Make a calibration curve for the fluorescence of Rhodamine B as a function of the concentration of AuNP that the Rhodamine is dissolved in
Take images of and measure the fluorescence of the supernatants of the samples synthesized on February 17, 2016 that had been incubated in Rhodamine overnight (i.e., the Rhodamine had been added on February 23, 2016)
Take images of the samples that were synthesized on February 23, 2016; incubate these samples in Rhodamine for two hours; take images of the samples after the incubation; and then measure the fluorescence of the supernatants of these samples
Make new AuNP fiber samples for use next week
Use HPLC to measure the impurities in acetylsalicylic acid
Protocol
We used the same Rhodamine and chloroauric acid samples as last week for all samples described today. We made a new lysozyme stock that was 46.79µM.
Incubation of Samples Synthesized on February 23, 2016 in Rhodamine B
Because we had incubated the samples that we synthesized on February 17, 2016 in Rhodamine B for 24 hours, we decided to incubate the samples that we synthesized on February 23, 2016 for 3.5 hours. Then, we could try to determine whether incubating the samples in Rhodamine for longer periods of time helped incorporate more Rhodamine into the fibers.
The following table shows the amount of Rhomdaine B added to each sample.
Table 1: Rhodamine B Added to Samples Synthesized on February 23, 2016
We then covered the tops of the test tubes in parafilm and incubated the samples at room temperature for 3.5 hours.
Fluorescence Measurements
First, we took fluorescence measurements of Rhodamine in varying concentrations of AuNP.
MICHAEL CAN YOU PUT THE CONCENTRATIONS/DILUTIONS OF AUNP AND RHODAMINE YOU USED FOR THE CALIBRATION CURVE HERE THANKS
Next, we took fluorescence measurements of the supernatants of each of the samples synthesized on February 17, 2016 after being incubated in Rhodamine for 24 hours. We then took fluorescence measurements of the supernatants of each of the samples synthesized on February 23, 2016 after being incubated in Rhodamine for 3.5 hours. For each measurement, we used 200µL of sample.
We also took one measurement of a 1µM Rhodamine standard (in DI water).
For all the fluorescence measurements, we used the following parameters:
Start 540 nm End 700 nm Ex: 520 nm
Synthesis of AuNP Fiber Samples
We made new AuNP fiber samples according to the following table. (Note that we did not add any Rhodamine. We are going to add the Rhodamine to the samples on March 1, 2016 after the samples have been incubated in the oven.)
Table 2: Components of the AuNP Fiber Samples Synthesized Today
Each of the samples had a total volume of 5mL and were made in glass test tubes. We sealed the test tubes by covering the top with aluminum foil and then taping the foil down with labeling tape so that no vapors could escape.
We then incubated the samples in the oven at 80°C for four hours.
HPLC
MATT PUT YOUR PROTOCOL HERE PLZ THANKSALOT:)
Data, Observations, and Analysis
Calibration Curve for Fluorescence of Rhodamine B as a Function of AuNP Concentration
MICHAEL CAN YOU PUT THE CALIBRATION CURVE HERE WHEN YOU'RE DONE THAT WOULD BE SUPER DUPER COOL THX
Fluorescence Data for Supernatants of Samples
Since only measured the fluorescence of one Rhodamine standard, we could not make a calibration curve to determine the change in the concentration of Rhodamine in the supernatant based on our fluorescence measurements of the supernatants. Next week, we will measure Rhodamine standards so that we can determine the change in Rhodamine concentration.
Below is our raw fluorescence data for these samples. See the entry for March 2, 2016 for the analysis of these samples (we're going to take measurements of Rhodamine standards on March 2 so that we can analyze all of the supernatants from this week and in the future.)
Figure 1: Raw Fluorescence of Supernatants of Samples Synthesized on February 17, 2016 After 24 Hour Incubation in Rhodamine
The above image shows the raw fluorescence data for each sample we measured. The fluorescence is shown as a function of the wavelength of fluorescence (nm).
Figure 2: Raw Fluorescence of Supernatants of Samples Synthesized on February 23, 2016 After 2 Hour Incubation in Rhodamine
Images of Samples Synthesized on February 17, 2016 After 24 Hour Incubation in Rhodamine
Figure 1: Samples Synthesized on February 17, 2016 After 24 Hour Incubation in Rhodamine
This image was taken after the samples had been incubating in Rhodamine for 24 hours. From left to right, the samples are:
A_200uM_01
A_200uM_02
A_200uM_03
A_100uM_01
A_100uM_02
A_100uM_03
A_50uM_01
A_50uM_02
A_50uM_03
au_lys_after
The concentration of Rhodamine added to the A_200uM samples was 1µM. The concentration of Rhodamine added to the A_100uM samples was 0.1µM. The concentration of Rhodamine added to the A_50uM samples was 0.01µM. No Rhodamine was added to the au_lys_after sample; this sample was a blank.
The pink hue in the samples, especially the A_200uM samples, is due to the Rhodamine and not the presence of AuNP.
Images of Samples Synthesized on February 23, 2016 Before Addition in Rhodamine
Figure 2a-c: Samples Synthesized on February 23, 2016 Before Addition in Rhodamine
Figure 2a: From Left to Right: Samples A_1uM_01, A_1uM_02, A_1uM_03, A_1uM_04 These samples were synthesized on February 23, 2016. They were going to be incubated in 1µM Rhodamine for 3.5 hours. This image was taken before the incubation in Rhodamine.Figure 2b: From Left to Right: Samples A_0.1uM_01, A_0.1uM_02, A_0.1uM_03, A_0.1uM_04 These samples were synthesized on February 23, 2016. They were going to be incubated in 0.1µM Rhodamine for 3.5 hours. This image was taken before the incubation in Rhodamine.Figure 2c: From Left to Right: Samples A_0.01uM_01, A_0.01uM_02, A_0.01uM_03, A_0.01uM_04 These samples were synthesized on February 23, 2016. They were going to be incubated in 0.01µM Rhodamine for 3.5 hours. This image was taken before the incubation in Rhodamine.
Images of Samples Synthesized on February 23, 2016 After 3.5 Hour Incubation in Rhodamine
Figure 3: Samples Synthesized on February 23, 2016 After 3.5 Hour Incubation in Rhodamine
From left to right, the samples in this image are:
A_0.01uM_02
A_0.01uM_01
A_0.01uM_03
A_0.01uM_04
A_0.1uM_03
A_0.1uM_02
A_0.1uM_01
A_0.1uM_04
A_1uM_03
A_1uM_02
A_1uM_01
A_1uM_04
This image was taken after the samples had been incubated in Rhodamine for 2 hours. We did not measure the fluorescence of the supernatants of these samples until they had incubated in Rhodamine for 3.5 hours.
All of the samples ending in _04 did not receive any Rhodamine. These were blanks.
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