Download: Protein Gel Guide
Finding the right protein gel electrophoresis system is critical to getting consistent and accurate results you need for your research. Electrophoresis buffers and reagents are important components of the protein electrophoresis system. We offer a range of SDS-PAGE buffers, native buffers and reagents for gel casting, sample preparation, running, and transferring gels. Find the recommended electrophoresis buffers and reagents for each gel system below.
SDS sample buffers or loading buffers
Protein samples prepared for SDS-PAGE analysis are denatured by heating in the presence of a sample buffer containing 1–2% SDS or LDS with or without a reducing agent such as 20 mM DTT, 2-mercaptoethanol (BME) or TCEP. Sample buffers or loading buffers also contain glycerol so that they are heavier than water and sink neatly to the bottom of the buffer-submerged well when added to a gel. In addition, a negatively charged, low-molecular weight dye is also included in the sample buffer that will migrate at the buffer-front, enabling one to monitor the progress of electrophoresis. The most common tracking dyes for sample loading buffers are bromophenol blue, phenol red, and Coomassie blue. The table below summaries common sample buffers used in the different gel buffer systems.
| Gel system | Denaturing sample buffer (final concentrations) | Sample preparation |
|---|---|---|
| Tris-glycine | Tris-glycine SDS sample buffer: Tris HCl (63 mM), glycerol (10%), SDS (2%), bromophenol blue (0.0025%), pH 6.8 | Heat samples at 85°C for 2–5 minutes with sample buffer |
| Bis-Tris | LDS sample buffer: Tris base (141 mM), Tris HCl (106 mM), LDS (2%), EDTA (0.51 mM), SERVA Blue G-250 (0.22 mM), phenol red (0.175 mM), pH 8.5 | Heat samples at 70°C for 10 minutes with sample buffer |
| Tris-Acetate | LDS sample buffer: Tris base (141 mM), Tris HCl (106 mM), LDS (2%), EDTA (0.51 mM), SERVA Blue G-250 (0.22 mM), phenol red (0.175 mM), pH 8.5 | Heat samples at 70°C for 10 minutes with sample buffer |
| Tris-Tricine | Tricine SDS sample buffer: Tris HCl (450 mM), glycerol (12%), SDS (4%), Coomassie Blue G (0.00075%), phenol red (0.0025%), pH 8.45 | Heat samples at 85°C for 2–5 minutes with sample buffer |
| Zymogram | Tris-glycine SDS: Tris HCl (63 mM), glycerol (10%), SDS (2%), bromophenol blue (0.0025%), pH 6.8 | Do not heat samples, load immediately onto gel after adding to sample |
For fluorescence applications, we recommend that you use sample buffers that do not contain dyes that interfere in fluorescent channels (e.g., bromophenol blue). Fluorescent Compatible Sample Buffer does not contain bromophenol blue or other interfering dyes and can be used with reducing or nonreducing SDS-PAGE applications.
 | Find SDS sample buffer recipes in gel-specific product manuals in the documents tab. |
Native sample buffers
Protein samples prepared for native PAGE applications are prepared in sample buffers or loading buffers that do not contain detergents to maintain the native state of the proteins. The loading buffers contain glycerol so that they are heavier than water and sink neatly to the bottom of the buffer-submerged well when added to a gel. In addition, a negatively charged, low-molecular weight dye is also included in the sample buffer that will migrate at the buffer-front, enabling one to monitor the progress of electrophoresis.
When preparing samples for native applications, protein samples should not be heated and loaded immediately onto the gel after mixing with the sample buffer.
| Gel system | Native sample buffer (final concentrations) |
|---|---|
| Tris-glycine | Native sample buffer: Tris HCl (100 mM), glycerol (10%), bromophenol blue (0.00025%), pH 8.6 |
| Tris-acetate | Native sample buffer: Tris HCl (100 mM), glycerol (10%), bromophenol blue (0.00025%), pH 8.6 |
| IEF | IEF sample buffer pH 3-7: Lysine (40 mM), glycerol (15%) IEF sample buffer pH 3-10: Arginine (20 mM), Lysine (20 mM), glycerol (15%) |
| NativePAGE Bis-Tris | NativePAGE Sample Buffer (4X): Bis-Tris (50 mM), 6 N HCl, NaCl (50 mM), Glycerol (10%), Ponceau S (0.001%), pH 7.2 |
| Find native sample buffer recipes in gel-specific product manuals in the documents tab. |
SDS running buffers
In protein electrophoresis (discontinuous buffer system), the primary anion in the gel is different (or discontinuous) from the primary anion in the running buffer. The tables below have the recommended SDS running buffers or native running buffers for each of the different gel chemistry buffering systems.
| Gel system | SDS running buffer |
|---|---|
| Tris-glycine | Tris-glycine SDS: Tris base (25 mM), glycine (192 mM), SDS (0.1%), pH 8.3 |
| Bis-Tris | MES SDS: MES (50 mM), Tris base (50 mM), SDS (0.1%), EDTA (1 mM), pH 7.3 MOPS SDS: MOPS (50 mM), Tris base (50 mM), SDS (0.1%), EDTA (1 mM), pH 7.7 |
| Tris-acetate | Tris-acetate SDS: Tris base (50 mM), Tricine (50 mM), SDS (0.1%), pH 8.24 |
| Tris-tricine | Tricine-SDS: Tris base (100 mM), tricine (100 mM), SDS (0.1%), pH 8.3 |
Native running buffers
| Gel system | Native running buffer |
|---|---|
| Tris-glycine | Tris-glycine native buffer: Tris base (25 mM), glycine (192 mM), pH 8.3 |
| Tris-acetate | Tris-glycine native buffer: Tris base (25 mM), glycine (192 mM), pH 8.3 |
| IEF | IEF cathode buffer pH 3-7: Lysine (40 mM) IEF cathode buffer pH 3-10: Arginine (20 mM), lysine (20 mM) IEF anode buffer: phosphoric acid 85% (7 mM) |
| Zymogram | Tris-glycine SDS: Tris base (25 mM), glycine (192 mM), SDS (0.1%), pH 8.3 |
| Find SDS running buffer recipes and native running buffer recipes in gel specific product manuals in the documents tab. |
NuPAGE Bis-Tris SDS buffers & reagents
| Protein sample | Gel chemistry | Sample buffers | Running buffer | Transfer buffer |
|---|---|---|---|---|
| Broad-range MW (6-400 kDa) | Bis-tris | Denaturing: LDS sample buffer Reducing: Sample reducing agent | Small to medium-sized proteins (6–260 kDa): MES SDS buffer Medium to large-size proteins (14–260 kDa): MOPs SDS buffer Reduced samples: Antioxidant | Bis-tris transfer buffer |
Bolt Bis-Tris Plus SDS buffers & reagents
| Protein sample | Gel chemistry | Sample buffers | Running buffer | Transfer buffer |
|---|---|---|---|---|
| Broad-range MW (6-400 kDa) | Bis-tris | Denaturing: LDS sample buffer Reducing: Sample reducing agent | Small to medium-sized proteins (6–260 kDa): MES SDS buffer Medium to large-size proteins (14–260 kDa): MOPs SDS buffer Reduced samples: Antioxidant | Bis-tris transfer buffer |
Why LDS sample buffers
Bolt and NuPAGE LDS Sample Buffers are formulated with Coomassie G250 and phenol red as tracking dyes instead of bromophenol blue. Coomassie (SERVA) G250 gives a sharp dye front with both MES and MOPS SDS running buffers and migrates much closer to the moving ion front than bromophenol blue. Bromophenol blue runs more slowly than some peptides with the MES SDS Running Buffer. This ensures that small peptides do not run off the gel. The concentration of the tracking dye (Coomassie G250) is increased in Bolt and NuPAGE LDS Sample Buffers to enhance viewing of the dye front. LDS is used instead of SDS to allow the formulation of a 4X solution. The 4X solution provides the ability to have higher protein concentrations and less dilution of your protein sample.
Tris-glycine buffers & reagents
| Protein sample | Gel chemistry | Sample buffers | Running buffer | Transfer buffer |
|---|---|---|---|---|
| Broad-range MW (6-400 kDa) | Tris-glycine | Denaturing: Tris-glycine SDS sample buffer Native: Tris-glycine native sample buffer | Denaturing: Tris-glycine SDS buffer Native: Tris-glycine native buffer | Tris-glycine transfer buffer |
Tris-acetate buffers & reagents
| Protein sample | Gel chemistry | Sample buffers | Running buffer | Transfer buffer |
|---|---|---|---|---|
| High MW (40-500 kDa) | Tris-acetate | Denaturing: LDS sample buffer Native: Tris-glycine native sample buffer | Denaturing: Tris-acetate SDS buffer Native: Tris-glycine native buffer | Bis-tris transfer buffer |
Tricine SDS buffers & reagents
| Protein sample | Gel chemistry | Sample buffers | Running buffer | Transfer buffer |
|---|---|---|---|---|
| Low MW (2.5-40 kDa) | Tricine | Denaturing: Tricine SDS sample buffer | Tricine SDS buffer | Tris-glycine transfer buffer |
IEF gel recommended buffers
| Gel chemistry | Sample buffers | Running buffer |
|---|---|---|
| IEF gel |
Zymography gel buffers & reagents
| Gel type | Sample buffer | Running buffer | Gel development buffer | Gel renaturing buffer |
|---|---|---|---|---|
| Zymogram gel | Tris-glycine SDS sample buffer | Tris-glycine SDS running buffer | Zymogram development buffer | Zymogram renaturing buffer |
NativePAGE Bis-Tris gel buffers & reagents
| Gel type | Sample buffer | Running buffer | Gel development buffer |
|---|---|---|---|
| NativePAGE Bis-Tris Gel | Bis-Tris Transfer Buffer |
Gel casting
When pouring your own polyacrylamide gels, choosing high-quality reagents can help you achieve optimal electrophoresis results. SureCast buffers and reagents make it quick and easy to make high quality handcast Tris-glycine gels.
SDS-PAGE gel recipe for pour-your-own Tris-glycine gels
Prepare resolving gel solution according to the volumes in the following table. The volumes provided in the table are for a single gel. Scale volumes proportionally based on the number of gels to be cast.
| Polyacrylamide % | |||||||||
|---|---|---|---|---|---|---|---|---|---|
| Solution | 4% | 6% | 8% | 10% | 12% | 14% | 16% | 18% | 20% |
| SureCast Acrylamide (40%) | 0.8mL | 1.2mL | 1.6mL | 2.0mL | 2.4mL | 2.8mL | 3.3mL | 3.6mL | 4.0mL |
| SureCast Resolving Buffer | 2.0mL | 2.0mL | 2.0mL | 2.0mL | 2.0mL | 2.0mL | 2.0mL | 2.0mL | 2.0mL |
| Distilled water | 5.1mL | 4.7mL | 4.3mL | 3.9mL | 3.5mL | 3.1mL | 2.7mL | 2.3mL | 1.9mL |
| 10% SureCast Ammonium Persulfate (APS) | 80µL | 80µL | 80µL | 80µL | 80µL | 80µL | 80µL | 80µL | 80µL |
| SureCast TEMED** | 8µL | 8µL | 8µL | 8µL | 8µL | 8µL | 8µL | 8µL | 8µL |
**Add this last and mix well just before the gel is to be poured
Prepare stacking gel solution according to the following table. The volumes provided in the table are for a single gel. Scale volumes proportionally based on the number of gels to be cast. Note: Solutions do not require degassing.
| Solution | 4% |
|---|---|
| SureCast Acrylamide (40%) | 0.30 mL |
| SureCast Stacking Buffer | 0.75 mL |
| Distilled water | 1.92 mL |
| 10% SureCast Ammonium Persulfate (APS) | 30 µL |
| SureCast TEMED*** | 3 µL |
***Add this last and mix well just before the gel is to be poured
| Learn more about gel casting and find additional SDS-PAGE gel recipes. |
Ordering quick links
NuPAGE Bis-Tris Buffers
Bolt Bis-Tris Plus Buffers
Tris-Glycine Buffers
Tris-Acetate Buffers
Tricine System Buffers
IEF Buffers
Zymogram Buffers
NativePAGE Bis-Tris Buffers
Handcast Buffers & Reagents
SDS sample buffers or loading buffers
Protein samples prepared for SDS-PAGE analysis are denatured by heating in the presence of a sample buffer containing 1–2% SDS or LDS with or without a reducing agent such as 20 mM DTT, 2-mercaptoethanol (BME) or TCEP. Sample buffers or loading buffers also contain glycerol so that they are heavier than water and sink neatly to the bottom of the buffer-submerged well when added to a gel. In addition, a negatively charged, low-molecular weight dye is also included in the sample buffer that will migrate at the buffer-front, enabling one to monitor the progress of electrophoresis. The most common tracking dyes for sample loading buffers are bromophenol blue, phenol red, and Coomassie blue. The table below summaries common sample buffers used in the different gel buffer systems.
| Gel system | Denaturing sample buffer (final concentrations) | Sample preparation |
|---|---|---|
| Tris-glycine | Tris-glycine SDS sample buffer: Tris HCl (63 mM), glycerol (10%), SDS (2%), bromophenol blue (0.0025%), pH 6.8 | Heat samples at 85°C for 2–5 minutes with sample buffer |
| Bis-Tris | LDS sample buffer: Tris base (141 mM), Tris HCl (106 mM), LDS (2%), EDTA (0.51 mM), SERVA Blue G-250 (0.22 mM), phenol red (0.175 mM), pH 8.5 | Heat samples at 70°C for 10 minutes with sample buffer |
| Tris-Acetate | LDS sample buffer: Tris base (141 mM), Tris HCl (106 mM), LDS (2%), EDTA (0.51 mM), SERVA Blue G-250 (0.22 mM), phenol red (0.175 mM), pH 8.5 | Heat samples at 70°C for 10 minutes with sample buffer |
| Tris-Tricine | Tricine SDS sample buffer: Tris HCl (450 mM), glycerol (12%), SDS (4%), Coomassie Blue G (0.00075%), phenol red (0.0025%), pH 8.45 | Heat samples at 85°C for 2–5 minutes with sample buffer |
| Zymogram | Tris-glycine SDS: Tris HCl (63 mM), glycerol (10%), SDS (2%), bromophenol blue (0.0025%), pH 6.8 | Do not heat samples, load immediately onto gel after adding to sample |
For fluorescence applications, we recommend that you use sample buffers that do not contain dyes that interfere in fluorescent channels (e.g., bromophenol blue). Fluorescent Compatible Sample Buffer does not contain bromophenol blue or other interfering dyes and can be used with reducing or nonreducing SDS-PAGE applications.
| Find SDS sample buffer recipes in gel-specific product manuals in the documents tab. |
Native sample buffers
Protein samples prepared for native PAGE applications are prepared in sample buffers or loading buffers that do not contain detergents to maintain the native state of the proteins. The loading buffers contain glycerol so that they are heavier than water and sink neatly to the bottom of the buffer-submerged well when added to a gel. In addition, a negatively charged, low-molecular weight dye is also included in the sample buffer that will migrate at the buffer-front, enabling one to monitor the progress of electrophoresis.
When preparing samples for native applications, protein samples should not be heated and loaded immediately onto the gel after mixing with the sample buffer.
| Gel system | Native sample buffer (final concentrations) |
|---|---|
| Tris-glycine | Native sample buffer: Tris HCl (100 mM), glycerol (10%), bromophenol blue (0.00025%), pH 8.6 |
| Tris-acetate | Native sample buffer: Tris HCl (100 mM), glycerol (10%), bromophenol blue (0.00025%), pH 8.6 |
| IEF | IEF sample buffer pH 3-7: Lysine (40 mM), glycerol (15%) IEF sample buffer pH 3-10: Arginine (20 mM), Lysine (20 mM), glycerol (15%) |
| NativePAGE Bis-Tris | NativePAGE Sample Buffer (4X): Bis-Tris (50 mM), 6 N HCl, NaCl (50 mM), Glycerol (10%), Ponceau S (0.001%), pH 7.2 |
| Find native sample buffer recipes in gel-specific product manuals in the documents tab. |
SDS running buffers
In protein electrophoresis (discontinuous buffer system), the primary anion in the gel is different (or discontinuous) from the primary anion in the running buffer. The tables below have the recommended SDS running buffers or native running buffers for each of the different gel chemistry buffering systems.
| Gel system | SDS running buffer |
|---|---|
| Tris-glycine | Tris-glycine SDS: Tris base (25 mM), glycine (192 mM), SDS (0.1%), pH 8.3 |
| Bis-Tris | MES SDS: MES (50 mM), Tris base (50 mM), SDS (0.1%), EDTA (1 mM), pH 7.3 MOPS SDS: MOPS (50 mM), Tris base (50 mM), SDS (0.1%), EDTA (1 mM), pH 7.7 |
| Tris-acetate | Tris-acetate SDS: Tris base (50 mM), Tricine (50 mM), SDS (0.1%), pH 8.24 |
| Tris-tricine | Tricine-SDS: Tris base (100 mM), tricine (100 mM), SDS (0.1%), pH 8.3 |
Native running buffers
| Gel system | Native running buffer |
|---|---|
| Tris-glycine | Tris-glycine native buffer: Tris base (25 mM), glycine (192 mM), pH 8.3 |
| Tris-acetate | Tris-glycine native buffer: Tris base (25 mM), glycine (192 mM), pH 8.3 |
| IEF | IEF cathode buffer pH 3-7: Lysine (40 mM) IEF cathode buffer pH 3-10: Arginine (20 mM), lysine (20 mM) IEF anode buffer: phosphoric acid 85% (7 mM) |
| Zymogram | Tris-glycine SDS: Tris base (25 mM), glycine (192 mM), SDS (0.1%), pH 8.3 |
| Find SDS running buffer recipes and native running buffer recipes in gel specific product manuals in the documents tab. |
NuPAGE Bis-Tris SDS buffers & reagents
| Protein sample | Gel chemistry | Sample buffers | Running buffer | Transfer buffer |
|---|---|---|---|---|
| Broad-range MW (6-400 kDa) | Bis-tris | Denaturing: LDS sample buffer Reducing: Sample reducing agent | Small to medium-sized proteins (6–260 kDa): MES SDS buffer Medium to large-size proteins (14–260 kDa): MOPs SDS buffer Reduced samples: Antioxidant | Bis-tris transfer buffer |
Bolt Bis-Tris Plus SDS buffers & reagents
| Protein sample | Gel chemistry | Sample buffers | Running buffer | Transfer buffer |
|---|---|---|---|---|
| Broad-range MW (6-400 kDa) | Bis-tris | Denaturing: LDS sample buffer Reducing: Sample reducing agent | Small to medium-sized proteins (6–260 kDa): MES SDS buffer Medium to large-size proteins (14–260 kDa): MOPs SDS buffer Reduced samples: Antioxidant | Bis-tris transfer buffer |
Why LDS sample buffers
Bolt and NuPAGE LDS Sample Buffers are formulated with Coomassie G250 and phenol red as tracking dyes instead of bromophenol blue. Coomassie (SERVA) G250 gives a sharp dye front with both MES and MOPS SDS running buffers and migrates much closer to the moving ion front than bromophenol blue. Bromophenol blue runs more slowly than some peptides with the MES SDS Running Buffer. This ensures that small peptides do not run off the gel. The concentration of the tracking dye (Coomassie G250) is increased in Bolt and NuPAGE LDS Sample Buffers to enhance viewing of the dye front. LDS is used instead of SDS to allow the formulation of a 4X solution. The 4X solution provides the ability to have higher protein concentrations and less dilution of your protein sample.
Tris-glycine buffers & reagents
| Protein sample | Gel chemistry | Sample buffers | Running buffer | Transfer buffer |
|---|---|---|---|---|
| Broad-range MW (6-400 kDa) | Tris-glycine | Denaturing: Tris-glycine SDS sample buffer Native: Tris-glycine native sample buffer | Denaturing: Tris-glycine SDS buffer Native: Tris-glycine native buffer | Tris-glycine transfer buffer |
Tris-acetate buffers & reagents
| Protein sample | Gel chemistry | Sample buffers | Running buffer | Transfer buffer |
|---|---|---|---|---|
| High MW (40-500 kDa) | Tris-acetate | Denaturing: LDS sample buffer Native: Tris-glycine native sample buffer | Denaturing: Tris-acetate SDS buffer Native: Tris-glycine native buffer | Bis-tris transfer buffer |
Tricine SDS buffers & reagents
| Protein sample | Gel chemistry | Sample buffers | Running buffer | Transfer buffer |
|---|---|---|---|---|
| Low MW (2.5-40 kDa) | Tricine | Denaturing: Tricine SDS sample buffer | Tricine SDS buffer | Tris-glycine transfer buffer |
IEF gel recommended buffers
| Gel chemistry | Sample buffers | Running buffer |
|---|---|---|
| IEF gel |
Zymography gel buffers & reagents
| Gel type | Sample buffer | Running buffer | Gel development buffer | Gel renaturing buffer |
|---|---|---|---|---|
| Zymogram gel | Tris-glycine SDS sample buffer | Tris-glycine SDS running buffer | Zymogram development buffer | Zymogram renaturing buffer |
NativePAGE Bis-Tris gel buffers & reagents
| Gel type | Sample buffer | Running buffer | Gel development buffer |
|---|---|---|---|
| NativePAGE Bis-Tris Gel | Bis-Tris Transfer Buffer |
Gel casting
When pouring your own polyacrylamide gels, choosing high-quality reagents can help you achieve optimal electrophoresis results. SureCast buffers and reagents make it quick and easy to make high quality handcast Tris-glycine gels.
SDS-PAGE gel recipe for pour-your-own Tris-glycine gels
Prepare resolving gel solution according to the volumes in the following table. The volumes provided in the table are for a single gel. Scale volumes proportionally based on the number of gels to be cast.
| Polyacrylamide % | |||||||||
|---|---|---|---|---|---|---|---|---|---|
| Solution | 4% | 6% | 8% | 10% | 12% | 14% | 16% | 18% | 20% |
| SureCast Acrylamide (40%) | 0.8mL | 1.2mL | 1.6mL | 2.0mL | 2.4mL | 2.8mL | 3.3mL | 3.6mL | 4.0mL |
| SureCast Resolving Buffer | 2.0mL | 2.0mL | 2.0mL | 2.0mL | 2.0mL | 2.0mL | 2.0mL | 2.0mL | 2.0mL |
| Distilled water | 5.1mL | 4.7mL | 4.3mL | 3.9mL | 3.5mL | 3.1mL | 2.7mL | 2.3mL | 1.9mL |
| 10% SureCast Ammonium Persulfate (APS) | 80µL | 80µL | 80µL | 80µL | 80µL | 80µL | 80µL | 80µL | 80µL |
| SureCast TEMED** | 8µL | 8µL | 8µL | 8µL | 8µL | 8µL | 8µL | 8µL | 8µL |
**Add this last and mix well just before the gel is to be poured
Prepare stacking gel solution according to the following table. The volumes provided in the table are for a single gel. Scale volumes proportionally based on the number of gels to be cast. Note: Solutions do not require degassing.
| Solution | 4% |
|---|---|
| SureCast Acrylamide (40%) | 0.30 mL |
| SureCast Stacking Buffer | 0.75 mL |
| Distilled water | 1.92 mL |
| 10% SureCast Ammonium Persulfate (APS) | 30 µL |
| SureCast TEMED*** | 3 µL |
***Add this last and mix well just before the gel is to be poured
| Learn more about gel casting and find additional SDS-PAGE gel recipes. |
Ordering quick links
NuPAGE Bis-Tris Buffers
Bolt Bis-Tris Plus Buffers
Tris-Glycine Buffers
Tris-Acetate Buffers
Tricine System Buffers
IEF Buffers
Zymogram Buffers
NativePAGE Bis-Tris Buffers
Handcast Buffers & Reagents
Resources
For Research Use Only. Not for use in diagnostic procedures.