The lampricides TFM and niclosamide are added to streams to control invasive larval sea lamprey (Petromyzon marinus) populations in the Laurentian Great Lakes. Lampricide effectiveness depends upon TFM and niclosamide bioavailability which is influenced by both abiotic and biotic factors. For example, at lower pH, TFM bioavailability is higher because a greater proportion exists as un-ionized TFM (TFM-OH), which easily crosses the gills. At higher pH, however, the negatively charged ionized species of TFM (TFM-O⁻) predominates, which is less easily taken-up, meaning more TFM must be applied. Although water alkalinity does not directly affect TFM speciation, as a buffer it influences how much expired water crossing the gills is acidified by CO₂ and metabolic acid excretion. In poorly buffered waters, greater acidification of the expired water increases TFM bioavailability in the gill microenvironment than in better buffered, higher alkalinity waters where more TFM must be applied. Hence, sea lamprey and non-target fishes such as lake sturgeon (Acipenser fulvescens) are more sensitive to lampricides in low pH, low alkalinity waters. Differences in gill structure and microenvironment acidification might also explain why TFM sensitivity of young-of-the-year lake sturgeon approaches or exceeds that of sea lamprey in higher alkalinity waters. Other biotic factors such as body size and metabolic rate also contribute to differences in lampricide sensitivity. We conclude that better understanding of the abiotic and biotic factors influencing lampricide bioavailability can be used to refine treatment protocols to improve lampricide effectiveness and to better protect non-target fishes from lampricide toxicity.