User:Moira M. Esson/Notebook/CHEM-571/2013/09/25: Difference between revisions

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==Entry title==
==Objectives==
* Insert content here...
# Run electrophoresis(SDS-PAGE) of pepsin and pepstatin samples prepared on [[User:Moira_M._Esson/Notebook/CHEM-571/2013/09/24|2013/09/24]]
# Finish running UV-Vis samples of pepsin and pepstatin samples prepared on [[User:Moira_M._Esson/Notebook/CHEM-571/2013/09/24|2013/09/24]]
<br>
==Electrophoresis==
*Electrophoresis was run on a Bio-Rad mini protean system with pre-cast TGX gels.
*Used Coomassive blue staining for the development
<br>
'''General Protocol''':
# Prepare the gel and assemble the electrophoresis cell
#*Refer [http://www.bio-rad.com/webroot/web/pdf/lsr/literature/Bulletin_1658100.pdf here] for exact details on how to assemble the unit.
#*Remove comb and tape from the gel
#*Rinse the walls with running buffer (30.0 Tris, 44.1g glycine, 10g SDS, H<sub>2</sub>O to 1L)
#*Assemble cell
#*Fill the inner and outer buffer chambers with running buffer
#Load samples prepared yesterday
#*Note: Also loaded two hemeglobin samples for comparison and pepsin and pepstatin reactions left overnight(these were prepared in the same fashion as [[User:Moira_M._Esson/Notebook/CHEM-571/2013/09/24|2013/09/24]]. In total ran 12 samples.
#*Note: Loaded entire 20μL samples
#Run electrophoresis for 30 minutes at 200V
#Develop stain of the gel
#*Place gel in fixant solution (40% methanol, 10% acetic acid, 50% water) for 30 minutes
 
<br>
{| {{table}}
| align="center" style="background:#f0f0f0;"|'''Well Numbers'''
| align="center" style="background:#f0f0f0;"|'''Sample'''
|-
| 1||Hemoglobin
|-
| 2||Pepsin
|-
| 3||Pepstatin
|-
| 4||0.5hrs Pepsin
|-
| 5||0.5 hrs Pepstatin
|-
| 6||1 hr Pepsin
|-
| 7||1 hr Pepstatin
|-
| 8||1.5hrs Pepsin
|-
| 9||1.5 hrs Pepstatin
|-
| 10||2hrs Pepsin
|-
| 11||2hrs Pepstatin
|-
| 12||Hemoglobin
|-
|
|} 




Revision as of 10:20, 25 September 2013

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Objectives

  1. Run electrophoresis(SDS-PAGE) of pepsin and pepstatin samples prepared on 2013/09/24
  2. Finish running UV-Vis samples of pepsin and pepstatin samples prepared on 2013/09/24


Electrophoresis

  • Electrophoresis was run on a Bio-Rad mini protean system with pre-cast TGX gels.
  • Used Coomassive blue staining for the development


General Protocol:

  1. Prepare the gel and assemble the electrophoresis cell
    • Refer here for exact details on how to assemble the unit.
    • Remove comb and tape from the gel
    • Rinse the walls with running buffer (30.0 Tris, 44.1g glycine, 10g SDS, H2O to 1L)
    • Assemble cell
    • Fill the inner and outer buffer chambers with running buffer
  2. Load samples prepared yesterday
    • Note: Also loaded two hemeglobin samples for comparison and pepsin and pepstatin reactions left overnight(these were prepared in the same fashion as 2013/09/24. In total ran 12 samples.
    • Note: Loaded entire 20μL samples
  3. Run electrophoresis for 30 minutes at 200V
  4. Develop stain of the gel
    • Place gel in fixant solution (40% methanol, 10% acetic acid, 50% water) for 30 minutes



Well Numbers Sample
1 Hemoglobin
2 Pepsin
3 Pepstatin
4 0.5hrs Pepsin
5 0.5 hrs Pepstatin
6 1 hr Pepsin
7 1 hr Pepstatin
8 1.5hrs Pepsin
9 1.5 hrs Pepstatin
10 2hrs Pepsin
11 2hrs Pepstatin
12 Hemoglobin



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