InVivoMAb rat IgG1 isotype control, anti-horseradish peroxidase

CloneCatalog #Category
HRPNBE0088Isotype Controls
$95 - $3250

About InVivoMAb rat IgG1 isotype control, anti-horseradish peroxidase

The HRPN monoclonal antibody reacts with horseradish peroxidase (HRP). Because HRP is not expressed by mammals this antibody is ideal for use as an isotype-matched control for rat IgG1 antibodies in most in vivo and in vitro applications. This antibody can interfere with HRP detection based assays. If using downstream HRP based assays to analyze samples derived from treated animals, please consider using our alternative rat IgG1 isotype control antibody BE0290.

InVivoMAb rat IgG1 isotype control, anti-horseradish peroxidase Specifications

Isotype Rat IgG1, κ
  • 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 Purified from tissue culture supernatant in an animal free facility
Purification Protein G
RRID AB_1107775
Molecular Weight 150 kDa
Storage The antibody solution should be stored at the stock concentration at 4°C. Do not freeze.

Application References

InVivoMAb rat IgG1 Isotype control; anti-horseradish peroxidase (Clone: HRPN)

Goschl, L., et al. (2018). “A T cell-specific deletion of HDAC1 protects against experimental autoimmune encephalomyelitis.” J Autoimmun 86: 51-61. PubMed

Multiple sclerosis (MS) is a human neurodegenerative disease characterized by the invasion of autoreactive T cells from the periphery into the CNS. Application of pan-histone deacetylase inhibitors (HDACi) ameliorates experimental autoimmune encephalomyelitis (EAE), an animal model for MS, suggesting that HDACi might be a potential therapeutic strategy for MS. However, the function of individual HDAC members in the pathogenesis of EAE is not known. In this study we report that mice with a T cell-specific deletion of HDAC1 (using the Cd4-Cre deleter strain; HDAC1-cKO) were completely resistant to EAE despite the ability of HDAC1cKO CD4(+) T cells to differentiate into Th17 cells. RNA sequencing revealed STAT1 as a prominent upstream regulator of differentially expressed genes in activated HDAC1-cKO CD4(+) T cells and this was accompanied by a strong increase in phosphorylated STAT1 (pSTAT1). This suggests that HDAC1 controls STAT1 activity in activated CD4(+) T cells. Increased pSTAT1 levels correlated with a reduced expression of the chemokine receptors Ccr4 and Ccr6, which are important for the migration of T cells into the CNS. Finally, EAE susceptibility was restored in WT:HDAC1-cKO mixed BM chimeric mice, indicating a cell-autonomous defect. Our data demonstrate a novel pathophysiological role for HDAC1 in EAE and provide evidence that selective inhibition of HDAC1 might be a promising strategy for the treatment of MS.


Clemente-Casares, X., et al. (2016). “Expanding antigen-specific regulatory networks to treat autoimmunity.” Nature 530(7591): 434-440. PubMed

Regulatory T cells hold promise as targets for therapeutic intervention in autoimmunity, but approaches capable of expanding antigen-specific regulatory T cells in vivo are currently not available. Here we show that systemic delivery of nanoparticles coated with autoimmune-disease-relevant peptides bound to major histocompatibility complex class II (pMHCII) molecules triggers the generation and expansion of antigen-specific regulatory CD4(+) T cell type 1 (TR1)-like cells in different mouse models, including mice humanized with lymphocytes from patients, leading to resolution of established autoimmune phenomena. Ten pMHCII-based nanomedicines show similar biological effects, regardless of genetic background, prevalence of the cognate T-cell population or MHC restriction. These nanomedicines promote the differentiation of disease-primed autoreactive T cells into TR1-like cells, which in turn suppress autoantigen-loaded antigen-presenting cells and drive the differentiation of cognate B cells into disease-suppressing regulatory B cells, without compromising systemic immunity. pMHCII-based nanomedicines thus represent a new class of drugs, potentially useful for treating a broad spectrum of autoimmune conditions in a disease-specific manner.


Ellis, G. T., et al. (2015). “TRAIL+ monocytes and monocyte-related cells cause lung damage and thereby increase susceptibility to influenza-Streptococcus pneumoniae coinfection.” EMBO Rep 16(9): 1203-1218. PubMed

Streptococcus pneumoniae coinfection is a major cause of influenza-associated mortality; however, the mechanisms underlying pathogenesis or protection remain unclear. Using a clinically relevant mouse model, we identify immune-mediated damage early during coinfection as a new mechanism causing susceptibility. Coinfected CCR2(-/-) mice lacking monocytes and monocyte-derived cells control bacterial invasion better, show reduced epithelial damage and are overall more resistant than wild-type controls. In influenza-infected wild-type lungs, monocytes and monocyte-derived cells are the major cell populations expressing the apoptosis-inducing ligand TRAIL. Accordingly, anti-TRAIL treatment reduces bacterial load and protects against coinfection if administered during viral infection, but not following bacterial exposure. Post-influenza bacterial outgrowth induces a strong proinflammatory cytokine response and massive inflammatory cell infiltrate. Depletion of neutrophils or blockade of TNF-alpha facilitate bacterial outgrowth, leading to increased mortality, demonstrating that these factors aid bacterial control. We conclude that inflammatory monocytes recruited early, during the viral phase of coinfection, induce TRAIL-mediated lung damage, which facilitates bacterial invasion, while TNF-alpha and neutrophil responses help control subsequent bacterial outgrowth. We thus identify novel determinants of protection versus pathology in influenza-Streptococcus pneumoniae coinfection.


Grinberg-Bleyer, Y., et al. (2015). “Cutting edge: NF-kappaB p65 and c-Rel control epidermal development and immune homeostasis in the skin.” J Immunol 194(6): 2472-2476. PubMed

Psoriasis is an inflammatory skin disease in which activated immune cells and the proinflammatory cytokine TNF are well-known mediators of pathogenesis. The transcription factor NF-kappaB is a key regulator of TNF production and TNF-induced proinflammatory gene expression, and both the psoriatic transcriptome and genetic susceptibility further implicate NF-kappaB in psoriasis etiopathology. However, the role of NF-kappaB in psoriasis remains controversial. We analyzed the function of canonical NF-kappaB in the epidermis using CRE-mediated deletion of p65 and c-Rel in keratinocytes. In contrast to animals lacking p65 or c-Rel alone, mice lacking both subunits developed severe dermatitis after birth. Consistent with its partial histological similarity to human psoriasis, this condition could be prevented by anti-TNF treatment. Moreover, regulatory T cells in lesional skin played an important role in disease remission. Our results demonstrate that canonical NF-kappaB in keratinocytes is essential for the maintenance of skin immune homeostasis and is protective against spontaneous dermatitis.


Meisen, W. H., et al. (2015). “The Impact of Macrophage- and Microglia-Secreted TNFalpha on Oncolytic HSV-1 Therapy in the Glioblastoma Tumor Microenvironment.” Clin Cancer Res 21(14): 3274-3285. PubMed

PURPOSE: Oncolytic herpes simplex viruses (oHSV) represent a promising therapy for glioblastoma (GBM), but their clinical success has been limited. Early innate immune responses to viral infection reduce oHSV replication, tumor destruction, and efficacy. Here, we characterized the antiviral effects of macrophages and microglia on viral therapy for GBM. EXPERIMENTAL DESIGN: Quantitative flow cytometry of mice with intracranial gliomas (+/-oHSV) was used to examine macrophage/microglia infiltration and activation. In vitro coculture assays of infected glioma cells with microglia/macrophages were used to test their impact on oHSV replication. Macrophages from TNFalpha-knockout mice and blocking antibodies were used to evaluate the biologic effects of TNFalpha on virus replication. TNFalpha blocking antibodies were used to evaluate the impact of TNFalpha on oHSV therapy in vivo. RESULTS: Flow-cytometry analysis revealed a 7.9-fold increase in macrophage infiltration after virus treatment. Tumor-infiltrating macrophages/microglia were polarized toward a M1, proinflammatory phenotype, and they expressed high levels of CD86, MHCII, and Ly6C. Macrophages/microglia produced significant amounts of TNFalpha in response to infected glioma cells in vitro and in vivo. Using TNFalpha-blocking antibodies and macrophages derived from TNFalpha-knockout mice, we discovered TNFalpha-induced apoptosis in infected tumor cells and inhibited virus replication. Finally, we demonstrated the transient blockade of TNFalpha from the tumor microenvironment with TNFalpha-blocking antibodies significantly enhanced virus replication and survival in GBM intracranial tumors. CONCLUSIONS: The results of these studies suggest that FDA approved TNFalpha inhibitors may significantly improve the efficacy of oncolytic virus therapy.


Park, H. J., et al. (2015). “PD-1 upregulated on regulatory T cells during chronic virus infection enhances the suppression of CD8+ T cell immune response via the interaction with PD-L1 expressed on CD8+ T cells.” J Immunol 194(12): 5801-5811. PubMed

Regulatory T (Treg) cells act as terminators of T cell immuniy during acute phase of viral infection; however, their role and suppressive mechanism in chronic viral infection are not completely understood. In this study, we compared the phenotype and function of Treg cells during acute or chronic infection with lymphocytic choriomeningitis virus. Chronic infection, unlike acute infection, led to a large expansion of Treg cells and their upregulation of programmed death-1 (PD-1). Treg cells from chronically infected mice (chronic Treg cells) displayed greater suppressive capacity for inhibiting both CD8(+) and CD4(+) T cell proliferation and subsequent cytokine production than those from naive or acutely infected mice. A contact between Treg and CD8(+) T cells was necessary for the potent suppression of CD8(+) T cell immune response. More importantly, the suppression required cell-specific expression and interaction of PD-1 on chronic Treg cells and PD-1 ligand on CD8(+) T cells. Our study defines PD-1 upregulated on Treg cells and its interaction with PD-1 ligand on effector T cells as one cause for the potent T cell suppression and proposes the role of PD-1 on Treg cells, in addition to that on exhausted T cells, during chronic viral infection.


Sell, S., et al. (2015). “Control of murine cytomegalovirus infection by gammadelta T cells.” PLoS Pathog 11(2): e1004481. PubMed

Infections with cytomegalovirus (CMV) can cause severe disease in immunosuppressed patients and infected newborns. Innate as well as cellular and humoral adaptive immune effector functions contribute to the control of CMV in immunocompetent individuals. None of the innate or adaptive immune functions are essential for virus control, however. Expansion of gammadelta T cells has been observed during human CMV (HCMV) infection in the fetus and in transplant patients with HCMV reactivation but the protective function of gammadelta T cells under these conditions remains unclear. Here we show for murine CMV (MCMV) infections that mice that lack CD8 and CD4 alphabeta-T cells as well as B lymphocytes can control a MCMV infection that is lethal in RAG-1(-/-) mice lacking any T- and B-cells. gammadelta T cells, isolated from infected mice can kill MCMV infected target cells in vitro and, importantly, provide long-term protection in infected RAG-1(-/-) mice after adoptive transfer. gammadelta T cells in MCMV infected hosts undergo a prominent and long-lasting phenotypic change most compatible with the view that the majority of the gammadelta T cell population persists in an effector/memory state even after resolution of the acute phase of the infection. A clonotypically focused Vgamma1 and Vgamma2 repertoire was observed at later stages of the infection in the organs where MCMV persists. These findings add gammadelta T cells as yet another protective component to the anti-CMV immune response. Our data provide clear evidence that gammadelta T cells can provide an effective control mechanism of acute CMV infections, particularly when conventional adaptive immune mechanisms are insufficient or absent, like in transplant patient or in the developing immune system in utero. The findings have implications in the stem cell transplant setting, as antigen recognition by gammadelta T cells is not MHC-restricted and dual reactivity against CMV and tumors has been described.


Beug, S. T., et al. (2014). “Smac mimetics and innate immune stimuli synergize to promote tumor death.” Nat Biotechnol 32(2): 182-190. PubMed

Smac mimetic compounds (SMC), a class of drugs that sensitize cells to apoptosis by counteracting the activity of inhibitor of apoptosis (IAP) proteins, have proven safe in phase 1 clinical trials in cancer patients. However, because SMCs act by enabling transduction of pro-apoptotic signals, SMC monotherapy may be efficacious only in the subset of patients whose tumors produce large quantities of death-inducing proteins such as inflammatory cytokines. Therefore, we reasoned that SMCs would synergize with agents that stimulate a potent yet safe “cytokine storm.” Here we show that oncolytic viruses and adjuvants such as poly(I:C) and CpG induce bystander death of cancer cells treated with SMCs that is mediated by interferon beta (IFN-beta), tumor necrosis factor alpha (TNF-alpha) and/or TNF-related apoptosis-inducing ligand (TRAIL). This combinatorial treatment resulted in tumor regression and extended survival in two mouse models of cancer. As these and other adjuvants have been proven safe in clinical trials, it may be worthwhile to explore their clinical efficacy in combination with SMCs.


DeBerge, M. P., et al. (2014). “Soluble, but not transmembrane, TNF-alpha is required during influenza infection to limit the magnitude of immune responses and the extent of immunopathology.” J Immunol 192(12): 5839-5851. PubMed

TNF-alpha is a pleotropic cytokine that has both proinflammatory and anti-inflammatory functions during influenza infection. TNF-alpha is first expressed as a transmembrane protein that is proteolytically processed to release a soluble form. Transmembrane TNF-alpha (memTNF-alpha) and soluble TNF-alpha (solTNF-alpha) have been shown to exert distinct tissue-protective or tissue-pathologic effects in several disease models. However, the relative contributions of memTNF-alpha or solTNF-alpha in regulating pulmonary immunopathology following influenza infection are unclear. Therefore, we performed intranasal influenza infection in mice exclusively expressing noncleavable memTNF-alpha or lacking TNF-alpha entirely and examined the outcomes. We found that solTNF-alpha, but not memTNF-alpha, was required to limit the size of the immune response and the extent of injury. In the absence of solTNF-alpha, there was a significant increase in the CD8(+) T cell response, including virus-specific CD8(+) T cells, which was due in part to an increased resistance to activation-induced cell death. We found that solTNF-alpha mediates these immunoregulatory effects primarily through TNFR1, because mice deficient in TNFR1, but not TNFR2, exhibited dysregulated immune responses and exacerbated injury similar to that observed in mice lacking solTNF-alpha. We also found that solTNF-alpha expression was required early during infection to regulate the magnitude of the CD8(+) T cell response, indicating that early inflammatory events are critical for the regulation of the effector phase. Taken together, these findings suggest that processing of memTNF-alpha to release solTNF-alpha is a critical event regulating the immune response during influenza infection.


Perng, O. A., et al. (2014). “The degree of CD4+ T cell autoreactivity determines cellular pathways underlying inflammatory arthritis.” J Immunol 192(7): 3043-3056. PubMed

Although therapies targeting distinct cellular pathways (e.g., anticytokine versus anti-B cell therapy) have been found to be an effective strategy for at least some patients with inflammatory arthritis, the mechanisms that determine which pathways promote arthritis development are poorly understood. We have used a transgenic mouse model to examine how variations in the CD4(+) T cell response to a surrogate self-peptide can affect the cellular pathways that are required for arthritis development. CD4(+) T cells that are highly reactive with the self-peptide induce inflammatory arthritis that affects male and female mice equally. Arthritis develops by a B cell-independent mechanism, although it can be suppressed by an anti-TNF treatment, which prevented the accumulation of effector CD4(+) Th17 cells in the joints of treated mice. By contrast, arthritis develops with a significant female bias in the context of a more weakly autoreactive CD4(+) T cell response, and B cells play a prominent role in disease pathogenesis. In this setting of lower CD4(+) T cell autoreactivity, B cells promote the formation of autoreactive CD4(+) effector T cells (including Th17 cells), and IL-17 is required for arthritis development. These studies show that the degree of CD4(+) T cell reactivity for a self-peptide can play a prominent role in determining whether distinct cellular pathways can be targeted to prevent the development of inflammatory arthritis.


Walsh, K. B., et al. (2014). “Animal model of respiratory syncytial virus: CD8+ T cells cause a cytokine storm that is chemically tractable by sphingosine-1-phosphate 1 receptor agonist therapy.” J Virol 88(11): 6281-6293. PubMed

The cytokine storm is an intensified, dysregulated, tissue-injurious inflammatory response driven by cytokine and immune cell components. The cytokine storm during influenza virus infection, whereby the amplified innate immune response is primarily responsible for pulmonary damage, has been well characterized. Now we describe a novel event where virus-specific T cells induce a cytokine storm. The paramyxovirus pneumonia virus of mice (PVM) is a model of human respiratory syncytial virus (hRSV). Unexpectedly, when C57BL/6 mice were infected with PVM, the innate inflammatory response was undetectable until day 5 postinfection, at which time CD8(+) T cells infiltrated into the lung, initiating a cytokine storm by their production of gamma interferon (IFN-gamma) and tumor necrosis factor alpha (TNF-alpha). Administration of an immunomodulatory sphingosine-1-phosphate (S1P) receptor 1 (S1P1R) agonist significantly inhibited PVM-elicited cytokine storm by blunting the PVM-specific CD8(+) T cell response, resulting in diminished pulmonary disease and enhanced survival. IMPORTANCE: A dysregulated overly exuberant immune response, termed a “cytokine storm,” accompanies virus-induced acute respiratory diseases (VARV), is primarily responsible for the accompanying high morbidity and mortality, and can be controlled therapeutically in influenza virus infection of mice and ferrets by administration of sphingosine-1-phosphate 1 receptor (S1P1R) agonists. Here, two novel findings are recorded. First, in contrast to influenza infection, where the cytokine storm is initiated early by the innate immune system, for pneumonia virus of mice (PVM), a model of RSV, the cytokine storm is initiated late in infection by the adaptive immune response: specifically, by virus-specific CD8 T cells via their release of IFN-gamma and TNF-alpha. Blockading these cytokines with neutralizing antibodies blunts the cytokine storm and protects the host. Second, PVM infection is controlled by administration of an S1P1R agonist.


Weinlich, R., et al. (2013). “Protective roles for caspase-8 and cFLIP in adult homeostasis.” Cell Rep 5(2): 340-348. PubMed

Caspase-8 or cellular FLICE-like inhibitor protein (cFLIP) deficiency leads to embryonic lethality in mice due to defects in endothelial tissues. Caspase-8(-/-) and receptor-interacting protein kinase-3 (RIPK3)(-/-), but not cFLIP(-/-) and RIPK3(-/-), double-knockout animals develop normally, indicating that caspase-8 antagonizes the lethal effects of RIPK3 during development. Here, we show that the acute deletion of caspase-8 in the gut of adult mice induces enterocyte death, disruption of tissue homeostasis, and inflammation, resulting in sepsis and mortality. Likewise, acute deletion of caspase-8 in a focal region of the skin induces local keratinocyte death, tissue disruption, and inflammation. Strikingly, RIPK3 ablation rescues both phenotypes. However, acute loss of cFLIP in the skin produces a similar phenotype that is not rescued by RIPK3 ablation. TNF neutralization protects from either acute loss of caspase-8 or cFLIP. These results demonstrate that caspase-8-mediated suppression of RIPK3-induced death is required not only during development but also for adult homeostasis. Furthermore, RIPK3-dependent inflammation is dispensable for the skin phenotype.


Mohr, E., et al. (2010). “IFN-{gamma} produced by CD8 T cells induces T-bet-dependent and -independent class switching in B cells in responses to alum-precipitated protein vaccine.” Proc Natl Acad Sci U S A 107(40): 17292-17297. PubMed

Alum-precipitated protein (alum protein) vaccines elicit long-lasting neutralizing antibody responses that prevent bacterial exotoxins and viruses from entering cells. Typically, these vaccines induce CD4 T cells to become T helper 2 (Th2) cells that induce Ig class switching to IgG1. We now report that CD8 T cells also respond to alum proteins, proliferating extensively and producing IFN-gamma, a key Th1 cytokine. These findings led us to question whether adoptive transfer of antigen-specific CD8 T cells alters the characteristic CD4 Th2 response to alum proteins and the switching pattern in responding B cells. To this end, WT mice given transgenic ovalbumin (OVA)-specific CD4 (OTII) or CD8 (OTI) T cells, or both, were immunized with alum-precipitated OVA. Cotransfer of antigen-specific CD8 T cells skewed switching patterns in responding B cells from IgG1 to IgG2a and IgG2b. Blocking with anti-IFN-gamma antibody largely inhibited this altered B-cell switching pattern. The transcription factor T-bet is required in B cells for IFN-gamma-dependent switching to IgG2a. By contrast, we show that this transcription factor is dispensable in B cells both for IFN-gamma-induced switching to IgG2b and for inhibition of switching to IgG1. Thus, T-bet dependence identifies distinct transcriptional pathways in B cells that regulate IFN-gamma-induced switching to different IgG isotypes.