User Manual

3.3 SDS-PAGE Buffers
Table 3.2. Recommended Criterion precast gels and buffers for SDS-PAGE.
Gel Type Sample Buffer Running Buffer
Criterion TGX Laemmli (catalog #161-0737) Tris/glycine/SDS
Criterion TGX Stain-Free Optional: 2-mercaptoethanol (catalog #161-0732)
Criterion Tris-HCl (catalog #161-0710) or DTT
Criterion Stain Free (catalog #161-0611)
Criterion XT Bis-Tris XT sample buffer (catalog #161-0791) XT MES (catalog #161-0789)
Optional: XT reducing agent XT MOPS (catalog #161-0788)
(catalog #161-0792)
Criterion XT Tris-acetate XT Tricine (catalog #161-0790)
3.3.1 Running Buffers
See Appendix B for buffer formulations. Do not adjust pH.
3.4 Sample Preparation
1. Determine the appropriate concentration of sample to load (depends on the load volume and the
detection method used; see Chapter 12 for approximate stain sensitivities).
2. Dilute the sample with at least an equivalent volume of sample buffer with added reducing agent.
For nonreducing conditions, omit the reducing agent.
TGX and Tris-HCl Gels XT Gels (Bis-Tris and Tris-Acetate)
4.75 μl Laemmli sample buffer 5 μl XT sample buffer
0.25 μl β-mercaptoethanol 1 μl XT reducing agent
5 μl sample 1–14 μl sample
10 μl total volume Make up to 20 μl total volume with diH
2
O
3. Heat the diluted sample at 90–95°C for 5 min, or at 70°C for 10 min.
Tris/glycine/SDS (1x) 25 mM Tris, 192 mM glycine, 0.1% SDS
(pH 8.3) Dilute 100 ml 10x stock (catalog #161-0732) with 900 ml diH
2
O
XT MES (pH 6.4) Dilute 50 ml 20x stock (catalog #161-0789) with 950 ml diH
2
O
XT MOPS (pH 6.9) Dilute 50 ml 20x stock (catalog #161-0788) with 950 ml diH
2
O
XT Tricine (pH 8.2) Dilute 50 ml 20x stock (catalog #161-0790) with 950 ml diH
2
O
3.3.2 Sample Buffers
Laemmli 62.5 mM Tris-HCl, pH 6.8, 2% SDS, 25% glycerol, 0.01% (catalog #161-0737)
bromophenol blue, 5% β-mercaptoethanol or 100 mM DTT (added fresh)
XT Use XT sample buffer (catalog #161-0791) and XT reducing agent
(catalog #161-0792)
12 Technical Support: 1-800-4BIORAD • 1-800-424-6723 • www.bio-rad.com
Criterion Precast Gels