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Analysis of Two ELISA Formats and Antigen Preparations Using Polyclonal Antibodies Against Phomopsis longicolla. L. M. Brill, Former graduate research assistant, Department of Plant Pathology, University of Illinois at Urbana-Champaign, 1102 South Goodwin Avenue, Urbana 61801-4709, Present address: Department of Biology, University of California, 405 Hilgard Avenue, Los Angeles 90024-1606; R. D. McClary(2), and J. B. Sinclair(3). (2)(3)research specialist in agriculture, and professor, respectively, Department of Plant Pathology, University of Illinois at Urbana-Champaign, 1102 South Goodwin Avenue, Urbana 61801-4709. Phytopathology 84:173-179. Accepted for publication 11 October 1993. Copyright 1994 The American Phytopathological Society. DOI: 10.1094/Phyto-84-173.

Polyclonal antibodies (PAbs) were produced in New Zealand white rabbits with culture filtrate and mycelial extract immunogen preparations from the soybean (Glycine max) fungal pathogen Phomopsis longicolla. The PAbs were purified to the immunoglobin fraction and tested in indirect enzyme-linked immunosorbent assay (ELISA) and in double antibody sandwich-ELISA (DAS-ELISA). The PAbs raised to culture filtrate were more specific but less active in binding to members of the Diaporthe-Phomopsis complex than were those raised to the mycelial extract immunogen preparation. DAS-ELISA was more specific and 100-fold more sensitive in detecting members of the complex than was indirect ELISA. Variability in specificity between different PAbs was lower in DAS-ELISA compared with indirect ELISA. Immunization of one rabbit with culture filtrate over an extended time resulted in maximum anti-P. longicolla activity after three immunizations, and the activity became constant against most members of the complex at the same time. Reactivity to some cultures of P. longicolla was undetectable following the fourth and fifth immunizations, whereas reactivity to all of our other cultures of the complex remained high.