Mouse IL-5 ELISA Assay

$445.00

The Mouse IL-5 ELISA Assay is intended for the quantitative determination of mouse Interleukin 5 (IL-5) concentrations in cell culture supernates, serum, and plasma. The Mouse Interleukin 5 (IL-5) ELISA Assay Kit is for research use only and not to be used in diagnostic procedures.

Mouse IL-5 ELISA Assay

The Mouse IL-5 ELISA Assay is For Research Use Only

Size: 1×96 wells
Sensitivity: 7 pg/mL
Dynamic Range: 15.625 – 500 pg/ml
Incubation Time: 3.5 hours
Sample Type: Serum, Plasma, Cell Culture
Sample Size: 100 µl
Alternative Names: Interleukin 5, T-cell replacing factor, B-cell growth factor II, Eosinophil differentiation factor, Eosinophil colony stimulating factor


Assay Background

Mouse interleukin 5 (IL-5), also known as T-cell replacing factor, B-cell growth factor II, eosinophil differentiation factor and eosinophil colony stimulating factor, is a pleiotropic cytokine produced primarily by T cells (1-4). It supports the proliferation and differentiation of mouse, but not human, B cells, and enhances IgM, IgG1, IgA, and IgE secretion. IL-5 chemoattracts and enhances the survival and the effector functions of mature eosinophils and synergizes with various colony stimulating factors to increase eosinophil progenitor production and eosinophil expansion. Because of its diverse effects on eosinophils, IL-5 is strongly implicated in the pathogenesis of asthma and other hypereosinophilic inflammatory conditions (4-6). Mouse IL-5 cDNA encodes a 133 amino acid (aa) residue precursor protein containing a hydrophobic signal peptide that is cleaved to yield a 113 aa residue mature protein. Native mouse IL-5 is a disulfide-linked homodimeric 40-45 kDa glycoprotein with N- and O-linked carbohydrate chains. Mature human IL-5 is approximately 70% identical at the amino acid level to mouse IL-5. Whereas mouse and human IL-5 are equally active on human cell lines, human IL-5 is much less active than mouse IL-5 in mouse cell assays (1-3). IL-5 is a main regulator of eosinopoiesis, eosinophil maturation and activation. The elevated production of this cytokine is reported to be related to asthma or hypereosinophilic syndromes.


Related Products

Mouse IL-6 ELISA Assay
Human IL-5 ELISA Assay

Additional Information

Assay Principle


The Eagle Biosciences Mouse Interleukin 5 (IL-5) ELISA Assay Kit employs the quantitative sandwich enzyme immunoassay technique. A monoclonal antibody specific for IL-5 has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and any IL-5 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-5 is added to the wells and binds to the combination of capture antibody-IL-5 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 colored product is formed in proportion to the amount of IL-5 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-5 standard dilutions and IL-5 sample concentration determined.

Assay Procedure


  1. Prepare all reagents and working standards as directed in the previous sections.
  2. Determine the number of microwell strips required to test the desired number of samples plus appropriate number of wells needed for running blanks and standards. Remove extra microwell strips from holder and store in foil bag with the desiccant provided at 2-8°C sealed tightly.
  3. Add 100µl of Standard, control, or sample, per well. Cover with the adhesive strip provided. Incubate for 1.5 hours at 37C.
  4. Aspirate each well and wash, repeating the process three times for a total of four washes. Wash by filling each well with Wash Buffer (350 µl) using a squirt bottle, manifold dispenser or auto-washer. Complete removal of liquid at each step is essential to good performance. After the last wash, remove any remaining Wash Buffer by aspirating or decanting. Invert the plate and blot it against clean paper towels.
  5. Add 100 µl of the working solution of Biotin-Conjugate to each well. Cover with a new adhesive strip and incubate 1 hour at 37C.
  6. Repeat the aspiration/wash.
  7. Add 100 µl of the working solution of Streptavidin-HRP to each well. Cover with a new adhesive strip and incubate for 30 minutes at 37C.   Avoid placing the plate in direct light.
  8. Repeat the aspiration/wash.
  9. Add 100 µl of Substrate Solution to each well. Incubate for 10-20 minutes at 37C. Avoid placing the plate in direct light.
  10. Add 100 µl of Stop Solution to each well. Gently tap the plate to ensure thorough mixing.
  11. Determine the optical density of each well immediately, using a microplate reader set to 450 nm.(optionally 630nm as the reference wave length;610-650nm is acceptable)

Typical Standard Curve


Mouse IL5 ELISA Standard curve

Documents

Product Manual


 

Please note: All documents above are for reference use only and should not be used in place of the documents included with this physical product. If digital copies are needed, please contact us.

Publications

References


  • Takatsu, K. and A. Tominaga (1991) Progress in Growth Factor Research 3:87.
  • Takatsu, K. (1998) Cytokine and Growth Factor Reviews 9:25.
  • Takatsu, K. et al. (1994) Adv. Immunol. 54:134.
  • Sanderson, C.J. (1992) Blood 79:3101.
  • Lee, N.A. and J.J. Lee (1997) Amer. J. Respir. Cell Mol. Biol. 16:497.
  • Yagi, H. et al. (1997) Br. J. Dermatol. 136:918

Product Citations