Interleukin 4 (IL-4) is a pleiotropic cytokine produced primarily by activated T lymphocytes, mast cells and basophils. 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. 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. Human IL-4 also shares approximately 30% amino acid sequence identity to human IL-13 and the two cytokines exhibit overlapping biological activities. 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.

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.

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. 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 .

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. 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. Soluble IL-4 R has been to shown to bind IL-4 with high affinity in solution.

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 combination of 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.

Figure 1:Schematic diagram of the assay

1. Aluminium pouches with a Microwell Plate coated with monoclonal antibody to human IL-4 (812)

2. 2 vials human IL-4 Standard lyophilized, 1000 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. 4 pieces Adhesive Films

12. package insert

NOTE: [96 Tests]

Unopened Kit：Store at 2 -8° C. Do not use past kit expiration date. opened/ReconstitutedReagents：Please refer to the datasheets for detail information.

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5. McKenzie, A.N.J. and G. Zurawski (1994) in Guidebook to Cytokines and Their Receptors,N.A. Nicola ed., Oxford Univ. Press, New York, p. 92.

6. Zurawski, G. and J.E. de Vries (1994) Immunol. Today 15:19.

7. Prete,G.et al.(1988) J. Immunol. 4193-4198