Development and validation of panoptic Meso scale discovery assay to quantify total systemic interleukin-6

Area of Science:

• Biochemistry
• Immunology
• Assay Development

Background:

• Interleukin-6 (IL-6) exists in various molecular weight forms, including non-complexed and high molecular weight complexes.
• Accurate measurement of total IL-6 is crucial for understanding patient responses to IL-6-targeted therapies.
• Current assays only detect the low molecular weight, non-complexed form of IL-6, limiting comprehensive assessment.

Purpose of the Study:

• To develop and validate a high-throughput assay for measuring total IL-6 (both complexed and non-complexed forms) in serum or plasma.
• To establish a standardized assay format suitable for clinical testing and large-scale applications.
• To overcome the limitations of existing assays that only measure a single form of IL-6.

Main Methods:

• Screening of commercial capture and detection antibodies against humanized IL-6 in an enzyme-linked immunosorbent assay (ELISA) format.
• Optimization of antibody pairs to maximize IL-6 recovery and minimize background in a 100% serum matrix.
• Transfer of the developed plate-based total IL-6 assay to the Meso Scale Discovery (MSD) platform for high-throughput clinical testing.

Main Results:

• A highly effective antibody pair was identified and validated on the MSD platform, demonstrating a lower limit of quantification of 9.77 pg/L in human serum.
• The assay exhibited excellent recovery (93.13-113.27%) and robust precision, with pooled coefficients of variation of 20.12% (inter-assay) and 8.67% (intra-assay).
• The novel assay successfully detected high molecular weight IL-6 forms in patient samples, which were undetectable by a commercial kit.

Conclusions:

• A novel panoptic IL-6 assay capable of measuring both high and low molecular weight forms has been developed and validated.
• This assay demonstrates significant potential for clinical utility in assessing total IL-6 levels.
• The MSD platform facilitates large-scale clinical testing, enabling broader application of this comprehensive IL-6 measurement tool.