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One obstacle in diagnostic pathology is the harmonization of one drug-one diagnostic tests for programmed death ligand-1 (PD-L1). There are many challenges in accurate comparisons of diagnostic tests, such as differences in the titer of each antibody, detection system and dynamic range of visualization. Our previously developed digital immunostaining technique is highly sensitive and quantitative with the ability to quantify particles that bind in a one-to-one fashion with antibody in each cell. Determining the differences in the titer of each antibody with digital immunostaining may be beneficial for future harmonized analysis. To demonstrate the accuracy of digital immunostaining, the present study compared the number of particles with ELISA and nCounter data from five cell lines. NCI-H460 exhib-ited the highest level of PD-L1 protein, followed by A549, PC-3, NCI-H1299, and NCI-H446 cells. In addition, the PD-L1 mRNA values determined by nCounter corresponded with the order of the protein levels determined by ELISA. The present study revealed that digital immunostaining for PD-L1 was highly associated with ELISA and nCounter data. Among the four antibodies tested, the titer of all but SP142 coincided with ELISA and nCounter data. These results indicated that our digital immunostaining technique may be beneficial for future harmonized analysis.

Quantitative immunohistochemical assay with novel digital immunostaining for comparisons of PD‑L1 antibodies