Optiview dab ihc detection kit

Interlaboratory reproducibility studies for the VENTANA PD-L1 (SP142) Assay were conducted to demonstrate the reproducibility of the assay in determining the PD-L1 status in UC and NSCLC tissue samples. In each study, 28 unique samples (equal number of cases above and below the cut-off) were stained at 3 external College of American Pathologists accredited laboratories on 5 nonconsecutive days over a period of at least 20 days. Staining was conducted on a single BenchMark ULTRA instrument at each site, using 1 lot of PD-L1 (SP142) antibody, OptiView DAB IHC Detection Kit, and OptiView Amplification Kit. Before staining, slides were blinded and randomized. At each site, the stained slides were independently evaluated by 2 pathologists. The sample set comprised a total of 420 case slides (140 slides per site, 3 sites) and 840 reads (420 case slides, 2 pathologists) generated from 28 unique samples.

IHC was performed on 4 µm FFPE sections. MCPyV (clone CM2B4; cat sc-136172, Santa Cruz Biotechnology, Santa Cruz, CA; dilution 1:100) and CD3 (clone LN10; cat NCL-L-CD3-565, Newcastle, UK; dilution 1:200) staining was performed on the Leica Bond III Autostainer. For programmed death-ligand 1 (PD-L1), IHC clone SP263 was used (Assay v1.00.0001, using the VENTANA OptiView DAB IHC Detection Kit on the BenchMark ULTRA autostainer). Semiquantitative scoring was performed by a single pathologist blinded to the clinical data. T cells were scored by counting the mean number of positive cells in tumor per high-power field (0.55 mm diameter). PD-L1 scoring was performed using a standardized approach that has been described previously.12 (link) Briefly, two scores were derived; the tumor proportion score (TPS), which represents the percentage of tumor cells showing positive membranous expression of PD-L1, and the immune cell score (IC), which represents the proportion of tumor area occupied by PD-L1–positive immune cells. TPS and IC assessment was based on the entire tumor area on a single representative slide. A minimum of 50 tumor cells were required for a tumor to be considered assessable for TPS and IC. Four and three patients did not have sufficient material for PD-L1 and CD3 IHC, respectively, and were excluded from this analysis.