When possible, we analysed separately each mussel to increase the
statistical significance. This was feasible for the cellular viability test,
NRRA, comet assay and micronucleus test,while the quantity of soft tissues
necessary to perform the enzymatic activities and the oxidative
stress biomarkers did not allow this accuracy level. Thus, for the determination
of these biochemical responses, we pooled 3 whole mussels
(×3 replicates) to reach the indispensable quantity of tissue. The protocols
followed in this research are well standardized and efficaciously
used in many previous studies by our research group (Binelli et al.,
2009a,b, 2012; Parolini and Binelli, 2012, 2013).