A study published in PLoS ONE revealed the discovery of a unique multiplex detection method able to search for four common plant pathogens simultaneously.
Plant pathogens are a worldwide concern, with 57 countries exporting 106,000 metric tons in 2010. This means that a diseased crop has not only major health concerns but also international economic effects.
Current methods for plant pathogen detection, such as ELISA (enzyme-linked immunosorbent assay) and PCR (polymerase chain reaction) are effective but slow and require specialized skills. The industry needs an inexpensive, rapid, and high-throughput method for plant disease screening and control, to protect crops and consumers.
The present study has developed such a method to detect four pervasive plant pathogens – chili vein-banding mottle virus (CVbMV), melon yellow spot virus (MYSV), watermelon silver mottle virus (WSMoV), and Acidovorax avenae ssp. citruli (Aac), which affects certain cucurbits (the melon family). The researchers have created a multiplex detection method using microsphere immunoassays which cuts assay time down to only one hour.
Eight antibodies for these pathogens were chosen to be used in the assay, provided by the National Center for Genetic Engineering and Biotechnology and Kasetsart University, both located in Thailand:
- Mouse MAb 11E5, for Aac
- Rabbit PAb MPC, for Aac
- Mouse MAb 1B4, for CVbMV
- Mouse MAb 1G8, for CVbMV
- Mouse MAb 2D6, for WSMoV
- Rabbit PAb A3, for WSMoV
- Rabbit PAb MYSV6, for WSMoV and MYSV
- Mouse MAb 5E7, for MYSV
To optimize the assay for accuracy, all possible combinations of antibodies and conjugations were coupled to microsphere sets and tested. They found that the following combinations illustrated in the figure below, showing the assay procedure, provided the highest signal strengths for detection.
The figure is reproduced from Charlermroj R, Himananto O, Seepiban C, Kumpoosiri M, Warin N, et al. (2013) Multiplex Detection of Plant Pathogens Using a Microsphere Immunoassay Technology. PLoS ONE 8(4): e62344. doi:10.1371/journal.pone.0062344 and under the Creative Commons Attribution License.
The research assay was developed using xMAP® multiplexing technologies designed by Luminex Corporation. A specific MagPlex® microsphere set was obtained to be coupled with the antibodies to capture each pathogen.
The development of this new multiplex immunoassay will allow researchers to conduct accurate assays at a much faster rate. The work sets precedence for this method of simultaneous detection of multiple plant pathogens, and shows Luminex to be the leader in flexible and open multiplexing technologies.