Yu:CBB

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(CBB)
Current revision (14:28, 22 December 2009) (view source)
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{| border="0" cellpadding="10"
{| border="0" cellpadding="10"
|+'''Solution recipe for 500mL Calmodulin-Binding Buffer.'''
|+'''Solution recipe for 500mL Calmodulin-Binding Buffer.'''
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|width="600px" valign="top"|1. Measure 12.5mL of 1M Tris pH 8.0 into a 500mL beaker for a final concentration of 25mM Tris pH 8.0
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|width="600px" valign="top"|1. Measure 12.5mL of 1M Tris pH 8.0 into 100mL millipore H<sub>2</sub>O in a 500mL beaker
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|width="100px" valign="top"|12.5mL 1M Tris pH 8.0
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|width="150px" valign="top"|25mM Tris-HCl
|-
|-
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|2. Add 15mL of 5M NaCl for a final concentration of 150mM NaCl
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|2. Add 15mL of 5M NaCl
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|15mL 5M NaCl
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|150mM NaCl
|-
|-
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|3. Add 5mL of 10% NP-40 for a final concentration of 0.1% NP-40
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|3. Add 500 μL of 1M Magnesium Acetate
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|5mL 10% NP-40
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|1mM MgOAc
|-
|-
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|4. Add 500μL of 0.5M EDTA for a final concentration of 0.5mM EDTA
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|4. Add 500 μL of 1M Imidazole
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|500μL 0.5M EDTA
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|1mM Imidazole
 +
|-
 +
|5. Add 1 mL 1M CaCl<sub>2</sub>
 +
|2mM CaCl<sub>2</sub>
|-
|-
|5. Bring volume to 500mL with millipore H<sub>2</sub>O in a graduated cylinder
|5. Bring volume to 500mL with millipore H<sub>2</sub>O in a graduated cylinder
|Volume to 500mL
|Volume to 500mL
|-
|-
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|6. Complete with protease inhibitors immediately prior to use:
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|6. Complete immediately prior to use with 0.696 μL β-mercaptoethanol per 1 mL CBB
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|Complete:
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|10mM β-mercaptoethanol
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|-
+
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|colspan="2" align="center"|
+
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{|border="0" cellpadding="5"
+
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|+'''Protease Inhibitors for CBB'''
+
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|Add 1μL of DTT per 1mL CBB
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|1mM DTT
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|-
+
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|Add 1μL of Imidazole per 1mL CBB
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|1mM Imidazole
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|-
+
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|Add 1μL of magnesium acetate per 1mL CBB
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|1mM MgOAc
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|}
+
|-
|-
|7. Keep completed CBB on ice at all times
|7. Keep completed CBB on ice at all times
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|
 
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Current revision

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Solution recipe for 500mL Calmodulin-Binding Buffer.
1. Measure 12.5mL of 1M Tris pH 8.0 into 100mL millipore H2O in a 500mL beaker 25mM Tris-HCl
2. Add 15mL of 5M NaCl 150mM NaCl
3. Add 500 μL of 1M Magnesium Acetate 1mM MgOAc
4. Add 500 μL of 1M Imidazole 1mM Imidazole
5. Add 1 mL 1M CaCl2 2mM CaCl2
5. Bring volume to 500mL with millipore H2O in a graduated cylinder Volume to 500mL
6. Complete immediately prior to use with 0.696 μL β-mercaptoethanol per 1 mL CBB 10mM β-mercaptoethanol
7. Keep completed CBB on ice at all times
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