Development of a Planar Waveguide Microarray for the Monitoring and Early Detection of Five Harmful Algal Toxins in Water and Cultures Sara E. McNamee, † Christopher T. Elliott, † Brett Greer, † Michael Lochhead, ‡ and Katrina Campbell* ,† † Institute for Global Food Security, School of Biological Sciences, Queen’s University, Stranmillis Road, Belfast BT9 5AG, United Kingdom ‡ MBio Diagnostics Inc., 5603 Arapahoe Avenue, Boulder, Colorado 80303, United States ABSTRACT: A novel multiplex microarray has been developed for the detection of five groups of harmful algal and cyanobacterial toxins found in marine, brackish, and freshwater environments including domoic acid (DA), okadaic acid (OA, and analogues), saxitoxin (STX, and analogues), cylindrospermopsin (CYN) and microcystins (MC, and analogues). The sensitivity and specificity were determined and feasibility to be used as a screening tool investigated. Results for algal/cyanobacterial cultures (n = 12) and seawater samples (n = 33) were compared to conventional analytical methods, such as high performance liquid chromatography (HPLC) and liquid chromatography tandem mass spectrom- etry (LC-MS/MS). Detection limits for the 15 min assay were 0.37, 0.44, 0.05, 0.08, and 0.40 ng/mL for DA, OA, STX, CYN, and MC, respectively. The correlation of data obtained from the microarray compared to conventional analysis for the 12 cultures was r 2 = 0.83. Analysis of seawater samples showed that 82, 82, 70, 82, and 12% of samples were positive (>IC 20 ) compared to 67, 55, 36, 0, and 0% for DA, OA, STX, CYN, and MC, respectively, for conventional analytical methods. The discrepancies in results can be attributed to the enhanced sensitivity and cross-reactivity profiles of the antibodies in the MBio microarray. The feasibility of the microarray as a rapid, easy to use, and highly sensitive screening tool has been illustrated for the five-plex detection of biotoxins. The research demonstrates an early warning screening assay to support national monitoring agencies by providing a faster and more accurate means of identifying and quantifying harmful toxins in water samples. 1. INTRODUCTION During recent decades, there has been an increase in the occurrences of harmful algal blooms (HAB) in many marine and freshwater systems. 1 HABs are caused by the production of biotoxins belonging to three taxa groups; diatoms, dino- flagellates, and cyanobacteria. There is also an increasing number of toxic algal species and algal toxins identified with the economic losses from HABs greater than before. 2 Several algal toxins are classified corresponding to their acute symptoms in humans and include amnesic shellfish poisoning (ASP) toxins, diarrheic shellfish poisoning (DSP) toxins, and paralytic shellfish poisoning (PSP) toxins. Cyanotoxins such as micro- cystins (MC) and cylindrospermopsin (CYN) initially found in freshwater environments are presenting in brackish and marine environments 3 and have been observed to be spreading from tropical to temperate environments. 4 Current methods of detection for marine biotoxins are primarily aimed for shellfish where regulations are in place within the EU (Regulation (EC) No. 853/2004) and worldwide (Codex, STAN 292-2008). No regulations are currently in place for the detection of marine biotoxins in aquatic samples. MCs are the only toxin with a guideline value of 1 μg/L in drinking water recommended by the World Health Organization (WHO). Therefore, many of the current methods do not have the required sensitivity to act as an early warning screening method for water quality monitoring. For marine biotoxins analytical methods such as HPLC and LC-MS/MS were originally developed for their detection in shellfish as an alternative to the mouse bioassay but have been adapted for detection in algal and seawater samples for PSP, OA and DA toxins. 4-12 Analytical tools such as HPLC with UV 13 or MS 14,15 detection have been developed for MC and LC-MS/MS for CYN detection. 4,16,17 Although analytical methods are available, it is acknowledged that these methods require skilled personnel, are labor intensive, can be hindered by the lack of available analytical standards and tend to be specific to a single toxin group. Immunological assays such as ELISA 18-23 and lateral flow devices 24-28 have shown promise as sensitive rapid commercially available screening tools (e.g., Abraxis, Jellet Rapid Testing Ltd., Neogen). The emergence of biosensor based immunological assays have shown many Received: August 25, 2014 Revised: October 15, 2014 Accepted: October 16, 2014 Published: October 31, 2014 Article pubs.acs.org/est © 2014 American Chemical Society 13340 dx.doi.org/10.1021/es504172j | Environ. Sci. Technol. 2014, 48, 13340-13349