Whole sediment toxicity testing is an increasingly used means of assessing concentration dependent responses of benthic biota to contaminants as it enables sediments of single contaminants to be studied. However, concerns exist regarding the means of establishing heavy metal concentration gradients in sediments for use in toxicity testing. Metal spiking causes a plethora of poorly monitored changes in sediment geochemistry. Such changes to sediment chemistry induced by metal spiking, the procedural adjustments to minimise these and the resultant effects to organism response were the focus of this research. The addition of Cu and Zn (Method 1) into three local estuarine sediments, decreased sediment pH and increased pore-water concentrations well beyond the range of naturally contaminated sediment. Spiking with Cd, however, produced little variation in the sediment geochemistry at concentrations of up to 15× sediment guidelines (22.5 µg/g). The extent of pore-water and pH disturbance decreased in sediments with a more robust binding potential. Unrealistic partitioning of metals to the pore-water, as occurred when sediment is spiked to any significant extent, increases the sensitivity of test species to this exposure pathway. Consequently, past methods of metal spiking sediment are likely to have resulted in overestimated contaminant toxicity. Neutralisation of metal-spiked sediment minimised the geochemical disturbance of metal addition, reducing levels of Cu and Zn in the pore-water by greater than an order of magnitude for sediments adjusted to pH 8 (Method 2). pH again decreased immediately after metal addition, although to a lesser degree than with Method 1. Adjustment of sediments to pH 8 required increasing amounts of base (OH-) over the spike metal concentration gradient, which contributed to different pH values at equilibration, and thus fluctuations in redox potential and pore-water metals over the concentration gradient. Despite pH neutralisation, concentrations of pore-water spiked metal remained above levels in naturally contaminated sediment. The use of pH adjustment was therefore insufficient to produce spiked sediment comparable to contaminated in situ sediment for toxicity tests. The serial dilution of high spiked metal concentration, pH 8 adjusted sediment (method 3) decreased the concentration of pore-water metals, while concurrently minimising geochemical variation over the concentration gradient. The serial sediment dilution method proved the best method for spiking Cu into sediment, producing pore-water metal concentrations and partition coefficients similar to that observed in naturally contaminated sediment. Procedural changes to Cu spike methods (i.e. Method 1, 2 & 3) substantially altered the observed toxic response of the estuarine bivalve Austriella cf plicifera. The exposure of A. cf plicifera to Cu spiked sediment using Method 2 and 3 eliminated the occurrence of bivalve mortality (up to 55%) observed in Cu-spiked sediment prepared using Method 1. The rate of bivalve reburial was highest for Cu-spiked sediment prepared using Method 3 (M3 up to M2 both up to M1). Complete inhibition of reburial was observed at lower spike concentrations in Method 1 sediment than Method 2 (Cu550 and Cu1100, respectively) with reburial merely reduced in Method 3 (Cu1100). The method of spiking also altered tissue accumulation, however, this was influenced by the degree of bivalve reburial. Changing lethal and sublethal response of A. cf plicifera with the method in which Cu was spiked suggests that whole-sediment toxicity tests employing questionable metal spiking procedures will not be reflective of organism responses in situ contamination. This has implications for the accuracy of sediment guideline concentrations established using concentration-response data in studies using poor metal spike procedures. The optimal spike method (M3) was used to study the effect of minor changes in pH (6.6, 7.2, 7.6) on bivalve sublethal response. Maximum levels of organism reburial were observed at sediment pH of 7.2. In pH 6.6 sediment higher pore-water Cu concentration caused slower reburial and for pH 7.6 sediment high AVS is thought to have produced a similar response. Optimised metal spiking procedures provide a proven reproducible means of producing metal contaminated environments, which more accurately resemble contaminated natural sediments.