Protein Gel and Membrane Stains

Whether you just need a quick visual confirmation or require a highly-sensitive stain to detect low-abundance proteins, we offer a variety of easy-to-use, effective protein stains for both in-gel and on-membrane detection. Check out our comprehensive collection of stains including Coomassie dye-based, silver, fluorescent, tagged fusion protein stains and PTM protein gel stains.

Polyacrylamide gel stains

 Coomassie stainingSilver stainingFluorescent protein staining
Sensitivity5-25 ng0.25-0.5 ng0.25-0.5 ng
Mode of actionCoomassie dye binds to basic and hydrophobic residues of proteins through non-covalent interactions, changing from dull reddish-brown to intense blue.Silver ions interact and bind with carboxylic acid groups (Asp and Glu), imidazole (His), sulfhydryls (Cys), and amines (Lys). Silver ions are reduced to metallic silver, resulting in brown-black color.Most fluorescent stains bind non-covalently to proteins either through the polypeptide backbone or through interaction with the SDS coat around the proteins
Typical protocol time10-135 min30-120 min60 min
DetectionVisualVisualUV or blue/green-light transilluminators or imaging instruments with appropriate filters
Compatibility with downstream applicationsMS and sequencing compatible, western blotting (only non-fixative methods)Certain formulations are MS compatibleMost stains are MS compatible, western blotting
AdvantagesQuick and simple staining protocols, inexpensiveLowest detection limits not requiring specialized equipmentBroad linear dynamic range with low detection limits
 Learn moreLearn moreLearn more

Coomassie dye stains

Coomassie dyes (also referred to as Coomassie blue or Coomassie brilliant blue) are common dyes used for the visualization of proteins separated by protein gel electrophoresis. Coomassie based stains offer easy staining protocols and provides good quantitative linearity with good sensitivity. Coomassie based dyes do not permanently chemically modify the target proteins rather interact via non-covalent interactions. Because no chemical modification occurs, excised protein bands can be completely de-stained and the recovered proteins can be used for analysis by mass spectrometry or sequencing.

In acidic conditions, Coomassie binds to basic and hydrophobic residues of proteins, changing in color from a dull reddish-brown to intense blue. As with all staining methods, Coomassie staining detects some proteins better than others, based on the chemistry of action and differences in protein composition. Thus, Coomassie staining can detect as little as 8–10 ng per band for some proteins and 25 ng per band for most proteins.

Two forms of the dye exists- R-250 (R standing for the reddish tint) and G-250 (G standing for the greenish tint). Both variants offer the same relative sensitivity and easy detection. Coomassie G-250 (also known as colloidal Coomassie dye) offers a faster staining protocols and eliminates the need for de-staining.

Silver dye stains

Silver staining is the most sensitive colorimetric method for detecting total protein. The technique involves the deposition of metallic silver onto the surface of a gel at the locations of protein bands. Silver ions (from silver nitrate in the staining reagent) interact and bind with certain protein functional groups. The strongest interactions occur with carboxylic acid groups (Asp and Glu), imidazole (His), sulfhydryls (Cys), and amines (Lys). Various sensitizer and enhancer reagents are essential for controlling the specificity and efficiency of silver ion binding to proteins and effective conversion (development) of the bound silver to metallic silver. The development process is essentially the same as for photographic film: silver ions are reduced to metallic silver, resulting in a brown-black color.

Silver staining protocols require several steps, which are affected by reagent quality as well as incubation times and thickness of the gel. An advantage of commercially available silver staining kits is that the formulations and protocols are optimized and consistently manufactured, helping to maximize consistency of results from experiment to experiment. Kits with optimized protocols are robust and easy to use, detecting less than 0.5 ng of protein in typical gels.

Silver stains use either glutaraldehyde or formaldehyde as the enhancer. These reagents can cause chemical crosslinking of the proteins in the gel matrix, limiting compatibility with de-staining and elution methods for analysis by mass spectrometry (MS). Therefore, optimization of sensitivity vs. protein recoverability is critical when employing silver staining as part of an MS workflow.

Silver stain formulations can be made such that protein bands stain black, blue-brown, red, or yellow, depending on their charge and other characteristics. This is particularly useful for differentiating overlapping spots on 2D gels.

Fluorescent dye stains

Fluorescent stains offer high sensitivity combined with simple staining protocols. Majority of fluorescent stains offer sensitivities similar to that obtained with silver staining techniques and can be used as a good alternative to silver staining in 1D or 2D electrophoresis. Unlike silver stains, fluorescent stains require equipment for visualization. Typical stains can be viewed with a standard UV or blue/green-light trans illuminator or with imaging equipment equipped with the appropriate filters. Fluorescent protein gel stains offer linear quantitation ranges, with some stains reporting over three orders of magnitude.

Nitrocellulose and PVDF membrane stains

 Pierce Reversible Protein Stain KitPonceau S Staining SolutionSYPRO Ruby Blot StainNo-Stain Protein Labeling Reagent
DetectionColorimetric
(blue)
Colorimetric
(pink-reddish)
Fluorescence
(Ex: 280 & 450 nm, Em: 618 nm)
Fluorescence
(Ex: 488 nm, Em: 590 nm)
Min. protein detected~25-50 ng~250 ng~2-8 ng~20 ng
Staining time~15 min~5 min~60 min~10 min
ApplicationsMonitoring transfer efficiency, total protein normalization after western blotting
Destaining before immunodetectionDestaining requiredDestaining requiredNot requiredNot required
Considerations Not recommended for fluorescent western blot detectionPVDF staining fixative required 
Cat. No.24580 (nitrocellulose)
24585 (PVDF)
A40000279 (500 mL)S11791 (200 mL)A44449
(40 reactions)
A44717
(10 reactions)

Total protein stains

Colorimetric stains

ProductLimit of detectionComponents (steps)*Time required**Mass spec. compatibleStaining agent
GelCode Blue Safe Protein Stain9 ng2 (5)2 hr 15 minYesG-250
GelCode Blue Stain Reagent8 ng1 (5)2 hr 15 minYesG-250
Imperial Protein Stain3 ng1 (5)2 hr 15 minYesR-250
Simply Blue Safe Stain>7 ng1 (5)12 min microwave
3 hr standard
YesG-250
PageBlue Protein Stain5 ng1 (5)30 min microwave
90 min standard
YesG-250
Colloidal Blue Staining< 10 ng1 (3)10 hr G-250
Pierce Silver Stain for Mass Spec0.25 ng6 (17)1 hr 13 minYesMetallic silver ions
Pierce Silver Stain Kit0.25 ng4 (15)2 hr 25 min Metallic silver ions
Pierce Color Silver Stain Kit0.1 ng4 (9)4 hr 5 minYes 
Pierce Zinc Reversible Stain Kit0.25 ng3 (4)14 minYes 
SilverXpress Silver Staining Kit0.86 ng5 (13)2 hr Metallic silver ions
SilverQuest Silver Staining Kit0.3 ng7 (8)1 hr 30 minYesMetallic silver ions
Pierce Reversible Protein Stain Kit for Nitrocellulose membranes25-50 ng3 (5)15 min  
Pierce Reversible Protein Stain Kit for PVDF Membranes25-50 ng4 (8)15 min  
Ponceau S Staining Solution~250 ng1 (3)15 min Ponceau S

Fluorescent stains

ProductLimit of detectionComponents (steps)*Time required**Mass spec. compatibleExcitation and emission maxima
SYPRO Ruby0.25- 1 ng1 (3)90 min microwave
18 hr standard
YesExcitation: 280, 450nm
Emission: 610 nm
SYPRO Orange4-8 ng1 (5)~1 hr Excitation: 300, 470nm Emission: 570 nm
SYPRO Red4-8 ng1 (5)~1 hr Excitation: 300, 550nm Emission: 630nm
SYPRO Tangerine1-4 ng1 (4)~1 hrYesExcitation: 300, 490nm Emission: 640nm
SYPRO Ruby blot stain2-8 ng1 (4)~1 hrYesExcitation: 280, 450nm Emission: 618nm
Coomassie Fluor Orange8 ng1 (4)~1 hr Excitation: 300, 470nm Emission: 570nm
Krypton Fluorescent Protein Stain0.25 ng1 (7)2 hr 40 minYesExcitation: 520nm
Emission: 580nm

*Some products require additional reagents to be supplied by user. The number of steps includes gel washes, solution changes, destaining steps, etc.
**Times include gel wash steps and all steps required for best sensitivity in a mini-gel. When less sensitivity is required, shorter staining and destaining times may be used with some protocols.

Functional group protein stains

ProductFunctional groupLimit of detectionComponents (steps)*Time required**Detection methodMass spec. compatible
Pro-Q Emerald 488 Glycoprotein Gel and Blot Stain KitPeriodate-oxidized carbohydrate groups4 ng glycoprotein / band2 (13)~6 hrsfluorescentYes
Pro-Q Emerald 300 Glycoprotein Gel and Blot Stain KitPeriodate-oxidized carbohydrate groups0.5 ng glycoprotein / band2 (10)~5 hrsUVYes
Pierce Glycoprotein StainSialic acid and other oxidizable carbohydrate groups160 ng5 (11)2 hr 40 mincolorimetricNot tested
Pro-Q Diamond Phosphoprotein Gel StainPhosphate groups attached to tyrosine, serine, or threonine residues1–16 ng phosphoprotein / band1 (5)~4-5 hrsfluorescentNot tested
Pro-Q Diamond Phosphoprotein Blot Stain KitPhosphate groups attached to tyrosine, serine, or threonine residues8-16 ng1 (6)~75 minfluorescentYes
Lumio Green Detection KitLumio, TC-tag1 pmole of a Lumio fusion protein3 (8)ImmediateUV, fluorescentNot tested
InVision™ His-tag In-gel Stain6X His-tag~0.5 picomole of a 6X His-tagged fusion protein2 (4)<3 hrsUV, fluorescentNot tested
6xHis Protein Tag Stain Kit6X His-tag200 ng4 (9)2 hr 20 minUV, fluorescentNot tested

*Some products require additional reagents to be supplied by user. The number of steps includes multiple gel washes, solution changes, destaining steps, etc.
**Times include gel wash steps and all steps required for best sensitivity in a mini-gel. When less sensitivity is required, shorter staining and destaining times may be used with some protocols.

Polyacrylamide gel stains

 Coomassie stainingSilver stainingFluorescent protein staining
Sensitivity5-25 ng0.25-0.5 ng0.25-0.5 ng
Mode of actionCoomassie dye binds to basic and hydrophobic residues of proteins through non-covalent interactions, changing from dull reddish-brown to intense blue.Silver ions interact and bind with carboxylic acid groups (Asp and Glu), imidazole (His), sulfhydryls (Cys), and amines (Lys). Silver ions are reduced to metallic silver, resulting in brown-black color.Most fluorescent stains bind non-covalently to proteins either through the polypeptide backbone or through interaction with the SDS coat around the proteins
Typical protocol time10-135 min30-120 min60 min
DetectionVisualVisualUV or blue/green-light transilluminators or imaging instruments with appropriate filters
Compatibility with downstream applicationsMS and sequencing compatible, western blotting (only non-fixative methods)Certain formulations are MS compatibleMost stains are MS compatible, western blotting
AdvantagesQuick and simple staining protocols, inexpensiveLowest detection limits not requiring specialized equipmentBroad linear dynamic range with low detection limits
 Learn moreLearn moreLearn more

Coomassie dye stains

Coomassie dyes (also referred to as Coomassie blue or Coomassie brilliant blue) are common dyes used for the visualization of proteins separated by protein gel electrophoresis. Coomassie based stains offer easy staining protocols and provides good quantitative linearity with good sensitivity. Coomassie based dyes do not permanently chemically modify the target proteins rather interact via non-covalent interactions. Because no chemical modification occurs, excised protein bands can be completely de-stained and the recovered proteins can be used for analysis by mass spectrometry or sequencing.

In acidic conditions, Coomassie binds to basic and hydrophobic residues of proteins, changing in color from a dull reddish-brown to intense blue. As with all staining methods, Coomassie staining detects some proteins better than others, based on the chemistry of action and differences in protein composition. Thus, Coomassie staining can detect as little as 8–10 ng per band for some proteins and 25 ng per band for most proteins.

Two forms of the dye exists- R-250 (R standing for the reddish tint) and G-250 (G standing for the greenish tint). Both variants offer the same relative sensitivity and easy detection. Coomassie G-250 (also known as colloidal Coomassie dye) offers a faster staining protocols and eliminates the need for de-staining.

Silver dye stains

Silver staining is the most sensitive colorimetric method for detecting total protein. The technique involves the deposition of metallic silver onto the surface of a gel at the locations of protein bands. Silver ions (from silver nitrate in the staining reagent) interact and bind with certain protein functional groups. The strongest interactions occur with carboxylic acid groups (Asp and Glu), imidazole (His), sulfhydryls (Cys), and amines (Lys). Various sensitizer and enhancer reagents are essential for controlling the specificity and efficiency of silver ion binding to proteins and effective conversion (development) of the bound silver to metallic silver. The development process is essentially the same as for photographic film: silver ions are reduced to metallic silver, resulting in a brown-black color.

Silver staining protocols require several steps, which are affected by reagent quality as well as incubation times and thickness of the gel. An advantage of commercially available silver staining kits is that the formulations and protocols are optimized and consistently manufactured, helping to maximize consistency of results from experiment to experiment. Kits with optimized protocols are robust and easy to use, detecting less than 0.5 ng of protein in typical gels.

Silver stains use either glutaraldehyde or formaldehyde as the enhancer. These reagents can cause chemical crosslinking of the proteins in the gel matrix, limiting compatibility with de-staining and elution methods for analysis by mass spectrometry (MS). Therefore, optimization of sensitivity vs. protein recoverability is critical when employing silver staining as part of an MS workflow.

Silver stain formulations can be made such that protein bands stain black, blue-brown, red, or yellow, depending on their charge and other characteristics. This is particularly useful for differentiating overlapping spots on 2D gels.

Fluorescent dye stains

Fluorescent stains offer high sensitivity combined with simple staining protocols. Majority of fluorescent stains offer sensitivities similar to that obtained with silver staining techniques and can be used as a good alternative to silver staining in 1D or 2D electrophoresis. Unlike silver stains, fluorescent stains require equipment for visualization. Typical stains can be viewed with a standard UV or blue/green-light trans illuminator or with imaging equipment equipped with the appropriate filters. Fluorescent protein gel stains offer linear quantitation ranges, with some stains reporting over three orders of magnitude.

Nitrocellulose and PVDF membrane stains

 Pierce Reversible Protein Stain KitPonceau S Staining SolutionSYPRO Ruby Blot StainNo-Stain Protein Labeling Reagent
DetectionColorimetric
(blue)
Colorimetric
(pink-reddish)
Fluorescence
(Ex: 280 & 450 nm, Em: 618 nm)
Fluorescence
(Ex: 488 nm, Em: 590 nm)
Min. protein detected~25-50 ng~250 ng~2-8 ng~20 ng
Staining time~15 min~5 min~60 min~10 min
ApplicationsMonitoring transfer efficiency, total protein normalization after western blotting
Destaining before immunodetectionDestaining requiredDestaining requiredNot requiredNot required
Considerations Not recommended for fluorescent western blot detectionPVDF staining fixative required 
Cat. No.24580 (nitrocellulose)
24585 (PVDF)
A40000279 (500 mL)S11791 (200 mL)A44449
(40 reactions)
A44717
(10 reactions)

Total protein stains

Colorimetric stains

ProductLimit of detectionComponents (steps)*Time required**Mass spec. compatibleStaining agent
GelCode Blue Safe Protein Stain9 ng2 (5)2 hr 15 minYesG-250
GelCode Blue Stain Reagent8 ng1 (5)2 hr 15 minYesG-250
Imperial Protein Stain3 ng1 (5)2 hr 15 minYesR-250
Simply Blue Safe Stain>7 ng1 (5)12 min microwave
3 hr standard
YesG-250
PageBlue Protein Stain5 ng1 (5)30 min microwave
90 min standard
YesG-250
Colloidal Blue Staining< 10 ng1 (3)10 hr G-250
Pierce Silver Stain for Mass Spec0.25 ng6 (17)1 hr 13 minYesMetallic silver ions
Pierce Silver Stain Kit0.25 ng4 (15)2 hr 25 min Metallic silver ions
Pierce Color Silver Stain Kit0.1 ng4 (9)4 hr 5 minYes 
Pierce Zinc Reversible Stain Kit0.25 ng3 (4)14 minYes 
SilverXpress Silver Staining Kit0.86 ng5 (13)2 hr Metallic silver ions
SilverQuest Silver Staining Kit0.3 ng7 (8)1 hr 30 minYesMetallic silver ions
Pierce Reversible Protein Stain Kit for Nitrocellulose membranes25-50 ng3 (5)15 min  
Pierce Reversible Protein Stain Kit for PVDF Membranes25-50 ng4 (8)15 min  
Ponceau S Staining Solution~250 ng1 (3)15 min Ponceau S

Fluorescent stains

ProductLimit of detectionComponents (steps)*Time required**Mass spec. compatibleExcitation and emission maxima
SYPRO Ruby0.25- 1 ng1 (3)90 min microwave
18 hr standard
YesExcitation: 280, 450nm
Emission: 610 nm
SYPRO Orange4-8 ng1 (5)~1 hr Excitation: 300, 470nm Emission: 570 nm
SYPRO Red4-8 ng1 (5)~1 hr Excitation: 300, 550nm Emission: 630nm
SYPRO Tangerine1-4 ng1 (4)~1 hrYesExcitation: 300, 490nm Emission: 640nm
SYPRO Ruby blot stain2-8 ng1 (4)~1 hrYesExcitation: 280, 450nm Emission: 618nm
Coomassie Fluor Orange8 ng1 (4)~1 hr Excitation: 300, 470nm Emission: 570nm
Krypton Fluorescent Protein Stain0.25 ng1 (7)2 hr 40 minYesExcitation: 520nm
Emission: 580nm

*Some products require additional reagents to be supplied by user. The number of steps includes gel washes, solution changes, destaining steps, etc.
**Times include gel wash steps and all steps required for best sensitivity in a mini-gel. When less sensitivity is required, shorter staining and destaining times may be used with some protocols.

Functional group protein stains

ProductFunctional groupLimit of detectionComponents (steps)*Time required**Detection methodMass spec. compatible
Pro-Q Emerald 488 Glycoprotein Gel and Blot Stain KitPeriodate-oxidized carbohydrate groups4 ng glycoprotein / band2 (13)~6 hrsfluorescentYes
Pro-Q Emerald 300 Glycoprotein Gel and Blot Stain KitPeriodate-oxidized carbohydrate groups0.5 ng glycoprotein / band2 (10)~5 hrsUVYes
Pierce Glycoprotein StainSialic acid and other oxidizable carbohydrate groups160 ng5 (11)2 hr 40 mincolorimetricNot tested
Pro-Q Diamond Phosphoprotein Gel StainPhosphate groups attached to tyrosine, serine, or threonine residues1–16 ng phosphoprotein / band1 (5)~4-5 hrsfluorescentNot tested
Pro-Q Diamond Phosphoprotein Blot Stain KitPhosphate groups attached to tyrosine, serine, or threonine residues8-16 ng1 (6)~75 minfluorescentYes
Lumio Green Detection KitLumio, TC-tag1 pmole of a Lumio fusion protein3 (8)ImmediateUV, fluorescentNot tested
InVision™ His-tag In-gel Stain6X His-tag~0.5 picomole of a 6X His-tagged fusion protein2 (4)<3 hrsUV, fluorescentNot tested
6xHis Protein Tag Stain Kit6X His-tag200 ng4 (9)2 hr 20 minUV, fluorescentNot tested

*Some products require additional reagents to be supplied by user. The number of steps includes multiple gel washes, solution changes, destaining steps, etc.
**Times include gel wash steps and all steps required for best sensitivity in a mini-gel. When less sensitivity is required, shorter staining and destaining times may be used with some protocols.

Resources