Dendritics generated rat monoclonal antibody (mAb) that recognizes mouse plasmacytoid dendritic cells (pDCs). The target molecule was found to be BST2 (bone marrow stromal cell antigen 2). This antibody, named 120G8, stains a small subset of CD11c-low spleen cells with high specificity. This population produces high amounts of IFNα upon in vitro viral stimulation. Both ex vivo- and in vitro-derived 120G8+ cells display a phenotype identical with that of mouse pDCs (B220-high Ly6C-high Gr1-low CD11b- CD11c-low). Mice treated with 120G8 mAb are depleted of B220-high Ly6C-high CD11c-low cells and have a much reduced ability to produce IFNα in response to in vivo CpG stimulation. mAb 120G8 stains all and only B220-high Ly6C-high CD11c-low pDC in all lymphoid organs. Immunohistochemical studies performed with this mAb indicate that pDC are
located in the T cell area of spleen, lymph nodes, and Peyer’s patches. Using 120G8 mAb in immunofluorescence studies demonstrates mouse strain- and organ-specific differences in the frequency of pDCs and other DC subsets (Asselin-Paturel C et al, 2003 ; J. Immunol., 172:6466; Blasius AI, 2006, J. Immunol., 177:3260 ; Goubier A et Al, 2008, Immunity, 29:464-475).
Specificity: mouse pDCs/IFN producing cells (IPC) (extracellular domain)
Immunogen: mouse plasmatocytoid DCs (pDCs)
Species cross-reactivity: nd
Applications tested: Flow cytometry, in vivo depletion, IHC
Usage recommendation: This monoclonal antibody may be used between at 1-10 µg/ml. For pDCs in vivo depletion in Balb /c mice, mAb 120G8 was used between 50-200 µg /
At Absolute Antibody we have developed a collection of recombinant engineered antibodies against clinically relevant proteins, including homologues of current therapeutic targets. We can match the antibody species to the host organism and tailor the effector function, similar to how pharmaceutical companies develop human therapeutics.
Antibodies against proteins involved in co-stimulation and other aspects of immune cell regulation are of particular interest to therapeutics developers. Some have already entered the clinic, with more in the development pipeline. However, many aspects of immune cell signalling are still unknown, and researchers require ever more advanced tools to tap into this potential.
At Absolute Antibody, we use recombinant technology to provide superior monoclonal antibody reagents at competitive prices. In particular we can modify antibody species and isotype for greater flexibility in vivo, for example we can readily generate mouse-anti-mouse or rat-anti-rat antibodies.
Why go recombinant?
Because of their recombinant manufacture our antibodies show minimal batch-to-batch variability and have potential for customisation. We can convert any antibody into any format allowing us to offer each specificity in a range of species, isotypes and subtypes. This means our customers may ‘build’ an antibody to best suit their experiment.
Choose primary antibody format to suit your secondary reagent
Choose antibody species to be compatible with your model organism
Choose antibody isotype to investigate your chosen host responses (includes IgM and all IgG subtypes)
Choose from a range of custom engineering options such as our Fc Silent format with reduced FcR binding to remove Fc receptor function, or other such formats found in the literature (e.g. IgG1-LALA, IgG1-D265A)
Choose one of our listed antibodies, or apply our recombinant technology to your own clone. All our antibody services are royalty-free.
Although the immune checkpoint protein B7-H1/PD-L1 is a commonly used cancer biomarker, controversy remains over the predictive and prognostic utility of immunohistochemical methods. Download our white paper which shows that we can reliably measure B7-H1/PD-L1 in conditioned media supernates and cell lysates from a number of cancer cell lines.
Data from our white paper indicate that:
We can reliably measure B7-H1/PD-L1 in a number of human cancer cell lines
Soluble B7-H1/PD-L1 expression levels are coincident with loss of PTEN tumor suppressor or increased activation of the phosphoinositide 3-kinase pathway
The immune system fights off pathogens, but this defensive force can be pathogenic itself when hyperactive, resulting in autoimmune diseases such as lupus and multiple sclerosis. Consequently, the body has developed multiple mechanisms to suppress the immune system when necessary.
One method of immunosuppression is the PD-1 pathway. This pathway is activated in response to the mobilization of the immune system. The receptor PD-1 is expressed on the surface of activated lymphocytes. Similarly, its ligand, PD-L1, is expressed by antigen-presenting cells in response to cytokine signaling. When PD-L1 is bound to PD-1, downstream signaling undoes the phosphorylation events associated with activation, thereby reverting lymphocytes to an inactive state [1, 2].
Tumor cells take advantage of the PD-1 pathway to evade the immune system . Consequently, many pharmaceutical companies have been developing drugs to inhibit PD-1 and PD-L1. In clinical trials, many patients have shown strong responses to these therapies [4-10]. For these drugs to be most effective, a high number of CD8+ T cells must already be at the tumor site, ready to be mobilized after inhibition of the PD-1 pathway .Recent investigations have uncovered promising methods of increasing the efficacy of PD1 inhibitors. One method is combining PD1 inhibitors with other drugs, such as HDAC inhibitors  and anti-CTLA4 antibodies . Another method is using biomarkers to predict response to therapy [13,14]. Recent work has also suggested that GSK3 inhibitors can enhance effects of immunotherapy .Though these results have been encouraging, several challenges remain, including the mitigation of autoimmune effects and how to overcome drug resistance.
1. Ohegbulam et al. (2015) Human cancer immunotherapy with antibodies to the PD-1 and PD-L1 pathway. Trends Mol Med. 21:24-33.
2. Haanen, J. (2013) Immunotherapy of Melanoma. EJC Suppl 11:97-105.
3. Yao et al. (2013) Advances in targeting cell surface signaling molecules for immune modulation. Nat Rev Drug Discov. 12:130-146.
4. Topalian, S. et al. (2012) Safety, activity, and immune correlates of anti-PD-1 antibody in cancer. N. Engl. J. Med. 366:2443-2454.
5. Hamid, O. et al. (2013) Safety and tumor responses with lambrolizumab (anti-PD-1) in melanoma. N. Engl. J. Med. 369:134-144.
6. Topalian, S. L. et al. (2012) Safety, activity, and immune correlates of anti-PD-1 antibody in cancer. N. Engl. J. Med. 366:2443–2454
7. Brahmer, J. R. et al. (2012) Safety and activity of anti-PD-L1 antibody in patients with advanced cancer. N. Engl. J. Med. 366:2455–2465
8. Hamid, O. et al. (2013) Safety and tumor responses with lambrolizumab (anti-PD-1) in melanoma. N. Engl. J. Med. 369:134–144
9. Wolchok, J. D. et al. (2013) Nivolumab plus ipilimumab in advanced melanoma. N. Engl. J. Med. 369:122–133
10. Topalian, S. L. et al. (2014) Survival, durable tumor remission, and long-term safety in patients with advanced melanoma receiving nivolumab. J. Clin. Oncol. 32:1020–1030
11. Tumeh, P. et al. (2014) PD-1 blockade induces responses by inhibiting adaptive immune response. Nature. 515:568-71.
12. Woods, D. et al. (2015) HDAC inhibition upregulates PD-1 ligands in melanoma and augments immunotherapy with PD-1 blockade. Cancer Immunol Res 12:1375-85.
13. Chakravarti, N., & Prieto, V. G. (2015). Predictive factors of activity of anti-programmed death-1/programmed death ligand-1 drugs: immunohistochemistry analysis. Translational Lung Cancer Research, 4:743–751.
14. Barak, V. et al. (2015) Assessing response to new treatments and prognosis in melanoma patients, by the biomarker S-100B. Anticancer Res. 35:6755-60.
15. Taylor, A. et al. (2014) Glycogen synthase kinase 3 inactivation drives T-bet-mediated downregulation of co-receptor PD-1 to enhance CD8+ cytolytic T cell responses. Immunity 44:274-86.
Using our Neuron Isolation Kit, you can now rapidly isolate viable and pure neurons from both neonatal and adult mouse brain by depletion of non-neuronal cells. The high purity of the isolated neurons relies on the specificity of the non-neuronal cell surface marker used in the Neuron Isolation kit.
Download this free scientific poster today and see scientific data about the expression of the non-neuronal cell marker:
In different neural cell types
At different mouse brain regions
During different developmental stages (from E14 to P7)
Amyloid β-protein (Aβ42) oligomerization is an early event in Alzheimer’s disease (AD). Current diagnostic methods using sequence-specific antibodies against less toxic fibrillar and monomeric Aβ42 run the risk of overdiagnosis. Hence, conformation-specific antibodies against neurotoxic Aβ42 oligomers have garnered much attention for developing more accurate diagnostics. Antibody 24B3, highly specific for the toxic Aβ42 conformer that has a turn at Glu22 and Asp23, recognizes a putative Aβ42 dimer, which forms stable and neurotoxic oligomers more potently than the monomer. 24B3 significantly rescues Aβ42-induced neurotoxicity, whereas sequence-specific antibodies such as 4G8 and 82E1, which recognizes the N-terminus, do not. The ratio of toxic to total Aβ42 in the cerebrospinal fluid of AD patients is significantly higher than in control subjects as measured by sandwich ELISA using antibodies 24B3 and 82E1. Thus, 24B3 may be useful for AD diagnosis and therapy.
The research group led by Professor Yuichi Oike at Kumamoto University reported that expression of angiopoietin-like protein 2 (ANGPTL2) increases in pathologically-remodeled hearts of mice and humans, while decreased cardiac ANGPTL2 expression occurs in physiological cardiac remodelling induced by endurance training in mice. Mice overexpressing ANGPTL2 in heart show cardiac dysfunction caused by both inactivation of AKT and sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA)2a signalling and decreased myocardial energy metabolism. Conversely, Angptl2 knockout mice exhibit increased left ventricular contractility and upregulated AKT-SERCA2a signalling and energy metabolism. Finally, ANGPTL2-knockdown in mice subjected to pressure overload ameliorates cardiac dysfunction. Overall, these studies suggest that therapeutic ANGPTL2 suppression could antagonize development of heart failure.
Poor quality, unverified antibodies are a major factor underlying the crisis of reproducibility in research, resulting in an estimated $800 million of waste each year and a ten-fold spike in retractions over the past decade.
More antibody validation protocols would identify non-specific reagents – a major source of irreproducible research – with the inclusion of negative controls. This article presents an overview of one such method: siRNA knockdown. The authors outline a general protocol, the knockdown mechanism, and tips for evaluating knockdown experiments.