Purpose: Biomarker analysis has played an increasingly important role in drug development both as markers of activity and characterization of disease. Biomarker levels can vary depending on matrix and disease state. An added complication is that the quantitation of biomarkers is highly dependent on the analytical platform and the reagents used to generate the data. Data generated from one assay may not necessarily match or correlate with data from another assay. Of particular interest in biomarkers is the potential difference in quantitation between measurements in serum versus plasma . Here we present the results of a platform comparison for the analysis of CCL18 levels in serum and plasma from three different platforms: ELISA, IMPACT, and Ella [2-4].
Methods: Fifty-eight matched sets of human serum and plasma were measured for CCL18 using the Quantikine® ELISA kit for human CCL18/PARC (R&D Systems) and the IMPACT (Immunological Multi-Parameter Chip Technology), a fully automated protein-array chip system (Roche Diagnostics GmbH). Additionally, the same 58 serum samples were analyzed using the Ella™ assay for CCL18 (ProteinSimple).
Results: Comparison of serum versus plasma using either the Quantikine ELISA or the IMPACT showed good agreement indicating no bias due to matrix. The average %difference between serum and plasma for the Quantikine ELISA was -5.3% with r=0.972, P=0.473; for the IMPACT, it was -10.3% with r=0.968, P=0.393. Comparing the data from the Quantikine ELISA to the IMPACT showed a very good correlation, with r=0.939, P< 0.0001 for serum and r=0.919, P< 0.0001 for plasma, however the quantitation using the IMPACT resulted in concentration values that averaged 3.95 times higher than for the ELISA. Subsequently, the same serum sample set was analyzed using Ella cartridges from ProteinSimple. Data was highly correlative with the ELISA (r=0.950, P=0.317) and the IMPACT (r=0.942, P< 0.0001), although similar to previous results, the IMPACT concentrations averaged 3.78 times higher whereas the average %difference between the ELISA and Ella was 11%.
Conclusion: There was good agreement between serum and plasma concentrations for matched samples, indicating that either matrix is suitable for measurement of CCL18 levels. The Quantikine ELISA and the ProteinSimple Ella cartridges, which use the same antibody pair and standard, showed good agreement in concentration. The IMPACT assay, which uses both a different antibody pair and standard, results in concentration values nearly four times higher although the data is correlative. These kinds of assay differences need to be understood and taken into account when comparing data from different sources. For biomarkers, where frequently the assessment is relative change from baseline rather than absolute value, differences in quantitation are acceptable as long as the data is correlative. If it is, then final selection of a platform may be guided by other factors such as cost, ease of use, and availability.
1) O’Neal W, et al. Comparison of serum, EDTA plasma, and P100 plasma for luminex-based biomarker multiplex assays in patients with chronic obstructive pulmonary disease in the SPIROMICS study. J. Transl Med. 2014; 12:1-9.
2) Yeung D, Ciotti S, Purushothama S, Gharakhani E, Kuesters G, Schlain B, Shen C, Donaldson D, Mikulskis A. Evaluation of highly sensitive immunoassay technologies for quantitative measurements of sub-pg/mL levels of cytokines in human serum. J. Immunol. Methods 2016; 437:53-63.
3) Claudon A, Vergnaud P, Valverde C, Mayr A, Klause U, Garnero P. New Automated Multiplex Assay for Bone Turnover Markers in Osteoporosis. Clin. Chem. 2008; 54:1554-1563.
4) Dysinger M, Marusov G, Fraser S. Quantitative analysis of four protein biomarkers: An automated microfluidic cartridge-based method and its comparison to colorimetric ELISA. J. Immunol. Methods 2017; 451:1-10.