User:Dhea Patel/Notebook/Experimental Biological Chemistry Notebook/2012/10/24

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(Autocreate 2012/10/24 Entry for User:Dhea_Patel/Notebook/Experimental_Biological_Chemistry_Notebook)
Current revision (00:53, 7 December 2012) (view source)
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==Objective==
==Objective==
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#To make biocompatible protein films with specific reactive sites
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*to run Fluorescence on Luminol-HRP assays.
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#Explore the solution conditions (pH, ionic strength, buffer, stabilizers)  that produce stable protein – gold nanoparticle suspensions
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 +
==Description==
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 +
Please refer to table for volumes and concentrations of HRP, Luminol, Phenol, Buffer, and hydrogen peroxide used.
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{| {{table}}
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| align="center" style="background:#f0f0f0;"|''''''
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| align="center" style="background:#f0f0f0;"|'''Volume and Stock Concentration Added to Cuvette'''
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| align="center" style="background:#f0f0f0;"|''''''
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| align="center" style="background:#f0f0f0;"|''''''
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| align="center" style="background:#f0f0f0;"|''''''
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| align="center" style="background:#f0f0f0;"|'''Volume Added to Cuvette [µL]'''
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| align="center" style="background:#f0f0f0;"|''''''
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|-
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| Trial||Luminol||H<sub>2</sub>O<sub>2</sub>||4-Iodophenol||HRP||Water||0.833mM Sodium Carbonate Buffer
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|-
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| 1||454 µL at 0.564 mM||112.7 µL at 5 mM||13.2 µL at 18 mM||33 µL at 9.6 µM||1033 µL||x
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|-
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| 2||454 µL at 0.564 mM||112.7 µL at 5 mM||13.2 µL at 18 mM||33 µL at 9.6 µM||1033 µL||x
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|-
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| 3||454 µL at 0.564 mM||112.7 µL at 5 mM||13.2 µL at 18 mM||33 µL at 9.6 µM||2000 µL||x
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|-
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| 4||454 µL at 0.564 mM||112.7 µL at 5 mM||13.2 µL at 18 mM||66 µL at 0.96 µM||2000 µL||x
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|-
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| 5||454 µL at 0.564 mM||112.7 µL at 5 mM||13.2 µL at 18 mM||33 µL at 2.3 µM||x||1000 µL
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|-
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| 6||454 µL at 0.564 mM||112.7 µL at 5 mM||13.2 µL at 18 mM||16.5 µL at 2.3 µM||x||1000 µL
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|-
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| 7||454 µL at 0.564 mM||112.7 µL at 5 mM||13.2 µL at 18 mM||16.5 µL at 0.96 µM||x||1000 µL
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|-
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| 8||454 µL at 0.564 mM||112.7 µL at 5 mM||13.2 µL at 18 mM||16.5 µL at 2.3 µM||x||2000 µL
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|}
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==Data==
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[[Image:Trail_6.png]]
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*All the trails, except trial 6 had a straight peak, showing do drop off of the fluorescence. Trail 6, containing, 454uL 0.564mM Luminol, 112.7uL 5mM hydrogen peroxide, 16.5uL 0.9uM HRP, 13.2 uL 18mM 4-Iodophenol, and 1000uL Carbonate Buffer, showed a distinct drop off. Tests should be run, keeping the molarity of 0.9uM constant and varying other factors to determine the optimum molar ratio.
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Objective

  • to run Fluorescence on Luminol-HRP assays.

Description

Please refer to table for volumes and concentrations of HRP, Luminol, Phenol, Buffer, and hydrogen peroxide used.

' Volume and Stock Concentration Added to Cuvette ' ' ' Volume Added to Cuvette [µL] '
TrialLuminolH2O24-IodophenolHRPWater0.833mM Sodium Carbonate Buffer
1454 µL at 0.564 mM112.7 µL at 5 mM13.2 µL at 18 mM33 µL at 9.6 µM1033 µLx
2454 µL at 0.564 mM112.7 µL at 5 mM13.2 µL at 18 mM33 µL at 9.6 µM1033 µLx
3454 µL at 0.564 mM112.7 µL at 5 mM13.2 µL at 18 mM33 µL at 9.6 µM2000 µLx
4454 µL at 0.564 mM112.7 µL at 5 mM13.2 µL at 18 mM66 µL at 0.96 µM2000 µLx
5454 µL at 0.564 mM112.7 µL at 5 mM13.2 µL at 18 mM33 µL at 2.3 µMx1000 µL
6454 µL at 0.564 mM112.7 µL at 5 mM13.2 µL at 18 mM16.5 µL at 2.3 µMx1000 µL
7454 µL at 0.564 mM112.7 µL at 5 mM13.2 µL at 18 mM16.5 µL at 0.96 µMx1000 µL
8454 µL at 0.564 mM112.7 µL at 5 mM13.2 µL at 18 mM16.5 µL at 2.3 µMx2000 µL

Data

Image:Trail_6.png

  • All the trails, except trial 6 had a straight peak, showing do drop off of the fluorescence. Trail 6, containing, 454uL 0.564mM Luminol, 112.7uL 5mM hydrogen peroxide, 16.5uL 0.9uM HRP, 13.2 uL 18mM 4-Iodophenol, and 1000uL Carbonate Buffer, showed a distinct drop off. Tests should be run, keeping the molarity of 0.9uM constant and varying other factors to determine the optimum molar ratio.


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