SYMPOSIA PAPER Published: 01 January 1995

Enzyme-Linked Immunosorbent Assay for Quantification of Var. Crystalline Protein in Some Commercial Formulations


The widespread use of microbial pesticides for forest insect control necessitates determination of the delta-endotoxin protein levels in commercial formulations to optimize their use-patterns in the field. As the analyte is biochemical and/or biological in nature, classical analytical procedures are often not applicable for this purpose, and new types of analytical methods need to be developed. A sandwich type enzyme-linked immunosorbent assay (ELISA) was developed to detect and quantify Bacillus thuringiensis var. kurstaki delta-endotoxin in nine commercial formulations that are being used for forest insect control in North America. Formulation ingredients (formulants) interfered with the toxin response in three of the nine formulations, and the total protein needed to be separated from formulants for ELISA. Total protein levels were consistently higher (about 40 to 60%), in all formulations than the delta-endotoxin content because of the presence of inactive proteins. Results of three types of force-feeding bioassays [using spruce budworm, Choristoneura fumiferana (Clemens.)] viz., with diluted formulations, formulation extracts, and redissolved protein precipitates, showed that the formulants contributed to bioactivity and had to be removed to assess toxicity of the BTK protein alone. The delta-endotoxin content of formulations correlated better with the LD50's (ng of total protein/larva required for 50% mortality) of the redissolved protein precipitates of formulations than with those of the diluted formulations and formulation extracts. As more microbial pesticides are being introduced for forest insect control, it is crucial that analytical methods are in place for quality control of commercial formulations, both from the standpoint of improved efficacy and environmental safety.

Author Information

Sundaram, KMS
Forestry Canada, Forest Pest Management Institute, Sault Ste. Marie, Ontario, Canada
Sundaram, A
Forestry Canada, Forest Pest Management Institute, Sault Ste. Marie, Ontario, Canada
Gee, SJ
Pesticide Biotechnology Laboratory, Department of Entomology and Environmental Toxicology, University of California, Davis, California, USA
Harrison, RO
Immunosystems, Inc., Scarborough, Maine, USA
Hammock, BD
University of California, Davis, California, USA.
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Developed by Committee: E35
Pages: 297–312
DOI: 10.1520/STP13017S
ISBN-EB: 978-0-8031-5298-4
ISBN-13: 978-0-8031-1890-4