User:Keyun Wang/Notebook/Experimental Biological Chemistry I/2012/11/20: Difference between revisions
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==Purpose== | ==Purpose== | ||
* ADA protein were transferred from dialysis tubing into 15mL falcon tubes | |||
* Make Au/ADA samples with the following mole ratios: 60 - 70 - 80 - 90 - 100 - 110 - 120 - 130 - 140 - 150, with ADA fraction 2 after dialysis. | * Make Au/ADA samples with the following mole ratios: 60 - 70 - 80 - 90 - 100 - 110 - 120 - 130 - 140 - 150, with ADA fraction 2 after dialysis. | ||
* | * Un-dialyzed Au/ADA samples and Au/HRP made on [[User:Keyun Wang/Notebook/Experimental Biological Chemistry I/2012/11/14|2012/11/14]] were run on UV-vis spectrometer and Atomic Absorption Spectrometer in order to compare spectra results for Au/ADA, Au/HRP, Au/Lysozyme, and Au/BSA. | ||
* | * Resuspend Au/HRP samples in different concentration and pH of Tris buffer to test ionic strength. | ||
==Procedure for Dialysis== | |||
* Dialysis beaker containing dialysis tubing enclosed with ADA protein fractions were taken from the 4°C cold room into room temperature lab room. | |||
* The dialysis clips were taken off from dialysis tubing. ADA protein fractions were poured into a sterile 15mL falcon tubes. | |||
* This process was repeated for all three protein fractions: ADA fraction 1&3 made on [[User:Keyun Wang/Notebook/Experimental Biological Chemistry I/2012/11/06|2012/11/06]], ADA fraction 2 made on [[User:Keyun Wang/Notebook/Experimental Biological Chemistry I/2012/11/06|2012/11/06]], and ADA fraction purified on [[User:Keyun Wang/Notebook/Experimental Biological Chemistry I/2012/10/03|2012/10/03]]. | |||
* ADA fractions in 15mL falcon tubes were stored in 4°C refrigerator. | |||
==Procedure for making dialyzed Au/ADA samples== | |||
* Au/ADA samples were made with the following mole ratios: | |||
60 - 70 - 80 - 90 - 100 - 110 - 120 - 130 - 140 - 150 | |||
* Stock solution of HAuCl<sub>4</sub> was made on [[User:Keyun Wang/Notebook/Experimental Biological Chemistry I/2012/09/05|2012/09/05]] with a concentration of 10.5uM. | |||
* Stock solution for ADA was the ADA protein fraction made in [[User:Keyun Wang/Notebook/Experimental Biological Chemistry I/2012/10/03|2012/10/03]] after dialysis. The ADA stock solution has a concentration of 58.36μM. | |||
* Volume of ADA protein was set at 137.1uL and a range of HAuCl<sub>4</sub> was used from 45.71μL to 114.3μL. Water was added to the sample to increase the volume of sample to 8mL. Volumes of each reactants are shown in table below: | |||
{| {{table}} | |||
| align="center" style="background:#f0f0f0;"|'''Au/ADA ratio''' | |||
| align="center" style="background:#f0f0f0;"|'''ADA added[uL]''' | |||
| align="center" style="background:#f0f0f0;"|'''HAuCl4 Added [uL]''' | |||
| align="center" style="background:#f0f0f0;"|'''Water Added[uL]''' | |||
| align="center" style="background:#f0f0f0;"|'''[ADA]final[uM]''' | |||
| align="center" style="background:#f0f0f0;"|'''[HAuCl4]final[uM]''' | |||
|- | |||
| 60||137.1||45.71||7817.2||1||60 | |||
|- | |||
| 70||137.1||53.3||7809.6||1||70 | |||
|- | |||
| 80||137.1||60.9||7802||1||80 | |||
|- | |||
| 90||137.1||68.6||7794.4||1||90 | |||
|- | |||
| 100||137.1||76.2||7786.8||1||100 | |||
|- | |||
| 110||137.1||83.8||7779.1||1||110 | |||
|- | |||
| 120||137.1||91.4||7771.5||1||120 | |||
|- | |||
| 130||137.1||99||7763.9||1||130 | |||
|- | |||
| 140||137.1||106.7||7756.3||1||140 | |||
|- | |||
| 150||137.1||114.3||7748.7||1||150 | |||
|- | |||
| | |||
|} | |||
* The samples were made in 15mL non-sterile falcon tubes. After all reactants were added, samples are capped and wrapped around with aluminum foil. | |||
* Samples were placed in incubator at 85°C for 4 hours then cooled down to room temperature. | |||
==Procedure for Running UV-vis spectrometer and Atomic Absorption spectrometer on Au/ADA and Au/HRP samples== | |||
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Revision as of 12:14, 27 November 2012
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Purpose
Procedure for Dialysis
Procedure for making dialyzed Au/ADA samples
60 - 70 - 80 - 90 - 100 - 110 - 120 - 130 - 140 - 150
Procedure for Running UV-vis spectrometer and Atomic Absorption spectrometer on Au/ADA and Au/HRP samples |