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bioRxiv · 10.1101/2022.02.21.481075

Consistent measurement of LAG-3 expression across multiple staining platforms with the 17B4 antibody clone

Wojcik, J. B.·Desai, K.·Avraam, K.·Vandebroek, A.·Dillon, L. M.·Giacomazzi, G.·Rypens, C.·Benci, J.
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Abstract

ContextAn immunohistochemistry (IHC) assay developed to detect lymphocyte-activation gene 3 (LAG-3), a novel immune checkpoint inhibitor target, has demonstrated high analytical precision and interlaboratory reproducibility using a Leica staining platform, but has not been investigated on other IHC staining platforms. ObjectiveTo evaluate the performance of LAG-3 IHC assays using the 17B4 antibody clone across widely used IHC staining platforms: Agilent/Dako Autostainer Link 48 (ASL-48) and VENTANA BenchMark ULTRA (VBU) compared with Leica BOND-RX (BOND-RX). DesignEighty formalin-fixed paraffin-embedded melanoma tissue blocks were cut into consecutive sections and evaluated using staining platform-specific IHC assays with the 17B4 antibody clone. Duplicate testing was performed on the BOND-RX platform to assess intraplatform agreement. LAG-3 expression using a numerical score was evaluated by a pathologist and with a digital scoring algorithm. LAG-3 positivity was determined from manual scores using a [≥] 1% cutoff. ResultsLAG-3 IHC staining patterns and intensities were visually similar across all 3 staining platforms. Pearson correlation was [≥] 0.88 for interplatform and BOND-RX intraplatform concordance when LAG-3 expression was evaluated with a numerical score determined by a pathologist. Correlation increased with a numerical score determined with a digital scoring algorithm (Pearson correlation [≥] 0.93 for all comparisons). Overall percentage agreement was [≥] 77.5% for interplatform and BOND-RX intraplatform comparisons when a [≥] 1% cutoff was used to determine LAG-3 positivity. ConclusionsData from this study demonstrate that LAG-3 expression can be robustly and reproducibly assessed across 3 major commercial IHC staining platforms using the 17B4 antibody clone.

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Wojcik, J. B., Desai, K., Avraam, K., Vandebroek, A., Dillon, L. M., Giacomazzi, G., Rypens, C., Benci, J.. 2022-02-22. Consistent measurement of LAG-3 expression across multiple staining platforms with the 17B4 antibody clone. https://doi.org/10.1101/2022.02.21.481075

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