have still to be fully elucidated. This study shows that CD3 expression levels differ from one T cell subset to another. CD4(+)Foxp3(-) T cells contain higher amounts of CD3 molecules than do CD4(+)Foxp3(+) and CD8(+) T cells in both mice and humans. The said differences correlate with the anti-CD3-mediated immune resetting that occurs in vivo after anti-CD3 administration in diabetic NOD mice. Additionally, transcriptome analysis demonstrates that CD4(+)Foxp3(+) Treg cells are significantly less responsive than are CD4(+)Foxp3(-) T cells to anti-CD3 treatment at a molecular level. Thus, heterogeneity in CD3 expression seems to confer to the various T cell subsets differing susceptibility to the in vivo tolerogenic anti-CD3-mediated modulation. These data shed new light on the molecular mechanism that underlies anti-CD3-mediated immune resetting and thus may open new opportunities to improve this promising treatment.


Kasahara, K., et al. (2014). “CD3 antibody and IL-2 complex combination therapy inhibits atherosclerosis by augmenting a regulatory immune response.” J Am Heart Assoc 3(2): e000719. PubMed

BACKGROUND: Accumulating evidence suggests that the balance between pathogenic effector T cells (Teffs) and regulatory T cells (Tregs) may be important for controlling atherosclerotic disease. We hypothesized that a combination therapy with anti-CD3 antibody (CD3-Ab) and IL-2/anti-IL-2 monoclonal antibody complex (IL-2 complex) aimed at increasing the ratio of Tregs to Teffs would effectively inhibit atherosclerosis in mice. METHODS AND RESULTS: We treated apolipoprotein E-deficient mice fed a high-cholesterol diet with vehicle, CD3-Ab, IL-2 complex, or their combination. Mice receiving the combination therapy had markedly reduced atherosclerotic lesions than mice treated with CD3-Ab or IL-2 complex alone. In addition, a striking increase in the Treg/Teff ratio of lymphoid organs and atherosclerotic lesions, along with plaque stabilization characterized by decreased macrophage content and increased collagen content was observed. The combination treatment also markedly reduced splenic Ly6C(high) inflammatory monocytes and might induce a favorable macrophage phenotype change in atherosclerotic lesions. CONCLUSIONS: Our results indicate that in addition to suppressing Teff responses, enhancing Treg-mediated immune responses is more efficacious in preventing atherosclerosis, suggesting a novel therapeutic approach for atherosclerosis.


Kita, T., et al. (2014). “Regression of atherosclerosis with anti-CD3 antibody via augmenting a regulatory T-cell response in mice.” Cardiovasc Res 102(1): 107-117. PubMed

AIMS: Although recent animal studies have investigated the cellular and molecular mechanisms underlying the process of atherosclerosis regression, it remains unknown whether adaptive immune responses including T cells are involved in this process. We investigated the role of T cells in atherosclerosis regression. METHODS AND RESULTS: LDL receptor-deficient mice were fed a high-cholesterol diet for 8 weeks to form atherosclerotic lesions and were then changed to a standard diet, and atherosclerosis was assessed 4 weeks later. Just before changing the diet, the mice received an iv injection of anti-CD3 antibody (CD3-Ab) or control immunoglobulin G for 5 consecutive days. CD3-Ab treatment regressed atherosclerosis and decreased the accumulation of macrophages and CD4(+) T cells in the plaques. CD3-Ab treatment also dramatically reduced CD4(+) T cells and increased the proportion of regulatory T cells (Tregs). Depletion of Tregs by anti-CD25 antibody injection abolished the regression of atherosclerosis seen in CD3-Ab-treated mice, indicating the essential role for Tregs in this process. CONCLUSION: CD3-Ab treatment induced rapid regression of established atherosclerosis via reducing CD4(+) T cells and increasing the proportion of Tregs. These findings suggest that therapeutic intervention for T-cell-mediated immune responses may represent a novel strategy to induce atherosclerosis regression in combination with lipid-lowering therapy.


Shiheido, H., et al. (2014). “Novel CD3-specific antibody induces immunosuppression via impaired phosphorylation of LAT and PLCgamma1 following T-cell stimulation.” Eur J Immunol 44(6): 1770-1780. PubMed

The activation of T cells is known to be accompanied by the temporary downmodulation of the TCR/CD3 complex on the cell surface. Here, we established a novel monoclonal antibody, Dow2, that temporarily induces downmodulation of the TCR/CD3 complex in mouse CD4(+) T cells without activating T cells. Dow2 recognized the determinant on CD3epsilon; however, differences were observed in the binding mode between Dow2 and the agonistic anti-CD3epsilon Ab, 145-2C11. An injection of Dow2 in vivo resulted in T-cell anergy, and prolonged the survival of cardiac allografts without a marked increase in cytokine release. The phosphorylated forms of the signaling proteins PLC-gamma1 and LAT in Dow2-induced anergic T cells were markedly decreased upon stimulation. However, the levels of phosphorylated LAT and PLCgamma1 in Dow2-induced anergic T cells could be rescued in the presence of the proteasome inhibitor MG-132. These results suggest that proteasome-mediated degradation is involved in hypophosphorylated LAT and PLCgamma1 in Dow2-induced anergic T cells. The novel CD3-specific Ab, Dow2, may provide us with a unique tool for inducing immunosuppression.


Grant, C. W., et al. (2013). “Testing agents for prevention or reversal of type 1 diabetes in rodents.” PLoS One 8(8): e72989. PubMed

We report the results of an independent laboratory’s tests of novel agents to prevent or reverse type 1 diabetes (T1D) in the non-obese diabetic (NOD) mouse, BioBreeding diabetes prone (BBDP) rat, and multiple autoimmune disease prone (MAD) rat models. Methods were developed to better mimic human clinical trials, including: prescreening, randomization, blinding, and improved glycemic care of the animals. Agents were suggested by the research community in an open call for proposals, and selected for testing by an NIDDK appointed independent review panel. Agents selected for testing to prevent diabetes at later stages of progression in a rodent model were a STAT4 antagonist (DT22669), alpha1 anti-trypsin (Aralast NP), celastrol (a natural product with anti-inflammatory properties), and a Macrophage Inflammatory Factor inhibitor (ISO-092). Agents tested for reversal of established T1D in rodent models were: alpha1 anti-trypsin (Aralast NP), tolerogenic peptides (Tregitopes), and a long-acting formulation of GLP-1 (PGC-GLP-1). None of these agents were seen to prevent or reverse type 1 diabetes, while the positive control interventions were effective: anti-CD3 treatment provided disease reversal in the NOD mouse, dexamethasone prevented T1D induction in the MAD rat, and cyclosporin prevented T1D in the BBDP rat. For some tested agents, details of previous formulation, delivery, or dosing, as well as laboratory procedure, availability of reagents and experimental design, could have impacted our ability to confirm prior reports of efficacy in preclinical animal models. In addition, the testing protocols utilized here provided detection of effects in a range commonly used in placebo controlled clinical trials (for example, 50% effect size), and thus may have been underpowered to observe more limited effects. That said, we believe the results compiled here, showing good control and repeatability, confirm the feasibility of screening diverse test agents in an independent laboratory.


Carpenter, P. A., et al. (2000). “Non-Fc receptor-binding humanized anti-CD3 antibodies induce apoptosis of activated human T cells.” J Immunol 165(11): 6205-6213. PubMed

Human trials in organ allografts have demonstrated that murine anti-CD3 mAbs are immunosuppressive. By mimicking Ag, anti-CD3 can produce T cell activation, anergy, or death. Activation of resting T cells in vivo results in dose-limiting cytokine release and is caused by Ab-mediated cross-linking of T cells and Fcgamma receptor (FcR)-bearing cells. With the goal of minimizing cytokine-induced toxicity, anti-CD3 have been engineered to lower Fc binding avidity. Preclinical murine studies have indicated that non-FcR-binding anti-CD3 can induce apoptosis of Ag-activated T cells. Since induction of T cell apoptosis may be an important mechanism of immunosuppression by anti-CD3, we tested whether Fc mutations affect the ability of anti-human CD3 to induce apoptosis of activated T cells. We compared wild-type murine anti-CD3, M291, and OKT3 and their humanized, FcR- and non-FcR-binding structural variants in quantitative assays of T cell apoptosis. Non-FcR-binding variants produced more sustainable phosphorylation of extracellular signal-regulated kinase-2, greater release of IFN-gamma, and more effectively caused activation-dependent T cell apoptosis. Non-FcR-binding variants dissociated more quickly from the T cell surface and caused less internalization of the TCR, which then remained available in greater abundance on the cell surface for signaling. Cross-linking of non-FcR-binding variants by antiglobulin enhanced TCR internalization and minimized induction of T cell apoptosis. We conclude that non-FcR-binding, humanized anti-CD3 have improved ability to induce apoptosis of activated T cells, presumably by allowing durable expression of the TCR and sustained signaling.