- Interleukin 4 (IL-4) is a pleiotropic cytokine produced primarily by activated T lymphocytes, mast cells and basophils (1-3). The sequence of human IL-4 cDNA predicts a 153 amino acid (aa) residue precursor protein containing a 24 aa residue signal peptide that is cleaved to form the mature protein (4). At the amino acid sequence level, mature human IL-4 is approximately 50% identical to mouse IL-4 and there is no species cross-reactivity between the two proteins (1, 2). Human IL-4 also shares approximately 30% amino acid sequence identity to human IL-13 and the two cytokines exhibit overlapping biological activities (5, 6). The gene for IL-4 has been mapped to human chromosome 5q, in close proximity to the genes for IL-3, IL-5, IL-13 and GM-CSF (1, 2).
IL-4 has multiple immune response-modulating functions on a variety of cell types. It is an important regulator of isotype switching, inducing IgE production in B lymphocytes. It is an important modulator of the differentiation of precursor T helper cells to the TH2 subset that mediates humoral immunity and modulates antibody production. In addition, IL-4 has also been shown to have anti-tumor activity both in vivo and in vitro (1-3).
The biological effects of IL-4 are mediated by specific cell surface receptor complexes. One type of functional IL-4 receptor complex consists of the IL-4-binding subunit (IL-4 R) and a second chain, designated the common c chain because it has also been identified as a component of the receptor complexes for IL-2, IL-7, IL-9 and IL-15 (7-9). A second type of functional IL-4 receptor complex, consisting of the IL-4 R and the more recently cloned IL-13 R , has also been proposed (10, 11).
Although IL-4 R does not bind IL-13 directly, it has been shown to complex with the low-affinity IL-13 R to form the functional high-affinity receptor complex for IL-13 (11, 12). In addition to the membrane-bound form of IL-4 R, a naturally occurring soluble form of IL-4 R has been identified in human and mouse biological fluids and in mouse cell culture supernates (13-15). Soluble IL-4 R has been to shown to bind IL-4 with high affinity in solution.
For the quantitative determination of human interleukin 4 (IL-4) concentrations in cell culture supernates, serum, and plasma.
Principle of the assay: This assay employs the quantitative sandwich enzyme immunoassay technique. A monoclonal antibody specific for IL-4 has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and any IL-4 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-4 is added to the wells and binds to the combinationof capture antibody-IL-4 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-4 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-4 standard dilutions and IL-4 sample concentration determined.
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- Sample Type: cell culture supernates, serum, and plasma
- Reagents: REAGENTS
1. Aluminum pouches with a Microwell Plate coated with monoclonal antibody to human IL-4 (8x12)
2. 2 vials human IL-4 Standard lyophilized, 500 pg/ml upon reconstitution
3. 2 vials concentrated Biotin-Conjugate anti-human IL-4 polyclonal antibody
4. 2 vials Streptavidin-HRP solution,
5. 1 bottle Standard /sample Diluent
6. 1 bottle Biotin-Conjugate antibody Diluent
7. 1 bottle Streptavidin-HRP Diluent
8. 1 bottle Wash Buffer Concentrate 20x (PBS with 1% Tween-20)
9. 1 vial Substrate Solution
10. 1 vial Stop Solution
11. 3 pieces Adhesive Films
12. package insert
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