- Prepare serial dilutions of 0.1 M Tris buffer stock solution at the following concentrations: 10 mM, 1 mM, 100 μM, 10 μM.
- Remake Au and BSA stock solutions to check for contamination.
- Resuspend Au/BSA fibers in different concentrations of pH 8 or 10 Tris buffer.
- Protocol for Resuspending Au/BSA Fibers
- Au/BSA solutions made on 2012/08/31 by Dr. Miller were obtained.
- The solutions were centrifuged at 3000 rpm, 25°C, for 5 min, such that the fibers formed a pellet at the bottom of the centrifuge tube.
- 1 mL of 10 mM pH 8 Tris buffer was added to each pellet and pipetted up and down to resuspend the fibers.
- Step two was repeated with Tris buffer at concentrations of 1 mM, 100 μM, and 10 μM on fibers with arbitrary Au/BSA mole ratios.
- Steps 2 and 3 were repeated with pH 10 Tris buffer.
- UV-vis spectroscopy was conducted on the resuspended fiber solutions.
- The AuNP/BSA solution ratios were considered arbitrary for this analysis, as the objective was to determine only whether AuNP/BSA fibers could be resuspended in Tris buffer. The presence of a peak at around 540 nm indicated the presence of AuNPs and therefore successful resuspension of the fibers.
- Absorbance of AuNP/BSA fibers resuspended in Tris buffer at concentrations of 100 μM, 10 μM, 1 mM, and 10 mM and pH of 8 or 10.
- The presence of a peak at around 540 nm for all solutions indicates that at least some AuNPs became resuspended in solution. However, as no absorbance spectra of pure Tris buffer has been taken, it may be that the peaks are a result of absorbance by the buffer.
- Abigail E. Miller 09:51, 17 September 2012 (EDT):why didn't you measure Tris buffer? plot absorbance versus tris buffer concentration? does that show any trend? you need to repeat this and demonstrate whether it is due to tris or not. First measure Tris buffer absorbance spectrum to see if it even abosrbance around 530 nm. then Measure Tris buffer as blanks (at each concentration) and then measure solutions with AuNPs right after mixing and then later.
- Please refer to Keyun Wang's entry for observations on making and centrifuging Au/BSA solutions.
- Please refer to Dhea Patel's entry for volumes and calculations used in remaking the Au/BSA solutions.