Bradford protein assay to determine protein concentration

In a standard assay to measure GA concentration, the calibration curve of GA was first conducted. Two hundred microliters of GA solutions (50, 100, 150, 200, 250, 300, 400, 450, and 500 μg/ml) in ultrapure water were prepared, and 200 μl of ultrapure water without protein was used as blank control. Two milliliters of Bradford reagent was added into the above solutions. After 5 min, the reaction solutions (1 ml) were transferred into a 96-well plate (five wells for each sample, 200 μl per well), and then, their absorbance at 595 nm was recorded by a microplate reader (Multiskan GO, Thermo Fisher Scientific, USA). The calibration curve plotting the linear relationship for GA concentration versus the solution absorbance at 595 nm was then obtained (fig. S2). The concentrations of GA in the synthesized GA/Pt or GA/Pt-NAS were then determined. Briefly, the sample containing GA/Pt or GA/Pt-NAS was diluted by ultrapure water to obtain a solution with protein concentration in the range of 50 to 500 μg/ml. After that, the same procedure was performed as aforementioned to measure the absorbance of the sample at 595 nm. GA/Pt or GA/Pt-NAS in ultrapure water without treatment was tested as a control to exclude the influence of Pt nanoparticles in GA on the absorbance. Last, the concentration of GA in the sample was calculated according to the calibration curve. The calibration curves for ProK, casein, and DNase I were measured by a similar protocol (fig. S2), and the protein concentrations in ProK/Pt, ProK/Pt-NAS, casein, DNase I/Pt, and DNase I/Pt-NAS were calculated according to the standard curves.

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