What you are given
- Acrylamide %: 10
- Gel volume: 10 mL
Formula: recipe by %T
Gels
How do you make an SDS-PAGE gel?
Each component is a fixed fraction of the total volume: acrylamide scaled from the 30% stock to the target percentage, buffer, SDS, then APS and TEMED last, with water making up the difference.
Worked example
Every number below is computed by the calculator, not typed in
The working
- Acrylamide:
V · (% / 30) = 10 · (10 / 30)=3.333 mL - Tris (1.5 M Tris-HCl pH 8.8):
V · 0.25 = 10 · 0.25=2.5 mL - SDS 10%:
V · 0.01=0.1 mL - APS 10%:
V · 0.01=0.1 mL - TEMED:
V · 0.0004=0.004 mL - Water (by difference):
V − (acrylamide + Tris + SDS + APS + TEMED)=3.963 mL
10% resolving gel, 10 mL total. Combine:
What each symbol means
- Vacryl: Acrylamide/bis stock volume (mL)
- V: Total gel volume (mL)
- P: Target acrylamide percentage (%)
- Pstock: Stock acrylamide percentage (30%) (%)
Source: SDS-PAGE recipe, fixed named stocks. A printable one-page version is on the formula sheet.
Where this usually goes wrong
- Adding APS and TEMED before you are ready to pour. Polymerisation starts immediately.
- Using resolving-gel buffer in the stacking layer, which ruins the stack.
- Handling unpolymerised acrylamide without gloves. It is a neurotoxin.
Try it yourself
Two problems, with the working revealed
Problem 1 SDS-PAGE gel
Work out the volume of every component.
- Gel layer: Stacking
- Acrylamide %: 4.5
- Gel volume: 8 mL
Show the answer and the working
4.5% stacking gel, 8 mL total. Combine:
- Acrylamide:
V · (% / 30) = 8 · (4.5 / 30)=1.2 mL - Tris (1.0 M Tris-HCl pH 6.8):
V · 0.125 = 8 · 0.125=1 mL - SDS 10%:
V · 0.01=0.08 mL - APS 10%:
V · 0.01=0.08 mL - TEMED:
V · 0.001=0.008 mL - Water (by difference):
V − (acrylamide + Tris + SDS + APS + TEMED)=5.632 mL
Open this in the calculator to change the numbers.
Problem 2 SDS-PAGE gel
Work out the volume of every component.
- Acrylamide %: 8
- Gel volume: 10 mL
Show the answer and the working
8% resolving gel, 10 mL total. Combine:
- Acrylamide:
V · (% / 30) = 10 · (8 / 30)=2.667 mL - Tris (1.5 M Tris-HCl pH 8.8):
V · 0.25 = 10 · 0.25=2.5 mL - SDS 10%:
V · 0.01=0.1 mL - APS 10%:
V · 0.01=0.1 mL - TEMED:
V · 0.0004=0.004 mL - Water (by difference):
V − (acrylamide + Tris + SDS + APS + TEMED)=4.629 mL
Open this in the calculator to change the numbers.
Do it with your own numbers.
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