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Sandwich ELISA analysis of Human TNF alpha Biotinylated MAb (Pierce product # M302B) was performed using components of Thermo Fisher Scientific Human TNF alpha Colorimetric ELISA Kit (Pierce product # EH3TNFA) by loading 50ul per well of Recombinant Human TNF alpha in octuplicate at 1000, 500, 250, 125, 62.5, 31.2, 15.6, and 0 pg/ml across a 5µg/ml rabbit anti-Human TNF pre-coated plate and incubated for 1 hour at room temperature. The plate was washed then incubated with 100 µl per well of Human TNF alpha Biotinylated MAb (M302B) in duplicate at 2, 1, 0.5, and 0.25 µg/ml for 1 hour at room temperature. The plate was washed and incubated with 100 µl per well of Ultra Streptavidin-HRP (Pierce product # N504) in all test wells at 1:1,500 for 30 minutes at room temperature. The plate was washed then incubated with 100ul per well of TMB substrate for 30 minutes at room temperature in the dark. The plate was then stopped with 0.16M sulfuric acid and absorbance was 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|
|Storage buffer||PBS with 4% BSA|
|Tested Applications||Dilution *|
|ELISA (ELISA)||Assay Dependent|
|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.
|ELISA (ELISA)||See 16 publications below|
M302B 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 M302B immunogen is recombinant human TNF alpha.
M302B detects TNF Alpha which has a predicted molecular weight of approximately 26 kDa.
The M302B TNFa antibody (biotinylated conjugate of clone 2TNF-H33) has successfully been paired as the detection antibody in a sandwich ELISA with capture antibody M303 (clone 2TNF-H34A) Typical dilutions for sandwich ELISA include 1 µg/ml for coating and 0.125 - 0.25 µg/ml for detection.
Biotinylated antibody M302B (clone 2TNF-H33) and antibody M303 (clone 2TNF-H34A) 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.
Cytokines across the night in chronic fatigue syndrome with and without fibromyalgia.
M302B 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.
M302B 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.
M302B 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.
M302B 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.
M302B 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.
M302B 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.
M302B 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.
M302B 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.
M302B 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.
M302B 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|
Thiothalidomides: novel isosteric analogues of thalidomide with enhanced TNF-alpha inhibitory activity.
M302B was used in ELISA to screen novel isosteric active thalidomide analogues
|Zhu X,Giordano T,Yu QS,Holloway HW,Perry TA,Lahiri DK,Brossi A,Greig NH||Journal of medicinal chemistry (46:5222)||2003|
Enhanced monocyte Th1 cytokine production in HCV-infected cryoglobulinemic patients.
M302B 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.
M302B 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.
M302B 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|
Suppression of TNF-alpha production in human mononuclear cells by an adenosine kinase inhibitor.
M302B was used in ELISA to study the ability of an adenosine kinase inhibitor to suppress TNF-alpha production in human mononuclear cells
|Eigler A,Matschke V,Hartmann G,Erhardt S,Boyle D,Firestein GS,Endres S||Journal of leukocyte biology (68:97)||2000|
Regulation of interleukin-12 by complement receptor 3 signaling.
M302B 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|
APC1 protein; cachectin; TNF superfamily, member 2; TNF, macrophage-derived; TNF, monocyte-derived; TNF-a; tumor necrosis factor alpha; tumor necrosis factor ligand 1F; tumor necrosis factor ligand superfamily member 2; tumor necrosis factor-alpha
DIF; TNF; TNF-alpha; TNFA; TNFSF2; TNLG1F