Direct ELISA analysis of TNF alpha was performed by coating wells of a 96-well plate with 100ul per well of human recombinant protein TNF alpha (Product #RTNFAI) diluted in carbonate/bicarbonate buffer (Product # 28382) at a concentration of 1ug/ml overnight at 4C. Wells of the plate were washed, blocked with starting blocking buffer (Product # 37538), and incubated with 100ul per well of a mouse anti-TNF alpha monoclonal antibody (Product # M303) at a dilution of 1:50, 1:100, 1:200, 1:400, 1:800, 1:1600 and 1:3200 for 90 minutes at 37C. The plate was washed, then incubated with 100ul per well of a HRP-conjugated goat anti-mouse IgG secondary antibody (Product # 62-6520) at a dilution of 1:2000 for 90 minutes at 37C. Detection was performed using 1-Step Ultra TMB substrate (Product # 34028) for 5-10 minutes at room temperature in the dark. The reaction was stopped with Stop solution (Product #N600), and absorbances were read on a spectrophotometer at 450-550nm.
|Tested species reactivity||Human|
|Published species reactivity||Human|
|Host / Isotype||Mouse / IgG1, kappa|
|Immunogen||Recombinant human TNF alpha|
|Tested Applications||Dilution *|
|Western Blot (WB)||Assay Dependent|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
M303 targets TNF Alpha in ELISA, and WB applications and shows reactivity with Human samples. The M303 antibody (clone 2TNF-H34A) has successfully been paired as the coating antibody in a sandwich ELISA with detection antibody M302B (biotinylated conjugate of clone 2TNG-H33).
The M303 immunogen is recombinant human TNF alpha.
M303 detects TNF Alpha which has a predicted molecular weight of approximately 26 kDa.
The M303 TNFa antibody (clone 2TNF-H34A) has successfully been paired as the coating antibody in a sandwich ELISA with detection antibody M302B (biotinylated conjugate of clone 2TNF-H33) Typical dilutions for sandwich ELISA include 1 µg/ml for coating and 0.125 - 0.25 µg/ml for detection.
Antibody M303 (clone 2TNF-H34A) and biotinylated antibody M302B (clone 2TNF-H33) have successfully been used in combination with recombinant TNFa protein STNFA in ELISA applications.
This gene encodes a multifunctional proinflammatory cytokine that belongs to the tumor necrosis factor (TNF) superfamily. This cytokine is mainly secreted by macrophages. It can bind to, and thus functions through its receptors TNFRSF1A/TNFR1 and TNFRSF1B/TNFBR. This cytokine is involved in the regulation of a wide spectrum of biological processes including cell proliferation, differentiation, apoptosis, lipid metabolism, and coagulation. This cytokine has been implicated in a variety of diseases, including autoimmune diseases, insulin resistance, and cancer. Knockout studies in mice also suggested the neuroprotective function of this cytokine.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.
Protein kinase c¿ regulates the expression of complement receptor Ig in human monocyte-derived macrophages.
M303 was used in blocking or activating experiment to examine the role of PKCalpha in CRIg expression in human monocyte-derived macrophages
|Ma Y,Usuwanthim K,Munawara U,Quach A,Gorgani NN,Abbott CA,Hii CS,Ferrante A||Journal of immunology (Baltimore, Md. : 1950) (194:2855)||2015|
Cytokines across the night in chronic fatigue syndrome with and without fibromyalgia.
M303 was used in ELISA to study the temporal cytokine profiles in chronic fatigue syndrome with and without fibromyalgia during sleep
|Nakamura T,Schwander SK,Donnelly R,Ortega F,Togo F,Broderick G,Yamamoto Y,Cherniack NS,Rapoport D,Natelson BH||Clinical and vaccine immunology : CVI (17:582)||2010|
Anandamide suppresses proliferation and cytokine release from primary human T-lymphocytes mainly via CB2 receptors.
M303 was used in ELISA to investigate the mechanism of human T cell regulation by anandamide
|Cencioni MT,Chiurchiù V,Catanzaro G,Borsellino G,Bernardi G,Battistini L,Maccarrone M||PloS one (5:null)||2010|
Efficient immobilization of a ligand antibody with high antigen-binding activity by use of a polystyrene-binding peptide and an intelligent microtiter plate.
M303 was used in ELISA to investigate an immobilization method for efficient and highly sensitive sandwich ELISA system with smaller amounts of ligand antibodies
|Kumada Y,Hamasaki K,Shiritani Y,Ohse T,Kishimoto M||Journal of biotechnology (142:135)||2009|
Sesquiterpene lactones induce distinct forms of cell death that modulate human monocyte-derived macrophage responses.
M303 was used in ELISA to study the role of sesquiterpene lactones in human monocyte-derived macrophage responses
|López-Antón N,Hermann C,Murillo R,Merfort I,Wanner G,Vollmar AM,Dirsch VM||Apoptosis : an international journal on programmed cell death (12:141)||2007|
Protein microarray platform for the multiplex analysis of biomarkers in human sera.
M303 was used in ELISA to develop a protein microarray platform for the multiplex analysis of biomarkers in human sera
|Urbanowska T,Mangialaio S,Zickler C,Cheevapruk S,Hasler P,Regenass S,Legay F||Journal of immunological methods (316:1)||2006|
The immunostimulant RU41740 from Klebsiella pneumoniae activates human cells in whole blood to potentially stimulate innate and adaptive immune responses.
M303 was used in ELISA to investigate the role of immunostimulant RU41740 from Klebsiella pneumoniae for innate and adaptive immune responses in human cells from whole blood
|Pedraza-Sánchez S,González-Hernández Y,Escobar-Gutiérrez A,Ramachandra L||International immunopharmacology (6:635)||2006|
Long-term treatment with etanercept significantly reduces the number of proinflammatory cytokine-secreting peripheral blood mononuclear cells in patients with rheumatoid arthritis.
M303 was used in ELISA to study the effect of etanercept treatment on proinflammatory cytokine-secreting peripheral blood mononuclear cells in rheumatoid arthritic patients
|Schotte H,Schlüter B,Willeke P,Mickholz E,Schorat MA,Domschke W,Gaubitz M||Rheumatology (Oxford, England) (43:960)||2004|
Thalidomide-based TNF-alpha inhibitors for neurodegenerative diseases.
M303 was used in ELISA to characterize the thalidomide as tumor necross factor-alpha inhibitors and its possible therapeutic effects on neurodegenerative diseases
|Greig NH,Giordano T,Zhu X,Yu QS,Perry TA,Holloway HW,Brossi A,Rogers JT,Sambamurti K,Lahiri DK||Acta neurobiologiae experimentalis (64:1)||2004|
Host factors impacting the innate response in humans to the candidate adjuvants RC529 and monophosphoryl lipid A.
M303 was used in ELISA to study the effect of genotypes on the innate response against the candidate adjuvants RC529 and monophosphoryl lipid A
|Tiberio L,Fletcher L,Eldridge JH,Duncan DD||Vaccine (22:1515)||2004|
Generation of three-dimensional pannus-like tissues in vitro from single cell suspensions of synovial fluid cells from arthritis patients.
M303 was used in ELISA to investigate the generation of three-dimensional pannus-like tissues in vitro from single cell suspensions of synovial fluid cells from arthritis patients
|Solomon S,Masilamani M,Mohanty S,Schwab JE,Boneberg EM,Illges H||Rheumatology international (24:71)||2004|
Activation of influenza virus-specific CD4+ and CD8+ T cells: a new role for plasmacytoid dendritic cells in adaptive immunity.
M303 was used in ELISA to investigate the role of plasmacytoid dendritic cells (pDCs) in adaptive immunity
|Fonteneau JF,Gilliet M,Larsson M,Dasilva I,Münz C,Liu YJ,Bhardwaj N||Blood (101:3520)||2003|
Enhanced monocyte Th1 cytokine production in HCV-infected cryoglobulinemic patients.
M303 was used in ELISA to investigate the augmented production of Th1-type cytokines in HCV-infected cryoglobulinemic patients
|Loffreda S,Muratori P,Muratori L,Mele L,Bianchi FB,Lenzi M||Journal of hepatology (38:230)||2003|
Enhanced interleukin-12 and CD40 ligand activities but reduced Staphylococcus aureus Cowan 1-induced responses suggest a generalized and progressively impaired type 1 cytokine pattern for human schistosomiasis.
M303 was used in ELISA to study the role of type 1 cytokine in chronic hepatosplenic schistosomiasis
|Montenegro SM,Abath FG,Domingues AL,Melo WG,Morais CN,Coutinho EM,Mahanty S,Wynn TA||Infection and immunity (70:5903)||2002|
Interferon--gamma control of EBV-transformed B cells: a role for CD8+ T cells that poorly kill EBV-infected cells.
M303 was used in ELISA to study the role of inteferon gamma in EBV-transformed B cells
|Shi Y,Lutz CT||Viral immunology (15:213)||2002|
Regulation of interleukin-12 by complement receptor 3 signaling.
M303 was used in ELISA to assess the interferon-alpha production in peripheral blood mononuclear cell
|Marth T,Kelsall BL||The Journal of experimental medicine (185:1987)||1997|