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Identification of Aldose Reductase 1 as a pine wood nematode secretory enzyme and generation and characterization of its monoclonal antibodies

Kim, A-Young, Osabutey, Angelina F., Yoon, Kyungjae Andrew, Choi, Bo-Hye, Lee, Si-Hyeock, Han, Hyerim, Koh, Young Ho
Journal of Asia-Pacific entomology 2019 v.22 no.1 pp. 233-238
Baculoviridae, Bursaphelenchus xylophilus, Pinus, aldehyde reductase, antigens, clones, enzyme-linked immunosorbent assay, genes, hybridomas, monoclonal antibodies, proteins, proteome, transcriptome
In order to identify the specific antigens for pine wood nematode (PWN, Bursaphelenchus xylophilus [Steiner and Buhrer] Nickle), we confirmed that one of the genes commonly found in the transcriptome, proteome and secretory proteins of PWN belonged to the Aldose Reductase (AR) family protein. 36.5 kDa PWN-AR1 was expressed and purified using Baculovirus Expression System. Total 1546 hybridoma fusion library was generated and screened for specificity to PWN-AR1 by Enzyme Linked Immunosorbent assay (ELISA). Nine clones showed strong immunoreactivity to PWN-AR1 were limited-diluted. Total 864 limited-diluted clones were further screened using PWN-AR1 by ELISA and 34 monoclonal antibody (Mab) clones were selected. 34 Mab clones were further screened using PWN extracts and a standard PWN-infected pine tree extract by ELISA. Finally nine clones were selected and their immunoreactivities to 4 different nematodes were examined by ELISA. Seven clones specifically recognized PWN while two clones recognized four nematodes. Our data suggested that PWN-AR1 is a PWN secretory enzyme while PWN is invading pine trees, Thus, PWN-AR1-Mabs could be used to develop diagnosis tools for PWN and its infected pine trees.