Fiber-optic microarray for simultaneous detection of multiple harmful algal bloom species.
Harmful algal blooms (HABs) are a serious threat to coastal resources, causing a variety of impacts on public health, regional economies, and ecosystems. Plankton analysis is a valuable component of many HAB monitoring and research programs, but the diversity of plankton poses a problem in discriminating toxic from nontoxic species using conventional detection methods. Here we describe a sensitive and specific sandwich hybridization assay that combines fiber-optic microarrays with oligonucleotide probes to detect and enumerate the HAB species Alexandrium fundyense, Alexandrium ostenfeldii, and Pseudo-nitzschia australis. Microarrays were prepared by loading oligonucleotide probe-coupled microspheres (diameter, 3 mum) onto the distal ends of chemically etched imaging fiber bundles. Hybridization of target rRNA from HAB cells to immobilized probes on the microspheres was visualized using Cy3-labeled secondary probes in a sandwich-type assay format. We applied these microarrays to the detection and enumeration of HAB cells in both cultured and field samples. Our study demonstrated a detection limit of approximately 5 cells for all three target organisms within 45 min, without a separate amplification step, in both sample types. We also developed a multiplexed microarray to detect the three HAB species simultaneously, which successfully detected the target organisms, alone and in combination, without cross-reactivity. Our study suggests that fiber-optic microarrays can be used for rapid and sensitive detection and potential enumeration of HAB species in the environment.
Ahn
Soohyoun
S
Department of Chemistry, Tufts University, 62 Talbot Ave., Medford, MA 02155, USA.
Kulis
David M
DM
Erdner
Deana L
DL
Anderson
Donald M
DM
Walt
David R
DR
D001483:Base Sequence; D015342:DNA Probes / Q000235:genetics; D056890:Eukaryota / Q000235:genetics* / Q000302:isolation & purification*; D005068:Eutrophication; D005336:Fiber Optic Technology; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods* / Q000706:statistics & numerical data; D055100:Optical Fibers; D010839:Phytoplankton / Q000235:genetics* / Q000302:isolation & purification*; D012623:Seawater / Q000382:microbiology; D012680:Sensitivity and Specificity
D015342:DNA Probes