title: Antibody array in a multiwell plate format for the sensitive and multiplexed detection of important plant pathogens.
abstract: The global seed market is considered to be an important industry with a total value of $10,543 million US dollars in 2012. Because plant pathogens such as bacteria and viruses cause a significant economic loss to both producers and exporters, the seed export industry urgently requires rapid, sensitive, and inexpensive testing for the pathogens to prevent disease spreading worldwide. This study developed an antibody array in a multiwell plate format to simultaneously detect four crucial plant pathogens, namely, a bacterial fruit blotch bacterium Acidovorax avenae subsp. citrulli (Aac), Chilli veinal mottle virus (ChiVMV, potyvirus), Watermelon silver mottle virus (WSMoV, tospovirus serogroup IV), and Melon yellow spot virus (MYSV, tospovirus). The capture antibodies specific to the pathogens were immobilized on each well at preassigned positions by an automatic microarrayer. The antibodies on the arrays specifically captured the corresponding pathogens present in the sample extracts. The presence of pathogens bound on the capture antibodies was subsequently detected by a cocktail of fluorescently conjugated secondary antibodies. The limits of detection of the developed antibody array for the detection of Aac, ChiVMV, WSMoV, and MYSV were 5 × 10(5) CFU/mL, 30 ng/mL, 1000 ng/mL, and 160 ng/mL, respectively, which were very similar to those of the conventional ELISA method. The antibody array in a multiwell plate format accurately detected plant pathogens in single and multiple detections. Moreover, this format enables easy handling of the assay at a higher speed of operation.
authors.affiliation: National Center for Genetic Engineering and Biotechnology (BIOTEC), National Science and Technology Development Agency (NSTDA) , 113 Thailand Science Park, Phahonyothin Road, Khlong Luang, Pathum Thani 12120, Thailand.
authors.forename: Ratthaphol
authors.initials: R
authors.lastname: Charlermroj
authors.forename: Orawan
authors.initials: O
authors.lastname: Himananto
authors.forename: Channarong
authors.initials: C
authors.lastname: Seepiban
authors.forename: Mallika
authors.initials: M
authors.lastname: Kumpoosiri
authors.forename: Nuchnard
authors.initials: N
authors.lastname: Warin
authors.forename: Oraprapai
authors.initials: O
authors.lastname: Gajanandana
authors.forename: Christopher T
authors.initials: CT
authors.lastname: Elliott
authors.forename: Nitsara
authors.initials: N
authors.lastname: Karoonuthaisiri
mesh_terms: D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000276:immunology; D002021:Buffers; D036022:Capsid Proteins / Q000235:genetics / Q000276:immunology; D042621:Comamonadaceae / Q000472:pathogenicity; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D004867:Equipment Design; D005456:Fluorescent Dyes / Q000737:chemistry; D057230:Limit of Detection; D046228:Microarray Analysis / Q000295:instrumentation / Q000379:methods*; D010935:Plant Diseases / Q000382:microbiology* / Q000821:virology*; D016133:Polymerase Chain Reaction; D017800:Potyvirus / Q000472:pathogenicity; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D017865:Tospovirus / Q000472:pathogenicity
chemical_list: D000911:Antibodies, Monoclonal; D002021:Buffers; D036022:Capsid Proteins; D005456:Fluorescent Dyes