Metals on metal implants have long been used in arthroplasties because of their robustness and low dislocation rate. Several relatively low-corrosion metals have been used in arthroplasty, including 316L stainless steel, titanium, and cobalt–chromium–molybdenum alloy. Debris from these implants, however, has been found to cause inflammatory responses leading to unexpected failure rates approaching 10% 7 years surgery. Safety assessment of these materials traditionally relies on the use of simple two-dimensional assays, where cells are grown on the surface of the material over a relatively short time frame. It is now well-known that the composition and stiffness of the extracellular matrix (ECM) have a critical effect on cell function. In this work, we have evaluated how cobalt ions influence the assembly of type I collagen, the principle component of the ECM in bone. We found that cobalt had a significant effect on collagen matrix formation, and its presence results in local variations in collagen density. This increase in heterogeneity causes an increase in localized mechanical properties but a decrease in the bulk stiffness of the material. Moreover, when collagen matrices contained cobalt ions, there was a significant change in how the cells interacted with the collagen matrix. Fluorescence images and biological assays showed a decrease in cell proliferation and viability with an increase in cobalt concentration. We present evidence that the cobalt ion complex interacts with the hydroxyl group present in the carboxylic terminal of the collagen fibril, preventing crucial stabilizing bonds within collagen formation. This demonstrates that the currently accepted toxicity assays are poor predictors of the longer-term biological performance of a material.