User:Matthew R Skorski/Notebook/471 - Exp BioChem/2016/02/03

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Objective

Today's objective was to measure the fluorescence and UV-Vis absorption of the supernatant from yesterday's samples. We also synthesized new fiber samples because not all of the samples from yesterday successfully formed fibers.

Protocol

Making New Samples

We followed the same protocol that is listed for February 2, 2016. We used the same stock solutions of gold and rhodamine. The only difference was that we made a fresh stock solution of lysozyme; the concentration of this stock was 39.98µM, so we used 138.98µL of this solution in each sample. We adjusted the volume of water added to 785.64µL to account for this change.

Measurements of Samples

First, we spun down all of the samples from yesterday at 13,000 RPM for 2 minutes. Then, we took fluorescence and UV-Vis measurements for the supernatants. We also took fluorescence measurements of some of the samples we synthesized today just before placing them in the oven. (We only measured samples to which we had added rhodamine.)

Fluorescence Measurements

The parameters for our fluorescence measurements were Start: 540 nm End: 700 nm Excitation: 520 nm. We used 200µL of sample for each measurement.

UV-Vis Absorbance Measurements

We used 0.5mL of sample diluted in 2.5mL of DI water for each measurement. We used Ocean Optics to measure the absorbance.

Rhodamine B absorbs light at a wavelength of 542nm.

Data and Analysis

We noticed that there was a peak shift in the fluorescence measurements for many of the samples that had rhodamine. A few possible explanations for this shift include:

  • FRET: When the sample is excited with light, energy in the rhodamine may be transferred to the nanoparticles, causing a peak shift
  • Rhodamine may have been acting as a reducing agent, like the lysozyme
  • A new compound may have been synthesized

We took UV-Vis absorbance measurements of the samples in order to determine whether new compounds had been synthesized.


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