The chloroauric acid used in the paper by Bakshi et al (2011)1 was not at hand at the time of experimentation. The gold, though interesting for subsequent experimentation, is not necessary to the creation of workable protein films, however. The purpose of this small-scale experiment is to merely gain familiarity with the materials of the experiment and to test the sensitivity of the experimental protocols (i.e., a very dry run)
Method
Bovine serum albumin
Bovine serum albumin (BSA: Calbiochem Omnipur, Fraction V, lyophilized powder) was prepared in a 0.015 mM solution with distilled water (10mL total volume). The solution was placed in a glass test tube and heated in an intermittently circulating water bath, loosely capped, for approximately 70 minutes at 52°C. The solution appeared clear with a small amount of 'froth' on the surface of the solution evident upon mixing, indicating it was not simply water.
A zein protein solution with total volume 10mL was prepared with:
Zein protein
-1 g zein (MP Biochemicals) [10% w/v]
-9 mL 95% ethanol [90% v/v]
-300 μL glycerol [30% on zein weight basis]
The zein solution appeared a vibrant orange-red that was fairly viscous -- much thinner in consistency than glycerol, rather like a light oil. The zein was chilled on ice for 80 minutes in a closed 15mL BD Falcon™ tube while the BSA solution was prepared.
In the paper, the suspension of nanoparticles was prepared by mixing variable concentrations of BSA with variable concentrations of chloroauric acid. The highest concentration of BSA used in the paper was 0.015 mM. Once the suspension of nanoparticles had been made, the BSA-Au solution was mixed into the zein solution at 10% v/v. At the point before mixing, the highest concentration of BSA would be at 0.0075 -- halved by equal parts mixing with the HAuCl4. The final concentration in the mixed protein solution of BSA would then be around 0.625 μM. For the small experiment performed here, the BSA solution before mixing with zein was at 0.015 mM and the concentration of BSA after mixing with zein was, naturally, twice the amount in the paper: 1.25μM.
Once mixed, the 11mL of solution was pipetted onto two glass 9cm petri dishes, 5.5mL into each. These were labeled accordingly and one was placed under a fume hood at room temperature (in the lab, just under 25°C) and the other was placed in an incubator at 37°C. They were left overnight.
1Bakshi, S. M.; Kaur, H.; Khullar, P.; Banipal, T. S.; Kaur, G.; Singh, N. Protein Films of Bovine Serum Albumin Conjugated Gold Nanoparticles: A Synthetic Route from Bioconjugated Nanoparticles to Biodegradable Protein Films. J. Phys. Chem. C2011, 115 (7), 2982-2992. [Link to the paper]
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