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Edman Sequencing 에 사용 가능한 buffer 와 권장 membrane

Edman Sequencing 에 사용 가능한 buffer 와 권장 membrane
1. Stock sample buffer solution ( 5x without urea )
0.5 M Sucrose 42.78 g
15% SDS 37.50 g
312.5 mM Tris 9.50 g
10mM Na.EDTA 0.925 g
Add DW of 225ml
Heat gently and Adjust pH 6.9 by 1N HCl
Finally, up-to 250ml, Keep at 4℃ (Aliquots)
2. 5x working sample buffer
Add
100ul of BME
100ul of BPB solution (0.05% v/v)
to 2ml of stock sample buffer
3. 2x working sample buffer
Add
1.5ml of DW
50ul of BPB (0.05% v/v)
to 1ml of stock sample buffer
4. Towbin buffer : Transfer buffer (1)
25 mM Tris 3.03 g
252 mM Glycine 14.4 g
20% Methanol 200 ml ( adjust % depend on protein MW )
Up-to 1 liter by DW
Pre-chill the buffer before use
‘Do not adjust pH
5. CAPs buffer : Transfer buffer (2)
10 mM CAPs 2.21 g
10% Methanol 100 ml
Up-to 1 liter by DW
Adjust to pH 11 by 1N(10N) NaOH solution
Pre-chill the buffer before use
6. The other recommendable transfer buffer is NuPAGE Transfer buffer: Invitrogen
7. Staining buffer for PVDF membrane ( Ponceus S/1.0% acetic acid )
0.02 % CBB R-250 10 mg
40% Methanol 20 ml
Up-to 50 ml by DW
8. Destaining buffer for PVDF membrane ( DW )
50% Methanol 40 ml
Up-to 100 ml by DW
9. Recommendable PVDF membranes
- Problott (Applied Biosystems)
- Immobilon-PSQ (Millipore)
- BioTrace (Pall)
- Sequi-Blot (BioRad)
10. Chemical reagent and solvent
- Recommend Chemical reagent to you as ultrapure grade
- Recommend solvent to you as analytical grade
8. Destaining buffer for PVDF membrane ( DW )
50% Methanol 40 ml
Up-to 100 ml by DW
9. Recommendable PVDF membranes
- Problott (Applied Biosystems)
- Immobilon-PSQ (Millipore)
- BioTrace (Pall)
- Sequi-Blot (BioRad)
10. Chemical reagent and solvent
- Recommend Chemical reagent to you as ultrapure grade
- Recommend solvent to you as analytical grade
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