Reporting the limits of detection and quantification for environmental DNA assays




Klymus, Katy E.
Merkes, Christopher M.
Allison, Michael J.
Goldberg, Caren S.
Helbing, Caren C.
Hunter, Margaret E.
Jackson, Craig A.
Lance, Richard F.
Mangan, Anna M.
Monroe, Emy M.

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Environmental DNA


Background Environmental DNA (eDNA) analysis is increasingly being used to detect the presence and relative abundance of rare species, especially invasive or imperiled aquatic species. The rapid progress in the eDNA field has resulted in numerous studies impacting conservation and management actions. However, standardization of eDNA methods and reporting across the field is yet to be fully established, with one area being the calculation and interpretation of assay limit of detection (LOD) and limit of quantification (LOQ). Aims Here, we propose establishing consistent methods for determining and reporting of LOD and LOQ for single‐species quantitative PCR (qPCR) eDNA studies. Materials & Methods/ Results We utilize datasets from multiple cooperating laboratories to demonstrate both a discrete threshold approach and a curve‐fitting modeling approach for determining LODs and LOQs for eDNA qPCR assays. We also provide details of an R script developed and applied for the modeling method. Discussion/Conclusions Ultimately, standardization of how LOD and LOQ are determined, interpreted, and reported for eDNA assays will allow for more informed interpretation of assay results, more meaningful interlaboratory comparisons of experiments, and enhanced capacity for assessing the relative technical quality and performance of different eDNA qPCR assays.



assay optimization, eDNA, qPCR, standardization


Klymus, K. E., Merkes, C. M., Allison, M. J., Goldberg, C. S., Helbing, C. C., Hunter, M. E., Jackson, C. A., Lance, R. F., Mangan, A. M., Monroe, E. M., Piaggio, A. J., Stokdyk, J. P., Wilson, C. C., & Richter, C. A. (2019). Reporting the limits of detection and quantification for environmental DNA assays. Environmental DNA, 2(3), 271-282.