Description

Nori Ferret TNFa ELISA Kit Summary

Alternative names for TNFa: TNF alpha, TNF-a, tumor necrosis factor alpha

Standard Curve

Kit Components
1. Pre-coated 96-well Microplate
2. Biotinylated Detection Antibody
3. Streptavidin-HRP Conjugate
4. Lyophilized Standards
5. TMB One-Step Substrate
6. Stop Solution
7. 20 x PBS
8. Assay Buffer

Other Materials Required but not Provided:
1. Microplate Reader capable of measuring absorption at 450 nm
2. Log-log graph paper or computer and software for ELISA data analysis
3. Precision pipettes (1-1000 µl)
4. Multi-channel pipettes (300 µl)
5. Distilled or deionized water

Protocol Outline
1. Prepare all reagents, samples and standards as instructed in the datasheet.
2. Add 100 µl of Standard or samples to each well and incubate 1 h at RT.
3. Add 100 µl of Working Detection Antibody to each well and incubate 1 h at RT.
4. Add 100 µl of Working Streptavidin-HRP to each well and incubate 20 min at RT.
5. Add 100 µl of Substrate to each well and incubate 5-30 min at RT.
6. Add 50 µl of Stop Solution to each well and read at 450 nm immediately.

Background: 

TNF-α, the prototypical member of the TNF protein superfamily, is a homotrimeric type-II membrane protein (1,2). Membrane bound TNF-α is cleaved by the metalloprotease TACE/ADAM17 to generate a soluble homotrimer (2). Both membrane and soluble forms of TNF-α are biologically active. TNF-α is produced primarily by macrophages, but it is produced also by a broad variety of cell types including lymphoid cells, mast cells, endothelial cells, cardiac myocytes, adipose tissue, fibroblasts, and neuronal tissue (1). Cellular response to TNF-α is mediated through interaction with receptors TNF-R1 and TNF-R2 and results in activation of pathways that favor both cell survival and apoptosis depending on the cell type and biological context. Activation of kinase pathways (including JNK, ERK (p44/42), p38 MAPK and NF-κB) promotes the survival of cells, while TNF-α mediated activation of caspase-8 leads to programmed cell death (1,2). TNF-α plays a key regulatory role in inflammation and host defense against bacterial infection, notably Mycobacterium tuberculosis (3). TNF-α causes many of the clinical problems associated with autoimmune disorders such as rheumatoid arthritis, ankylosing spondylitis, inflammatory bowel disease, psoriasis, hidradenitis suppurativa and refractory asthma. The role of TNF-α in autoimmunity is underscored by blocking TNF-α action to treat rheumatoid arthritis and Crohn’s disease (1,2,4).

References

1. Aggarwal, B.B. (2003) Nat Rev Immunol 3, 745-56.
2. Hehlgans, T. and Pfeffer, K. (2005) Immunology 115, 1-20.
3. Lin, P.L. et al. (2007) J Investig Dermatol Symp Proc 12, 22-5.
4. Brennan, F.M. and McInnes, I.B. (2008) J Clin Invest 118, 3537-45.

Product Citation

40.        Zucca E et al. (2016) Evaluation of amniotic mesenchymal cell derivatives on cytokine production in equine
alveolar macrophages: an in vitro approach to lung inflammation. Stem Cell Research and Therapy 7:137.   Article
Product used and cited: Nori Equine TNFα/IL-6/TGFβ1 ELISA kits. Impact factor: 4.731.

91.        Witonsky S et al. (2019) Can levamisole upregulate the equine cell-mediated macrophage (M1) dendritic
cell (DC1) T-helper 1 (CD4 Th1) T-cytotoxic (CD8) immune response in vitro? J Vet Internal Med 33(2):889-96.
Impact factor: 2.286.
Products used and cited: Nori Equine IL-6 ELISA Kit.

37.        Siemieniuch MJ et al. (2016) Type of inflammation differentially affects expression of interleukin 1β and 6, tumor
necrosis factor-α and toll-like receptors in subclinical endometritis in mares. PLOS ONE 11(5):e0154934. Doi:10.137
/journal.pone.0154934. Impact factor: 2.806.
Product used and cited: Nori Equine IL-6 ELISA kit.

48. Water EVD et al. (2016) The preventive effects of two nutraceuticals on experimentally induced acute synovitis. Equine Veterinary Journal DOI:10.1111/evj.12629. Impact factor: 2.40.

Product used and cited: Nori Equine IL-6 ELISA Kit GR106001, GR106088.

81.        Abass M et al. (2018) Local mepivacaine before castration of horses under medetomidine
isoflurane balanced anesthesia is effective to reduce perioperative nociception and cytokine release.
Equine Veterinary Journal, 50:733-738. Impact factor: 2.022
Produce used and cited: Nori Equine IL-6 and TNF-alpha ELISA Kit.

96.        Broeckx SY et al. (2019) The use of equine chondrogenic-induced mesenchymal stem cells as a treatment for osteoarthritis:
A randomized, double-blinded, placebo-controlled proof-of-concept study. Equine Vet J 51(6): 787-94. Impact factor: 2.022.
Products used and cited: Nori Equine IL-6 and TGFβ3 ELISA Kits.

110.        Hussein H et al. (2016) Cathepsin K inhibition renders equine bone marrow nucleated cells hypo-responsive
to LPS and unmethylated CpG stimulation in vitro. Comparative Immunology, Microbiology and Infectious Diseases
45: 40-7. Impact factor: 2.05.
Products used and cited: Nori Equine IL-1β, IL-6 and TNFα ELISA Kits.

126. Broeckx SY et al. (2019) Evaluation of an osteochondral fragment-groove procedure for induction of
metacarpophalangeal joint osteoarthritis in horses. Am J Vet Res 80(3):246-58. Impact factor: 1.07.
         Products used and cited: Nori Equine IL-6 and TGFb3 ELISA Kits (GR106001, GR106134).