International Proficiency Test Targeting a Large Panel of Botulinum Neurotoxin Sero- and Subtypes in Different Matrices

Detection of botulinum neurotoxins (BoNTs) involves a combination of technical challenges that call for the execution of inter-laboratory proficiency tests (PTs) to define the performance and ease of implementation of existing diagnostic methods regarding representative BoNT toxin-types spiked in clinical, food, or environmental matrices. In the framework of the EU project EuroBioTox, we organized an international proficiency test for the detection and quantification of the clinically relevant BoNT/A, B, E, and F sero- and subtypes including concentrations as low as 0.5 ng/mL. BoNTs were spiked in serum, milk, and soil matrices. Here, we evaluate the results of 18 laboratories participating in this PT. Participants have implemented a wide array of detection methods based on functional, immunological, and mass spectrometric principles. Methods implemented in this proficiency test notably included endopeptidase assays either coupled to mass spectrometry (Endopep-MS) or enzyme-linked immunosorbent assays (Endopep-ELISA). This interlaboratory exercise pinpoints the most effective and complementary methods shared by the greatest number of participants, also highlighting the importance of combining the training of selected methods and of distributing toxin reference material to reduce the variability of quantitative data.