Development of Monoclonal Antibodies Specific to Galectin of Pine Wood Nematode, Bursaphelenchus xylophilus (Steiner and Buhrer) Nickle and Their Utilization for Detection of Pine Wood Nematodes

Abstract

Currently, there is no available tool that rapidly diagnoses pine wood nematode (PWN)-infected pine trees in the field. In this study, we synthesized and purified PWN Galectin, which might be an antigen specific to PWN, using the Baculovirus expression system. We used PWN Galectin as an antigen for generating 1,464 fusion hybridoma cell lines secreting monoclonal antibodies (Mabs). Among them, we selected 62 fusion hybridoma cell lines showing high reactivity to PWN Galectin. We further selected 12 fusion hybridoma cell lines showing high reactivity to the standard PWN-infected pine tree phosphate buffered saline (PBS) extract. Additionally, two fusion hybridoma cell lines showing no or extremely low reactivity were used as controls. The selected fusion hybridoma cell lines were subjected to limiting dilutions for selecting and establishing Mab-secreting cell lines showing higher reactivity to the standard PWN-infected pine tree extract than to the standard normal pine tree PBS extract. Moreover, the selected fusion hybridoma cell lines were further selected based on their higher reactivity to PWN protein extracts than to three non-pathogenic nematode protein extracts. The Mab-secreting cell lines established in this study could be used to develop rapid diagnostic tools that can be used in the field or in laboratories for detecting PWN-infected pine trees or PWN.