Background Dental amalgam is a widely used restorative material containing 50% elemental mercury that emits mercury vapor. No randomized clinical trials have determined whether there are adverse immunologic effects associated with this low-level mercury exposure in children. The objective of this study was to evaluate a sub-population of the New England Children’s Amalgam Trial (NECAT) for in vitro manifestations of immunotoxic effects of dental amalgam. Methods A randomized clinical trial in which children requiring dental restorative treatment were randomized to either amalgam for posterior restorations or resin composite. A total of 66 children, aged 6–10 years, were assessed for total white cell numbers, T-cell, B-cell, neutrophil and monocyte responsiveness over a five-year period. Owing to the small number of participants, the study is exploratory in nature with limited statistical power. Results The mean number of tooth surfaces restored during the five-year period was 7.8 for the amalgam group and 10.1 for composite group. In the amalgam group there was a slight, but not statistically significant, decline in responsiveness of T-cells and monocytes at 5–7 days post treatment; no differences were consistently observed at 6, 12 or 60 months. Conclusions This study confirms that treatment of children with dental amalgams leads to increased, albeit low level, exposure to mercury. In this exploratory analysis of immune function, amalgam exposure did not cause overt immune deficits, although small transient effects were observed 5–7 days post restoration. Clinical implications These findings suggest that immunotoxic effects of amalgam restorations in children need not be a concern when choosing this restorative dental material.