User:Mary Mendoza/Notebook/CHEM572 Exp. Biological Chemistry II/2013/02/05 - Revision history

Mary Mendoza: /* Data */

2013-05-08T22:13:29Z

Data

←Older revision		Revision as of 22:13, 8 May 2013
Line 11:		Line 11:


-
-	~~==Data==~~
-	~~{\| {{table}}~~
-	~~\| align="center" style="background:#f0f0f0;"\|'''Concentration of Adenosine (uM)'''~~
-	~~\| align="center" style="background:#f0f0f0;"\|'''Average Velocity over 30s (nm/sec)'''~~
-	\|-
-	~~\| 100\|\|0.000345161~~
-	\|-
-	~~\| 10\|\|0.000109677~~
-	\|-
-	~~\| 1\|\|2.58065E-05~~
-	\|}
-
-	~~{\| {{table}}~~
-	~~\| align="center" style="background:#f0f0f0;"\|'''1/[adenosine] (1/uM)'''~~
-	~~\| align="center" style="background:#f0f0f0;"\|'''1/[velocity] (sec/nm)'''~~
-	\|-
-	~~\| 0.01\|\|2897.196262~~
-	\|-
-	~~\| 0.1\|\|9117.647059~~
-	\|-
-	~~\| 1\|\|38750~~
-	\|}
-
-
-
-	~~Lineweaver-Burk Plot of the data obtained today.~~
-	~~[[Image:Lineweaver-Burk_Plot_2013-02-05.png]]~~
-
-	~~{\| {{table}}~~
-	~~\| 1/Vmax\|\|4015.59923~~
-	\|-
-	~~\| '''Vmax'''\|\|'''0.000249029 nm/sec'''~~
-	\|-
-	~~\| 1/Km\|\|0.111964094~~
-	\|-
-	~~\| '''Km'''\|\|'''8.93143474 uM'''~~
-	\|}

Mary Mendoza: /* Procedure */

2013-05-08T22:13:08Z

Procedure

←Older revision		Revision as of 22:13, 8 May 2013
Line 10:		Line 10:


-	~~==Procedure==~~
-	~~#UVProbe was opened.~~
-	~~##Window > 1. Kinetics~~
-	~~##Methods Icon~~
-	~~###Wavelength: 265nm~~
-	~~###Duration: 300 seconds (5minutes)~~
-	~~###OK~~
-	~~#Shimadzu CPS-Controller was set to 25°C.~~
-	~~##wait for the temperature to raise to 25°C~~
-	~~#place the sample in the cell and click start.~~

-	*Phosphate buffer was used as a blank and was used to create the baseline.
-	*A spectrum of 1mM adenosine was taken. The absorbance exceeded the capabilities of the spectrometer, so 100μM adenosine (0.6mL 500μM stock adenosine diluted with 100μM by adding 2.4mL buffer) was used as the maximum concentration.
-	*Once it was determined that 100μM adenosine created a clear spectra, kinetics was run at 265nm using 2.38mL phosphate buffer, 600μL of 500μM stock adenosine, and 20μL ADA (as outlined in the table above).
-	**The velocity was determined by clicking Operations > Activity Table > dAbs. The average of the dAbs values is the average slope (velocity).
-	*Then, an assay was run at 265nm using 2.38mL phosphate buffer, 600μL of 50μM stock adenosine (to make a final concentration of 10μM), and 20μL ADA (as outlined in the table above).
-	**The absorbance was close to 0, indicating that the ideal adenosine concentration likely ranges between 10μM and 100μM.
-	*Then, an assay was run at 265nm using 2.38mL phosphate buffer, 600μL of 5μM stock adenosine (to make a final concentration of 1μM), and 20μL ADA (as outlined in the table above).
-	*From the spectra obtained today, the following tables were created:
-	**This table is the volumes of 5mM adenosine and phosphate buffer used to make the new adenosine stock solutions.
-	***[[Image:Screen_Shot_2013-02-05_at_9.01.20_PM.png]]
-	**This table is the updated table of the volumes and concentrations of solutions that will be added in the cuvette.
-	***[[Image:Screen_Shot_2013-02-05_at_9.01.33_PM.png]]

	==Data==		==Data==

Mary Mendoza: /* Calculations */

2013-05-08T22:12:53Z

Calculations

←Older revision		Revision as of 22:12, 8 May 2013
Line 8:		Line 8:


-	~~==Calculations==~~	+
-	*ADA Kinetics were run using the following volumes and concentrations.	+
-	**Note that the table has been adjusted based on the change in concentration of the adenosine stock solution.	+
-	~~[[Image:Screen_Shot_2013-02-05_at_11.59.42_AM.png]]~~	+
-	*The kinetics was run using UV-vis at 25°C at 265nm for 5 minutes.	+

	==Procedure==		==Procedure==

Mary Mendoza: /* ADA Kinetics Assay */

2013-05-08T22:12:43Z

ADA Kinetics Assay

←Older revision		Revision as of 22:12, 8 May 2013
Line 6:		Line 6:
	\| colspan="2"\|		\| colspan="2"\|
	<!-- ##### DO NOT edit above this line unless you know what you are doing. ##### -->		<!-- ##### DO NOT edit above this line unless you know what you are doing. ##### -->
-	~~==ADA Kinetics Assay==~~	+
-	* The objective of executing adenosine deaminase (ADA) kinetic assay is to obtain the slope or rate of the enzymatic reaction of the conversion of adenosine into inosine by ADA.	+
-	*The spectrophotometer used was the make of Shimadzu UV-2550 connected to a Shimadzu CPS-temperature controller.	+
-	* On the UV Probe system interface, the kinetics program was chosen under the Windows option. The parameters were adjusted from the method button.	+
-	* The rest of the procedure is taken from [[User:Dhea Patel/Notebook/CHEM 572: ADA&Inhibitor Kinetics/2013/02/05\|Dhea Patel's notebook]].	+

	==Calculations==		==Calculations==

Mary Mendoza: /* ADA Kinetics Assay */

2013-02-08T15:51:50Z

ADA Kinetics Assay

←Older revision		Revision as of 15:51, 8 February 2013
Line 7:		Line 7:
	<!-- ##### DO NOT edit above this line unless you know what you are doing. ##### -->		<!-- ##### DO NOT edit above this line unless you know what you are doing. ##### -->
	==ADA Kinetics Assay==		==ADA Kinetics Assay==
-	* The spectrophotometer used was the make of Shimadzu UV-2550 connected to a Shimadzu CPS-temperature controller.	+	* The objective of executing adenosine deaminase (ADA) kinetic assay is to obtain the slope or rate of the enzymatic reaction of the conversion of adenosine into inosine by ADA.
		+	*The spectrophotometer used was the make of Shimadzu UV-2550 connected to a Shimadzu CPS-temperature controller.
	* On the UV Probe system interface, the kinetics program was chosen under the Windows option. The parameters were adjusted from the method button.		* On the UV Probe system interface, the kinetics program was chosen under the Windows option. The parameters were adjusted from the method button.
	* The rest of the procedure is taken from [[User:Dhea Patel/Notebook/CHEM 572: ADA&Inhibitor Kinetics/2013/02/05\|Dhea Patel's notebook]].		* The rest of the procedure is taken from [[User:Dhea Patel/Notebook/CHEM 572: ADA&Inhibitor Kinetics/2013/02/05\|Dhea Patel's notebook]].

Mary Mendoza: /* ADA Kinetics Assay */

2013-02-08T05:58:23Z

ADA Kinetics Assay

←Older revision		Revision as of 05:58, 8 February 2013
Line 9:		Line 9:
	* The spectrophotometer used was the make of Shimadzu UV-2550 connected to a Shimadzu CPS-temperature controller.		* The spectrophotometer used was the make of Shimadzu UV-2550 connected to a Shimadzu CPS-temperature controller.
	* On the UV Probe system interface, the kinetics program was chosen under the Windows option. The parameters were adjusted from the method button.		* On the UV Probe system interface, the kinetics program was chosen under the Windows option. The parameters were adjusted from the method button.
-	* The rest of the procedure is taken from Dhea Patel's notebook.	+	* The rest of the procedure is taken from [[User:Dhea Patel/Notebook/CHEM 572: ADA&Inhibitor Kinetics/2013/02/05\|Dhea Patel's notebook]].

	==Calculations==		==Calculations==

Mary Mendoza: /* ADA Kinetics Assay */

2013-02-08T05:56:58Z

ADA Kinetics Assay

←Older revision		Revision as of 05:56, 8 February 2013
Line 9:		Line 9:
	* The spectrophotometer used was the make of Shimadzu UV-2550 connected to a Shimadzu CPS-temperature controller.		* The spectrophotometer used was the make of Shimadzu UV-2550 connected to a Shimadzu CPS-temperature controller.
	* On the UV Probe system interface, the kinetics program was chosen under the Windows option. The parameters were adjusted from the method button.		* On the UV Probe system interface, the kinetics program was chosen under the Windows option. The parameters were adjusted from the method button.
		+	* The rest of the procedure is taken from Dhea Patel's notebook.
		+
		+	==Calculations==
		+	*ADA Kinetics were run using the following volumes and concentrations.
		+	**Note that the table has been adjusted based on the change in concentration of the adenosine stock solution.
		+	[[Image:Screen_Shot_2013-02-05_at_11.59.42_AM.png]]
		+	*The kinetics was run using UV-vis at 25°C at 265nm for 5 minutes.
		+
		+	==Procedure==
		+	#UVProbe was opened.
		+	##Window > 1. Kinetics
		+	##Methods Icon
		+	###Wavelength: 265nm
		+	###Duration: 300 seconds (5minutes)
		+	###OK
		+	#Shimadzu CPS-Controller was set to 25°C.
		+	##wait for the temperature to raise to 25°C
		+	#place the sample in the cell and click start.
		+
		+	*Phosphate buffer was used as a blank and was used to create the baseline.
		+	*A spectrum of 1mM adenosine was taken. The absorbance exceeded the capabilities of the spectrometer, so 100μM adenosine (0.6mL 500μM stock adenosine diluted with 100μM by adding 2.4mL buffer) was used as the maximum concentration.
		+	*Once it was determined that 100μM adenosine created a clear spectra, kinetics was run at 265nm using 2.38mL phosphate buffer, 600μL of 500μM stock adenosine, and 20μL ADA (as outlined in the table above).
		+	**The velocity was determined by clicking Operations > Activity Table > dAbs. The average of the dAbs values is the average slope (velocity).
		+	*Then, an assay was run at 265nm using 2.38mL phosphate buffer, 600μL of 50μM stock adenosine (to make a final concentration of 10μM), and 20μL ADA (as outlined in the table above).
		+	**The absorbance was close to 0, indicating that the ideal adenosine concentration likely ranges between 10μM and 100μM.
		+	*Then, an assay was run at 265nm using 2.38mL phosphate buffer, 600μL of 5μM stock adenosine (to make a final concentration of 1μM), and 20μL ADA (as outlined in the table above).
		+	*From the spectra obtained today, the following tables were created:
		+	**This table is the volumes of 5mM adenosine and phosphate buffer used to make the new adenosine stock solutions.
		+	***[[Image:Screen_Shot_2013-02-05_at_9.01.20_PM.png]]
		+	**This table is the updated table of the volumes and concentrations of solutions that will be added in the cuvette.
		+	***[[Image:Screen_Shot_2013-02-05_at_9.01.33_PM.png]]
		+
		+	==Data==
		+	{\| {{table}}
		+	\| align="center" style="background:#f0f0f0;"\|'''Concentration of Adenosine (uM)'''
		+	\| align="center" style="background:#f0f0f0;"\|'''Average Velocity over 30s (nm/sec)'''
		+	\|-
		+	\| 100\|\|0.000345161
		+	\|-
		+	\| 10\|\|0.000109677
		+	\|-
		+	\| 1\|\|2.58065E-05
		+	\|}
		+
		+	{\| {{table}}
		+	\| align="center" style="background:#f0f0f0;"\|'''1/[adenosine] (1/uM)'''
		+	\| align="center" style="background:#f0f0f0;"\|'''1/[velocity] (sec/nm)'''
		+	\|-
		+	\| 0.01\|\|2897.196262
		+	\|-
		+	\| 0.1\|\|9117.647059
		+	\|-
		+	\| 1\|\|38750
		+	\|}
		+
		+
		+
		+	Lineweaver-Burk Plot of the data obtained today.
		+	[[Image:Lineweaver-Burk_Plot_2013-02-05.png]]
		+
		+	{\| {{table}}
		+	\| 1/Vmax\|\|4015.59923
		+	\|-
		+	\| '''Vmax'''\|\|'''0.000249029 nm/sec'''
		+	\|-
		+	\| 1/Km\|\|0.111964094
		+	\|-
		+	\| '''Km'''\|\|'''8.93143474 uM'''
		+	\|}

Mary Mendoza: /* ADA Kinetics Assay */

2013-02-05T17:44:52Z

ADA Kinetics Assay

←Older revision		Revision as of 17:44, 5 February 2013
Line 8:		Line 8:
	==ADA Kinetics Assay==		==ADA Kinetics Assay==
	* The spectrophotometer used was the make of Shimadzu UV-2550 connected to a Shimadzu CPS-temperature controller.		* The spectrophotometer used was the make of Shimadzu UV-2550 connected to a Shimadzu CPS-temperature controller.
-	* On the UV Probe system interface, the kinetics program was chosen under the ~~Tools~~ option. The parameters were adjusted from the method button.	+	* On the UV Probe system interface, the kinetics program was chosen under the Windows option. The parameters were adjusted from the method button.

Mary Mendoza: /* Entry title */

2013-02-05T17:40:34Z

Entry title

←Older revision		Revision as of 17:40, 5 February 2013
Line 6:		Line 6:
	\| colspan="2"\|		\| colspan="2"\|
	<!-- ##### DO NOT edit above this line unless you know what you are doing. ##### -->		<!-- ##### DO NOT edit above this line unless you know what you are doing. ##### -->
-	==~~Entry title~~==	+	==ADA Kinetics Assay==
-	* ~~Insert content here~~...	+	* The spectrophotometer used was the make of Shimadzu UV-2550 connected to a Shimadzu CPS-temperature controller.
		+	* On the UV Probe system interface, the kinetics program was chosen under the Tools option. The parameters were adjusted from the method button.

Mary Mendoza: Autocreate 2013/02/05 Entry for User:Mary_Mendoza/Notebook/CHEM572_Exp._Biological_Chemistry_II

2013-02-05T17:36:12Z

Autocreate 2013/02/05 Entry for User:Mary_Mendoza/Notebook/CHEM572_Exp._Biological_Chemistry_II

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{|{{table}} width="800"
|-
|style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;"> Project name</span>
|style="background-color: #F2F2F2" align="center"|<html><img src="/images/9/94/Report.png" border="0" /></html> [[{{#sub:{{FULLPAGENAME}}|0|-11}}|Main project page]]<br />{{#if:{{#lnpreventry:{{FULLPAGENAME}}}}|<html><img src="/images/c/c3/Resultset_previous.png" border="0" /></html>[[{{#lnpreventry:{{FULLPAGENAME}}}}{{!}}Previous entry]]<html>      </html>}}{{#if:{{#lnnextentry:{{FULLPAGENAME}}}}|[[{{#lnnextentry:{{FULLPAGENAME}}}}{{!}}Next entry]]<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html>}}
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==Entry title==
* Insert content here...


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