User:Keyun Wang/Notebook/Experimental Biological Chemistry I/2012/10/03 - Revision history

Keyun Wang: /* Procedure */

2012-10-27T03:39:22Z

Procedure

←Older revision		Revision as of 03:39, 27 October 2012
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	* UV-vis showed concentration protein elution at 280nm absorptivity, and indicated the elution test tube with most concentrated ADA protein.		* UV-vis showed concentration protein elution at 280nm absorptivity, and indicated the elution test tube with most concentrated ADA protein.
	* Two test tubes of ADA protein were collected in a 15mL falcon tube, and flow through of the protein are stored in 4C.		* Two test tubes of ADA protein were collected in a 15mL falcon tube, and flow through of the protein are stored in 4C.
		+	* For specific parameter for the column purifier, please refer to [[User:Melissa Novy/Notebook/CHEM-571/2012/10/03\|Melissa's Notebook]] for the details of the column purifier and conditions which the ADA protein was run at.

	==Discussion==		==Discussion==

Keyun Wang at 03:36, 27 October 2012

2012-10-27T03:36:09Z

←Older revision		Revision as of 03:36, 27 October 2012
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	==Procedure==		==Procedure==
-	* Column from ~~AKTA~~ purifier were washed with 100mL binding buffer at pH 7.5 with Tris, NaCl, and 30mM of imidazol.	+	* Column from purifier were washed with 100mL binding buffer at pH 7.5 with 500nM Tris, 200nM NaCl, and 30mM of imidazol.
-	*'''[[User:Abigail E. Miller\|Abigail E. Miller]] 17:56, 7 October 2012 (EDT)''':what are the tris nad NaCl concnetrations? what is AKTA purifier?	+
	* Protein supernatant filtered from yesterday was injected into column.		* Protein supernatant filtered from yesterday was injected into column.
	* Protein supernatant were run through the column with AKTA purifier. Flow through were collected in 50ml falcon tube just in case protein failed to bind to column.		* Protein supernatant were run through the column with AKTA purifier. Flow through were collected in 50ml falcon tube just in case protein failed to bind to column.
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	* UV-vis showed concentration protein elution at 280nm absorptivity, and indicated the elution test tube with most concentrated ADA protein.		* UV-vis showed concentration protein elution at 280nm absorptivity, and indicated the elution test tube with most concentrated ADA protein.
	* Two test tubes of ADA protein were collected in a 15mL falcon tube, and flow through of the protein are stored in 4C.		* Two test tubes of ADA protein were collected in a 15mL falcon tube, and flow through of the protein are stored in 4C.
-
-	*'''[[User:Abigail E. Miller\|Abigail E. Miller]] 17:56, 7 October 2012 (EDT)''':more information - flow rates? time? any specific parameters for the FPLC.

	==Discussion==		==Discussion==

Abigail E. Miller: /* Procedure */

2012-10-07T21:56:37Z

Procedure

←Older revision		Revision as of 21:56, 7 October 2012
Line 12:		Line 12:
	==Procedure==		==Procedure==
	* Column from AKTA purifier were washed with 100mL binding buffer at pH 7.5 with Tris, NaCl, and 30mM of imidazol.		* Column from AKTA purifier were washed with 100mL binding buffer at pH 7.5 with Tris, NaCl, and 30mM of imidazol.
		+	*'''[[User:Abigail E. Miller\|Abigail E. Miller]] 17:56, 7 October 2012 (EDT)''':what are the tris nad NaCl concnetrations? what is AKTA purifier?
	* Protein supernatant filtered from yesterday was injected into column.		* Protein supernatant filtered from yesterday was injected into column.
	* Protein supernatant were run through the column with AKTA purifier. Flow through were collected in 50ml falcon tube just in case protein failed to bind to column.		* Protein supernatant were run through the column with AKTA purifier. Flow through were collected in 50ml falcon tube just in case protein failed to bind to column.
Line 17:		Line 18:
	* UV-vis showed concentration protein elution at 280nm absorptivity, and indicated the elution test tube with most concentrated ADA protein.		* UV-vis showed concentration protein elution at 280nm absorptivity, and indicated the elution test tube with most concentrated ADA protein.
	* Two test tubes of ADA protein were collected in a 15mL falcon tube, and flow through of the protein are stored in 4C.		* Two test tubes of ADA protein were collected in a 15mL falcon tube, and flow through of the protein are stored in 4C.
		+
		+	*'''[[User:Abigail E. Miller\|Abigail E. Miller]] 17:56, 7 October 2012 (EDT)''':more information - flow rates? time? any specific parameters for the FPLC.

	==Discussion==		==Discussion==

Keyun Wang at 19:00, 3 October 2012

2012-10-03T19:00:15Z

←Older revision		Revision as of 19:00, 3 October 2012
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	{\|{{table}} width="800"		{\|{{table}} width="800"
	\|-		\|-
-	\|style="background-color: #EEE"\|[[Image:owwnotebook_icon.png\|128px]]<span style="font-size:22px;"> ~~Project name~~</span>	+	\|style="background-color: #EEE"\|[[Image:owwnotebook_icon.png\|128px]]<span style="font-size:22px;"> Experimental Biological Chemistry I</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>}}		\|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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	\| colspan="2"\|		\| colspan="2"\|
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-	==~~Entry title~~==	+	==Purpose==
-	* ~~Insert content here~~...	+	* Run protein supernatant through nickel-filled column in order to separate adenosine deaminase from the rest of protein in supernatant.
		+	* Remake Au/BSA solutions with different Gold to BSA ratios for experiments for next week.
		+
		+	==Procedure==
		+	* Column from AKTA purifier were washed with 100mL binding buffer at pH 7.5 with Tris, NaCl, and 30mM of imidazol.
		+	* Protein supernatant filtered from yesterday was injected into column.
		+	* Protein supernatant were run through the column with AKTA purifier. Flow through were collected in 50ml falcon tube just in case protein failed to bind to column.
		+	* 200mL elution buffer were run through the column while collecting the binded ADA protein in separate test tubes.
		+	* UV-vis showed concentration protein elution at 280nm absorptivity, and indicated the elution test tube with most concentrated ADA protein.
		+	* Two test tubes of ADA protein were collected in a 15mL falcon tube, and flow through of the protein are stored in 4C.
		+
		+	==Discussion==
		+	* Protein supernatant were run through the column. The nickel inside the column binds to the histidine head of Adenosine Deaminase protein. Adenosine Deaminase is the only protein among protein supernatant with histidine head, thus ADA protein should be the only protein sticking to the nickel column.
		+	* Once elution buffer were run, the high imidazol concentration within elution buffer is subject to competitive binding with the nickel in the column, allowing the nickel to release adenosine deaminase.
		+	* ADA in response is released from the column and collected into elution test tubes.


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-	\|}	+

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Keyun Wang: Autocreate 2012/10/03 Entry for User:Keyun_Wang/Notebook/Experimental_Biological_Chemistry_I

2012-10-03T18:45:01Z

Autocreate 2012/10/03 Entry for User:Keyun_Wang/Notebook/Experimental_Biological_Chemistry_I

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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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