Mouse IL-12p70 ELISA Kit

Principle of the Assay

This assay employs the quantitative sandwich enzyme immunoassay technique. A monoclonal antibody specific for IL-12p70 has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and any IL-12p70 present is bound by the immobilized antibody. Following incubation unbound samples are removed during a wash step, and then a detection antibody specific for IL-12p70 is added to the wells and binds to the combination of capture antibody- IL-12p70 in sample. Following a wash to remove any unbound combination, and enzyme conjugate is added to the wells. Following incubation and wash steps a substrate is added. A coloured product is formed in proportion to the amount of IL-12p70 present in the sample. The reaction is terminated by addition of acid and absorbance is measured at 450nm. A standard curve is prepared from seven IL-12p70 standard dilutions and IL-12p70 sample concentration determined.

For Use with serum, plasma and cell culture supernatants. For Research Use Only. Not for use in diagnostic procedures.

Target Information

Interleukin 12 (IL-12), also known as natural killer cell stimulatory factor (NKSF) or cytotoxic lymphocyte maturation factor (CLMF), is a pleiotropic cytokine originally identified in the medium of a human B-lymphoblastoid cell line.

Biologically active mouse IL-12 is a disulfide-linked, 70 kDa (p70) heterodimeric glycoprotein composed of a 40 kDa (p40) subunit and a 35 kDa (p35) subunit. While the p40 and p35 subunits by themselves do not have IL-12 activity, the p40 homodimer has been shown to bind the IL-12 receptor and is an IL-12 antagonist. Mature mouse p35 subunit is composed of 193 amino acid (aa) residues and contains seven cysteines plus one potential N-linked glycosylation site. Mature mouse p40 subunit has 313 aa, with 13 cysteines and five potential N-linked glycosylation sites. Mouse p35 and p40 subunits show 63% and 72% aa identity, respectively, to the human p35 and p40 subunits. Although mouse IL-12 is active on both human and mouse cells, human IL-12 is only active on human cells.

IL-12 has been shown to have multiple effects on T lymphocytes and natural killer (NK) cells. Some of these effects include the induction of IFN-γ and TNF production by T and NK cells, the enhancement of cytotoxic activity of T and NK cells and the stimulation of T and NK cell proliferation. IL-12 has also been shown to be a central mediator of the cell-mediated immune response by promoting Th1 development.

Cell surface staining for IL-12 on a human monocytic and mouse macrophage cell line has been reported, suggesting that membrane-associated IL-12 may exist. Cells known to produce IL-12 include macrophages, dendritic cells, monocytes, Langerhans cells, neutrophils, and keratinocytes. Although a human B cell line has been shown to produce IL-12, fresh B cells are apparently not producers of IL-12.

Materials Supplied

Kit Components	96 Wells Quantity/Size
Aluminium pouches with a Microwell Plate coated with monoclonal antibody to mouse IL-12p70 (8﹡12)	1 plate
Mouse IL-12p70 Standard lyophilized, 2000 pg/ml upon reconstitution	2 vials
Concentrated Biotin-Conjugate anti-mouse IL-12p70 monoclonal antibody	2 vials
Streptavidin-HRP solution	2 vials
Standard /sample Diluent	1 bottle
Biotin-Conjugate antibody Diluent	1 bottle
Streptavidin-HRP Diluent	1 bottle
Wash Buffer Concentrate 20x (PBS with 1% Tween-20)	1 bottle
Substrate Solution	1 vial
Stop Solution	1 vial
Adhesive Films	4 pieces
Product data sheet	1 copy

Storage

Store at 2 - 8°C

Performance Characteristics

REPEATABILITY	The coefficient of variation of both intra-assay and inter-assay were less than 10%.
SENSITIVITY	The minimum detectable dose was 7pg/mL.
ASSAY RANGE	15.6-1000 pg/mL
SPECIFICITY	This assay recognizes both natural and recombinant mouse IL-12p70. The factors listed below were prepared at 50ng/ml in Standard /sample Diluent and assayed for cross-reactivity and no significant cross-reactivity or interference was observed.

Factors assayed for cross-reactivity

Recombinant mouse	G-CSF, GM-CSF, IL-1α, IL-1β, IL-2 , IL-3, IL-4, IL-5, IL-6, IL-7, IL-9, IL-10, IL-12p35, IL-12p40 monomer, IL-12p70, IL-23, TNF-α
Recombinant human	G-CSF, IL-6, IL-6sR, IL-12p35, IL-12p40 monomer, IL-12p70, IL-23, GM-CSF

Data Analysis Assistance

We have partnered with MyAssays to offer you an easy to use and versatile tool to analyze the data you receive using our ELISA Kit. Click the link below to be directed to the data analysis tool provided by MyAssays specifically for BSKM1017.

https://www.myassays.com/bioss-mouse-il-12p70-elisa-kit.assay