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Optimization, Comparison, and Application of Colorimetric vs. Chemiluminescence Based Indirect Sandwich ELISA for Measurement of Human IL‐23

Abstract Currently, there is neither a published ELISA method nor it is clear whether chemiluminescence substrates would provide better sensitivity vs. colorimetric substrates for measuring human IL‐23–a recently described Type‐1 immunity associated cytokine. Initially, we optimized a colorimetric ELISA using p‐nitro‐phenyl phosphate substrate. Subsequently, we compared it with chemiluminescence substrates that provided ∼5‐fold enhanced sensitivity (mean sensitivity; 26.3 pg/mL vs. colorimetric assay, 131 pg/mL; p<0.01). Both methods were reliable, with <10% inter‐ and intra‐assay variations. We then found that the chemiluminescence method was useful in situations where human IL‐23 was not readily measurable by a colorimetric method.