Human IL-32 DuoSet ELISA Summary

Product Summary

This DuoSet ELISA Development kit contains the basic components required for the development of sandwich ELISAs to measure natural and recombinant human IL-32. The suggested diluent is suitable for the analysis of most cell culture supernate samples. Diluents for complex matrices, such as serum and plasma, should be evaluated prior to use in this DuoSet ELISA.

Preparation and Storage

Background: IL-32

IL-32 alpha is the shortest and most abundant of four potential splice variants of the proinflammatory cytokine IL-32 (previously called NK4) with a predicted unmodified size of 15 kDa. Potential modifications include myristoylation and N-glycosylation. Transfected IL-32 alpha was more likely to be cell-associated as compared to IL-32 beta, suggesting an intracellular function. IL-32 is induced by mitogens in peripheral lymphocytes, by IFN-gamma in epithelial cells, or by IL-12 with IL-18 in NK cells and in turn induces cytokine expression. No ortholog has been found in mouse.

Assay Procedure

Refer to the product datasheet for the complete assay procedure.

Bring all reagents and samples to room temperature before use. It is recommended that all samples, standards, and controls be assayed in duplicate.

1.     Prepare all reagents, standard dilutions, and samples as directed in the product insert.

2.     Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal.

3.     Add 50 μL of Assay Diluent to each well.

4.     Add 50 μL of Standard, control, or sample to each well. Cover with a plate sealer, and incubate at room temperature for 2 hours.

5.     Aspirate each well and wash, repeating the process 4 times for a total of 5 washes.

6.     Add 100 μL of Conjugate to each well. Cover with a new plate sealer, and incubate at room temperature for 2 hours.

7.     Aspirate and wash 5 times.

8.     Add 100 μL Substrate Solution to each well.

9.     Add 100 μL of Stop Solution to each well. Read at 450 nm within 30 minutes. Set wavelength correction to 540 nm or 570 nm.