Impact of Unconjugated estriol (uE3) assay interference on prenatal screening tests.

Background: Unconjugated estriol (uE3) is an important biomarker in second trimester prenatal screening. Previous studies from our laboratory identified rare interference in the Beckman uE3 assay due to anti-ALP antibodies, which could be mitigated with a scavenger or heat-inactivated ALP (hALP). In the current study, 160 de-identified patient samples previously submitted for the Quad screen with low uE3 multiples of the median (MoM ≤0.50) were investigated for potential interference.

Methods: A reagent pack spiking strategy with hALP was employed to understand if the interference could be identified and mitigated in a scalable manner. The 160 samples were measured using uE3 lot #920861 previously known to be subject to interference, lot #920861 spiked with hALP, and the vendor reformulated lot #922579. Samples were suspected to have interference if the percent difference in uE3 measurements was >50%. Pseudo-risks were calculated using a test patient environment to understand the screening impact due to the change in uE3 result.

Results: Seventeen of the 160 samples had uE3 results that were >50% different between the hALP spiked and non-spiked reagent pack. Both original lot #920861 with hALP and reformulated lot #922579 identified the same 17 patients as having interference in lot #920861. Analysis of screening risks using a test patient environment showed that assay interference could result in false positives for one trisomy 21 and three trisomy 18 post-test risk calculations.

Conclusion: Our experiment of reagent pack spiking with hALP produced similar uE3 results to a reformulated reagent designed to address potential interference, demonstrating that this is a feasible strategy to screen for interference in a scalable manner. The vendor-provided reformulation addressed anti-ALP interference and improved the performance of the screen.