Approaches to perform dose confirmations for non-target organism studies by insect bioassay

Monsanto Company has developed and marketed crops that produce insecticidal (Bt) proteins and are thereby protected against insect feeding damage. As part of the environmental safety assessment for these products, an assessment of potential risks to non-target organisms (NTO) is conducted. As part of these NTO studies, dose confirmations are performed by insect bioassay to assess the functional activity, stability and homogeneity of the Bt protein in the test diet.

Two approaches can be used to confirm the concentration of biologically active Bt protein in the test diet. One approach to confirming the concentration of the Bt protein by inverse-prediction from a standard curve prepared with a Bt protein reference standard. If the measured concentration is within 20% of the nominal concentration the dose is considered to be confirmed. However, if the result exceeds 20% of the nominal concentration, the measured concentration is reported. A second approach uses dose response curves prepared for the test diet and for a reference standard. The median effect values for the test diet are statistically compared with the median effect values for the reference standard. If the estimated median effect values are significantly different between the test diet and the reference standard, the concentration in the test diet is calculated using the ratio of the median effect values. An illustration and discussion of both approaches will be presented.