The quartz crystal microbalance (QCM) method can detect the amounts of proteins adsorbed on biomaterials by a frequency shift of an oscillating quartz crystal. In the present study, we observed the adsorption behavior of proteins onto a titanium surface by using 27-MHz QCM. As proteins, two cell adhesive proteins, fibronectin and collagen, and albumin were evaluated.
The QCM apparatus used was a 27-MHz AFFINIX Q
Nμ (Initium Co., Ltd., Tokyo, Japan) with 500-
μL cells. The temperature was maintained at 25±1℃ and the solution in the cells was stirred during the measurements. A titanium sensor was prepared by titanium sputter-coating onto an Au electrode. Bovine albumin, human plasma fibronectin and atelocollagen were dissolved in phosphate-buffered saline (PBS) solution (pH 7.4) at a concentration of 50
μg/mL. Each protein solution was injected into the PBS solution in the cells of the QCM apparatus. The decrease of frequency was monitored and the amounts of proteins adsorbed onto the titanium surface at 30 min after the injection were calculated by Sauerbrey's equation. The apparent reaction rate,
Kobs, was also obtained.
A slight frequency decrease was observed upon injection of albumin, and the frequency decrease for fibronectin adsorption was larger than that for albumin adsorption. Collagen significantly showed the largest decrease of frequency shift. Significant differences in the adsorbed amounts were seen among the three proteins. Namely, collagen exhibited the significantly largest adsorption amount on titanium, and albumin showed the significantly smallest amount.
Kobs for collagen was significantly the smallest, and that of albumin was significantly the largest. A small value of
Kobs corresponds to rapid adsorption of protein on titanium.
It is presumed that electrostatic interaction was dominant in the protein adsorption on titanium. Both albumin and fibronectin had more electrostatic repulsion to titanium based on their isoelectric points. In contrast, the attractive force increased between collagen and titanium because of the positively charged collagen. The QCM method was found to be a useful tool for monitoring not only the adsorption amounts of proteins but also adsorption behavior including the adsorption rate.
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