AlphaLISA versus ELISA-based detection of interleukin 18 in healthy subjects and patients with end-stage renal disease

Abstract

Background. Interleukin 18 (IL-18) is an important pro-inflammatory cytokine investigated in end-stage renal disease (ESRD) and several autoimmune and inflammatory diseases. The quantitative colorimetric sandwich ELISA test kit from MBL is the most frequently used ELISA kit for IL-18 detection. Recently, a new bead-based proximity assay named AlphaLISA was developed. The aim of this study was to compare the performance of these two kits and to evaluate aspects of sample handling on IL-18 measurements. Methods. Measurements of IL-18 were performed on plasma samples from 11 ESRD-patients in regular haemodialysis treatment and 10 healthy volunteers. Results. We found a significant difference between assays corresponding to a factor of 6.4 (6.0; 6.7), p < 0.0001. Agreement was excellent with intra-class correlation coefficient 0.95. MBL had lower inter-assay variation, batch-to-batch variation and day-to-day variation. Spiking with recombinant IL-18 resulted in a mean recovery of 91 ± 7 (MBL) vs. 135 ± 17% (AlphaLISA), p < 0.0001. Both kits performed equally well when linearity was investigated. IL-18 was stable over a period of 424 days when plasma samples were stored at − 80°C. When blood samples were stored at room temperature mean IL-18 concentration changed significantly between 0, 1, 6, and 24 hours after sampling (p = 0.0213). No significant change was found with storage at 5°C. Severe haemolysis affected IL-18 measurements, whereas repeated freezing and thawing showed no effect. Mean IL-18 concentration was significantly higher in ESRD-patients compared to healthy subjects regardless of assay. Conclusion. Assay performance was best with the MBL-kit, although AlphaLISA was less time consuming when measuring plasma IL-18.

Key Words::

• Alphascreen
• bead-based proximity
• chronic
• enzyme-linked immunosorbent assay
• high throughput
• IL-18
• kidney failure
• luminescence
• sandwich
• specimen handling

Acknowledgements

We would like to thank our lab-technician Ilse Rasmussen who performed all testing of the two kits in our lab. We would also like to thank Perkin Elmer for assistance and guidance in the use of AlphaLISA. Finally, we would like to acknowledge the Institute of Medical Biochemistry, Aarhus University for allowing us to use their lab facilities and the EnSpire™ plate reader. This study was supported with funding from Helen og Ejnar Bjørnows fond.

Declaration of interest: The authors report no conflicts of interest. The authors alone are responsible for the content and writing of the paper.