User:Moira M. Esson/Notebook/CHEM-571/2013/10/16: Difference between revisions
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== | ==Objectives== | ||
* | * Test the enzymatic/catalytic activity of HRP-AuNPs using luminol and UV/vis with the goal of determining the activity of HRP-AuNPs. | ||
* Calculate the absorbance prepared Citrate-Au solutions and BSA-Au solutions as standards (solutions were previously prepared on [[User:User:Moira M. Esson/Notebook/CHEM-571/2013/08/28|08/28/2013]] | |||
<br> | |||
==Catalytic activity testing== | |||
*Catalytic activity was tested using UV/vis spectroscopy. | |||
*3mL sample of luminol, H2O2 and buffer was placed in cuvette with micro-stir bar. After 1min., 100μL HRP-Au was added. | |||
*A spectrum was taken every 15seconds for twenty minutes. | |||
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'''Sample preparation''' | |||
*Stock solutions were prepared by [[User:Matt Hartings/Notebook/AU Biomaterials Design Lab/2013/10/16|Matt Harting's]] | |||
Buffer:0.6140g Tris in 1L, pH set to 8 with HCl ---> 5.1mM | |||
Luminol:Dissolve 12.9mg luminol in 300uL of DMSO and add to 50mL buffer---> 1.46mM | |||
H2O2: 312uL 30% H2O2 into 100mL buffer ---> Should be 45mM | |||
<br> | |||
*Preparation of HRP-AuNP samples: | |||
**General Description of the samples: | |||
***The final concentration of luminol is such that the absorbance at 333nm is less than one(this was confirmed using UV/vis) | |||
***The final concentration of hydrogen peroxide is fifty times that of the luminol. | |||
***Using the 60:1 [Au:HRP] solution, 100uL will be used | |||
***Add buffer so that the final concentration is 3mL | |||
'''Calculations for preparation of sample:''' | |||
#Luminol stock solution preparation 5mL 7.26E-5 M luminol: 0.261mL luminol stock added to 5mL buffer. | |||
#H2O2 stock preparation of 5mL 0.0038M(this is 50x more concentrated): 0.169mL stock in 5mL buffer. | |||
*Final 3mL solution of luminol, H2O2 and buffer was prepared by adding 1mL luminol, 1mL H2O2 and 1mL buffer. | |||
*Concentration of luminol was chosen based on previous luminol data on [[User:Moira M. Esson/Notebook/CHEM- 571/2013/10/01|10/01/2013]]. | |||
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'''Testing of HRP-AuNP activity''' | |||
Revision as of 13:58, 21 October 2013
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Objectives
Catalytic activity testing
Buffer:0.6140g Tris in 1L, pH set to 8 with HCl ---> 5.1mM Luminol:Dissolve 12.9mg luminol in 300uL of DMSO and add to 50mL buffer---> 1.46mM H2O2: 312uL 30% H2O2 into 100mL buffer ---> Should be 45mM
Calculations for preparation of sample: #Luminol stock solution preparation 5mL 7.26E-5 M luminol: 0.261mL luminol stock added to 5mL buffer. #H2O2 stock preparation of 5mL 0.0038M(this is 50x more concentrated): 0.169mL stock in 5mL buffer.
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