(P-IG-53) Validation of the Anti-D Assay for Pooled Plasma Using the ORTHO VISION® Swift and Swift MAX Testing Platform

Background/Case Studies: The Anti-D assay is critical for detecting and quantifying Anti-D antibodies in pooled plasma, ensuring the safety and quality of plasma-derived products. The ORTHO VISION® Swift testing platforms, utilizing ID-MTS Gel Cards, automates immunohematology testing for Anti-D. This study aimed to validate the assay’s specificity, analytical sensitivity, robustness, and resistance to interference from Anti-A and Anti-B antibodies, in accordance with ICH Q2(R2) guidelines.

Study

Design/Methods: Specificity was evaluated by testing three pooled plasma lots (2230501, 2230502, 2230503) and 30 pooled plasma samples for Anti-D antibodies, expecting negative results. Analytical sensitivity was assessed using production pool samples (lot 2230501) spiked with WHO Anti-D standard (NIBSC code: 01/572) at concentrations of 142.5, 71.25, 35.625, 28.5, 17.8125, 8.90625, and 0 mIU/mL, with five replicates per concentration tested over three days. Probit analysis determined the limit of detection (LOD). Robustness was tested by preparing two aliquots of 20 replicates at 3x LOD (26.71875 mIU/mL) and analyzing them on two instruments (S/N: 70003052, 50002420). Interference was evaluated by spiking pooled plasma samples at 3x LOD with Anti-A and Anti-B at no spike, 1:16, and 1:32 dilutions, testing three replicates per condition. Geometric titer means, titer scores, and precision (%CV ≤20% within-run, ≤30% intermediate) were calculated.

Results/Findings: The assay demonstrated excellent specificity, with all 33 samples (three pooled plasma lots and 30 pooled plasma samples) testing negative for Anti-D antibodies, consistent with expected results. Analytical sensitivity analysis established an LOD of 8.90625 mIU/mL, with 15/15 valid positive results at all spiked concentrations and 0/15 at 0 mIU/mL. Robustness was confirmed, as all 40 replicates across two instruments tested positive at 26.71875 mIU/mL, with geometric titer means of 1 and titer score %CV of 0% (Aliquot 1), 11.47% (Aliquot 2), and 8.01% (combined). Interference studies showed consistent Anti-D detection at 3x LOD across all Anti-A and Anti-B antibody concentrations (up to 1:16 and 1:32 dilutions), with no interference observed. No exceptions or deviations occurred during the study.

Conclusions: The Anti-D assay on the ORTHO VISION® Swift platforms was successfully validated for pooled plasma testing, exhibiting high specificity, analytical sensitivity, robustness, and resistance to interference from Anti-A and Anti-B antibodies. The assay is suitable for quantifying Anti-D antibodies in production pools, supporting quality control in plasma fractionation processes.