**Supplementary material**

#### **Supplemental Figure 1.**

*The assay workflow and experimental plate layout for kinase inhibition screening. Supplemental Figure A shows the assay workflow beginning with plating cells, incubating cells with kinase inhibitor, cell activation, assay steps and high-throughput data acquisition. Supplemental Figure B shows the plate map layout showing negative controls in red, sample wells in blue and positive controls in green. The plate map for plate 2 follows the layout of plate one.*

*Developing a Novel Multiplexed Immune Assay Platform to Screen Kinase Modulators… DOI: http://dx.doi.org/10.5772/intechopen.97304*


#### **Supplemental Figure 2.**

*Four MEK1/2 inhibitors that inhibit relevant markers. The screening results showed that all 4 compounds broadly inhibit the major relevant markers (CD69 and CD25 expression, and IFNγ and TNFα secretion) after 24-hour T cell activation. Supplement Figure 2A represents the inhibitory effect of MEK1/2 inhibitors on the expression of CD69 and CD25 on both T helper and T cytotoxic cells. Supplement Figure 2B illustrates the decreasing effects on the secretion of the cytokines IFNγ and TNFα. In the subsequent dosage test to confirm the screening results (Supplement Figure 2C), 24 IC50s were generated from 4 compounds and 6 endpoints.*
