The IL-4 is purified by proprietary chromatographic techniques.
IL-4 was lyophilized from a 0.2µm filtered concentrated (1mg/ml) solution in PBS, pH 7.4.
It is recommended to reconstitute the lyophilized IL-4 in sterile 18MΩ-cm H2O not less than 100µg/ml, which can then be further diluted to other aqueous solutions.
Lyophilized Interleukin-4 although stable at room temperature for 3 weeks, should be stored desiccated below -18°C. Upon reconstitution IL-4 should be stored at 4°C between 2-7 days and for future use below -18°C.
For long term storage it is recommended to add a carrier protein (0.1% HSA or BSA).
Please prevent freeze-thaw cycles.
(a) Analysis by RP-HPLC.
(b) Analysis by SDS-PAGE.
Amino acid sequence
1. UV spectroscopy at 280 nm using the absorbency value of 0.594 as the extinction coefficient for a 0.1% (1mg/ml) solution. This value is calculated by the PC GENE computer analysis program of protein sequences (IntelliGenetics).
2. Analysis by RP-HPLC, using a standard solution of IL-4 as a Reference Standard.
1. Title:Glutamate Released by Dendritic Cells as a Novel Modulator of T Cell Activation1
Publication:The Journal of Immunology, 2006, 177: 6695-6704.Copyright © 2006 by The American Association of Immunologists, Inc.
2.Title:CD26, adenosine deaminase, and adenosine receptors mediate costimulatory signals in the immunological synapse
Publication:Published online before print June 27, 2005, doi: 10.1073/pnas.0501050102 PNAS July 5, 2005 vol. 102 no. 27 9583-9588
3.Title:A Network Modeling Approach to Analysis of the Th2 Memory Responses Underlying Human Atopic Disease
Publication:doi: 10.4049/?jimmunol.0804125 The Journal of Immunology May 15, 2009 vol. 182 no. 10 6011-6021
4.Title:Defective IL-10 production in severe phenotypes of Crohn’s disease
Publication:Published online before print February 23, 2009, doi: 10.1189/jlb.1108698 May 2009 Journal of Leukocyte Biology vol. 85 no. 5 896-903
5.Title:Adenosine deaminase potentiates the generation of effector, memory, and regulatory CD4+ T cells .
Publication:Published online before print October 19, 2010, doi: 10.1189/jlb.1009696 January 2011 Journal of Leukocyte Biology vol. 89 no. 1 127-136 .
6.Title:Glutamate Released by Dendritic Cells as a Novel Modulator of T Cell Activation/
Safety Data Sheet
IL4 is produced mainly by a subpopulation of activated T-cells (Th2) which are the biologically most active helper cells for B-cells and which also secrete IL5 and IL6. Another subpopulation (Th1) also produces IL4 albeit to a lesser extent. Non-T/Non-B-cells of the lineage of mast cells also produce IL4.
IL4 is a protein of 129 amino acids (20 kDa) that is synthesized as a precursor containing a hydrophobic secretory signal sequence of 24 amino acids. IL4 is glycosylated at two arginine residues (positions 38 and 105) and contains six cysteine residues involved in disulfide bond formation. The disulfide bonds are essential for biological activity. Some glycosylation variants of IL4 have been described that differ in their biological activities. A comparison of murine and human IL4 shows that both proteins only diverge at positions 91-128.
An IL4 variant, Y124D, in which Tyr124 of the recombinant human protein (see also: Recombinant cytokines) is substituted by an aspartic acid residue, binds with high affinity to the IL4 receptor (KD = 310 pM). This variant is a powerful antagonist for the IL4 receptor system. It retains no detectable proliferative activity for T-cells and competitively inhibits IL4 dependent T-cell proliferation (K(i) = 620 pM) (see also: Factor-dependent cell lines). The existence of this mutant demonstrates that high affinity binding and signal generation can be uncoupled efficiently in a ligand. Y124D also acts as a powerful antagonist for the IL13 receptor. A naturally occurring alternative splice variant of IL4, designated IL4-delta-2, lacks Exon 2 sequences and has been found to act as a naturally occurring antagonist of IL4 actions.
The human IL4 gene contains four exons and has a length of approximately 10 kb. It maps to chromosome 5q23-31 (see also: 5q minus syndrome). The murine gene maps to chromosome 11. The IL4 gene is in close proximity to other genes encoding hematopoietic growth factors (see: GM-CSF, M-CSF, IL3, IL5). The distance between the IL4 and the IL5 gene is approximately 90-240 kb.
At the nucleotide level the human and the murine IL4 gene display approximately 70 % homology. The 5' region of the IL4 contains several sequence elements, designated CLE (conserved lymphokine element), that are binding sites for transcription factors controlling the expression of this and other genes (see also: gene expression). A sequence motif, called P sequence (CGAAAATTTCC) in the 5' region of the human IL4 gene (positions -79 - -69) is the binding site for a nuclear factor, called NF(P), mediating the response to T-cell activation signals (see also: cell activation).
The biological activities of IL4 are mediated by a specific receptor (Kdis = 20-100 pM) which is expressed at densities of 100-5000 copies/cell. The extracellular domain of the IL4 receptor is related to the receptors for Epo, IL6, and the beta chain of the IL2 receptor. It has been given the name CD124.
Two types of IL4 receptor (IL4R) exist: the type 1 receptor is a heterodimer consisting of CD132 and IL4R-alpha. The type 2 receptor is a heterodimer consisting of IL4R-alpha and IL13R-alpha-1.
The cDNA for the murine IL4 receptor encodes a transmembrane protein of 810 amino acids (including a secretory signal sequence). This receptor has a large intracellular domain of 553 amino acids. The human receptor has an extracellular domain of 207 amino acids, a transmembrane domain of 24 residues, and a large intracellular domain of 569 amino acids.
The IL4 receptor has been shown recently to contain the gamma subunit of the IL2 receptor as a signaling component. This gamma subunit is also associated with the receptors for IL4 and IL7 and probably also of IL13. These findings can explain the severity of the immune defect in X-linked immunodeficiency.