  Tris(hydroxymethyl)aminomethane (Tris) has been used as the buffer in
bioactivity testing for over two decades and has become a standard choice for
the scientific community. While it is believed to be non-interacting, the
extent of its interactions with titanium oxide surfaces has not been
systematically studied. Here, we use experimental (zeta potential measurements)
and computational (molecular dynamics) approaches to evaluate the interaction
of Tris with a rutile surface and how it affects the adsorption of other
molecules relevant in biomedical in vitro testing. We show that the interaction
of Tris with the rutile surface is strong and significantly affects the
interaction of other organic residues with the surface. These strong
interactions are compounded by the Tris concentration in the in vitro testing
protocol which is much higher compared to other components. Our findings
indicate that the kinetics observed in in vitro tests will be strongly
influenced by the presence of Tris as a buffering agent when compared to the
natural CO2 buffer in blood. These results reveal that considering the so-far
neglected active role of Tris in in vitro testing is critically needed and that
in vitro protocols using CO2 partial pressure as the buffering agent should
yield more reliable results.
