To assess the potential effects of Cry2Aa-expressing insect-resistant Bt rice on Chrysoperla sinica larvae, we conducted two tritrophic bioassays using a non-target (Laodelphax striatellus) and a target herbivore (Chilo suppressalis) as prey. None of the tested life-table parameters of C. sinica did differ when fed with L. striatellus nymphs reared on either Bt or control rice plants. Similarly, C. sinica larval survival and development were not affected when fed C. suppressalis larvae that were reared on Cry2Aa-contained artificial diet compared to those fed control diet. However, the 7-day larval weight was significantly decreased in the Bt treatment and none of the C. sinica larvae developed to the adult stage. To clarify whether the observed effects were due to the direct toxicity of Cry2Aa or prey-quality mediated, we conducted a dietary exposure assay in which the toxicity of Cry2Aa to C. sinica larvae was tested. Potassium arsenate (PA) was included as a positive control. None of the tested life-table parameters of C. sinica was adversely affected when fed Cry2Aa at 500 μg/ml sucrose solution. In contrast, C. sinica larvae were adversely affected by feeding on sucrose solution containing PA. In the feeding assays, exposure of C. sinica larvae to Cry2Aa was confirmed by ELISA. Our results demonstrate that C. sinica larvae are not sensitive to Cry2Aa at concentrations exceeding the levels that the larvae may encounter in Bt rice fields. Consequently the detrimental effects observed in the tritrophic studies using Bt rice-fed C. suppressalis as prey can be attributed to the decreased prey quality due to the sensitivity of C. suppressalis larvae to Cry2Aa.