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What is the difference between acrylamide and bisacrylamide?
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Muiz Roslin
What is the difference between acrylamide and bisacrylamide?
Hi Rasdin, Prem is correct, you need both (points for you Prem). Just to help out a little more, we generally use a 30% Bis-Acrylamide solution for our standard SDS-PAGE. so 87.6g Acrylamide + 2.4g Bis-Acrylamide in 300mls final water. Just remember that acrylamide is a cumulative neurotoxin and is very dangerous to inhale the powder. Cheers Ron
Hi Rasdin, Prem is correct, you need both (points for you Prem). Just to help out a little more, we generally use a 30% Bis-Acrylamide solution for our standard SDS-PAGE. so 87.6g Acrylamide + 2.4g Bis-Acrylamide in 300mls final water. Just remember that acrylamide is a cumulative neurotoxin and is very dangerous to inhale the powder. Cheers Ron
A agree with all comments. Both reagents are necessary for your gel. At this moment I work with 2 proteins (25 kDa and 27 kDa), and I use a 30% Acrylamide and 0,8% Bis Acrylamide solution (30 g Acrylamide + 0,8 gramas Bis). I recommend: Filter the solution and always using a fresh solution (3 months are good to use).
A agree with all comments. Both reagents are necessary for your gel. At this moment I work with 2 proteins (25 kDa and 27 kDa), and I use a 30% Acrylamide and 0,8% Bis Acrylamide solution (30 g Acrylamide + 0,8 gramas Bis). I recommend: Filter the solution and always using a fresh solution (3 months are good to use).
Hi Rasdin, Prem is correct, you need both (points for you Prem). Just to help out a little more, we generally use a 30% Bis-Acrylamide solution for our standard SDS-PAGE. so 87.6g Acrylamide + 2.4g Bis-Acrylamide in 300mls final water. Just remember that acrylamide is a cumulative neurotoxin and is very dangerous to inhale the powder. Cheers Ron
Hi Rasdin, Prem is correct, you need both (points for you Prem). Just to help out a little more, we generally use a 30% Bis-Acrylamide solution for our standard SDS-PAGE. so 87.6g Acrylamide + 2.4g Bis-Acrylamide in 300mls final water. Just remember that acrylamide is a cumulative neurotoxin and is very dangerous to inhale the powder. Cheers Ron
Relevant paragraphs from the National Diagnostics article: Control of the pore size of a polyacrylamide gel is accomplished by changing the T and C values. With increasing T, the pore size decreases in a nearly linear relationship. Higher percentage gels (higher T), with smaller pores, are used to separate smaller molecules. The relationship of C to pore size is more complex. Generally, the minimum pore size occurs when C is about 5% (a 19:1 gel). Decreasing C results in a more open pore structure because there are fewer crosslinker molecules. Increasing C beyond 5% also increases the pore size. This appears to be because of nonhomogeneous bundling of strands in the gel. Researchers have settled on C values of 5.0% (19:1 acrylamide/bis) for most forms of denaturing DNA and RNA electrophoresis and 3.3% (29:1) for most native DNA and RNA gels. For SDS-PAGE electrophoresis of proteins, the standard C value that has been adopted is 2.6% (37.5:1)."
Relevant paragraphs from the National Diagnostics article: Control of the pore size of a polyacrylamide gel is accomplished by changing the T and C values. With increasing T, the pore size decreases in a nearly linear relationship. Higher percentage gels (higher T), with smaller pores, are used to separate smaller molecules. The relationship of C to pore size is more complex. Generally, the minimum pore size occurs when C is about 5% (a 19:1 gel). Decreasing C results in a more open pore structure because there are fewer crosslinker molecules. Increasing C beyond 5% also increases the pore size. This appears to be because of nonhomogeneous bundling of strands in the gel. Researchers have settled on C values of 5.0% (19:1 acrylamide/bis) for most forms of denaturing DNA and RNA electrophoresis and 3.3% (29:1) for most native DNA and RNA gels. For SDS-PAGE electrophoresis of proteins, the standard C value that has been adopted is 2.6% (37.5:1)."
Here is an excellent article from National Diagnostics on this topic: https://nationaldiagnostics.com/electrophoresis/article/polyacrylamide-matrix "For discussions of the composition of polyacrylamide gels, a standard nomenclature has been widely adopted. In this nomenclature, T represents the total percentage concentration (w/v) of monomer (acrylamide plus crosslinker) in the gel. The term C refers to the percentage of the total monomer represented by the crosslinker. For example, an 8%, 19:1 (acrylamide/bisacrylamide) gel would have a T value of 8% and a C value of 5%."
Here is an excellent article from National Diagnostics on this topic: https://nationaldiagnostics.com/electrophoresis/article/polyacrylamide-matrix "For discussions of the composition of polyacrylamide gels, a standard nomenclature has been widely adopted. In this nomenclature, T represents the total percentage concentration (w/v) of monomer (acrylamide plus crosslinker) in the gel. The term C refers to the percentage of the total monomer represented by the crosslinker. For example, an 8%, 19:1 (acrylamide/bisacrylamide) gel would have a T value of 8% and a C value of 5%."
Acrylamide and Bisacrylamide, both are very important for proper polymerization. Acrylamide forms linear polymers whereas Bisacrylamide cross links these linear polymer. More numbers of cross links means smaller pore size. So, ratio of Acrylamide and Bisacrylamide determines pore size.
Acrylamide and Bisacrylamide, both are very important for proper polymerization. Acrylamide forms linear polymers whereas Bisacrylamide cross links these linear polymer. More numbers of cross links means smaller pore size. So, ratio of Acrylamide and Bisacrylamide determines pore size.
Acrylamide and Bisacrylamide, both are very important for proper polymerization. Acrylamide forms linear polymers whereas Bisacrylamide cross links these linear polymer. More numbers of cross links means smaller pore size. So, ratio of Acrylamide and Bisacrylamide determines pore size.
Acrylamide and Bisacrylamide, both are very important for proper polymerization. Acrylamide forms linear polymers whereas Bisacrylamide cross links these linear polymer. More numbers of cross links means smaller pore size. So, ratio of Acrylamide and Bisacrylamide determines pore size.
I'm assuming that you will also be analyzing a loading control protein within the size range of your two proteins of interest. Bax should be about 20 kDa, am I correct? You can try starting with a 12.5%, 37.5:1 (acrylamide/bisacrylamide) gel in that case to get good separation in that molecular weight range (20-50 kDa). I hope that helps.
I'm assuming that you will also be analyzing a loading control protein within the size range of your two proteins of interest. Bax should be about 20 kDa, am I correct? You can try starting with a 12.5%, 37.5:1 (acrylamide/bisacrylamide) gel in that case to get good separation in that molecular weight range (20-50 kDa). I hope that helps.
Hi Rasdin,
Prem is correct, you need both (points for you Prem). Just to help out a little more, we generally use a 30% Bis-Acrylamide solution for our standard SDS-PAGE. so 87.6g Acrylamide + 2.4g Bis-Acrylamide in 300mls final water. Just remember that acrylamide is a cumulative neurotoxin and is very dangerous to inhale the powder.
Cheers
Ron
Hi Rasdin,
Prem is correct, you need both (points for you Prem). Just to help out a little more, we generally use a 30% Bis-Acrylamide solution for our standard SDS-PAGE. so 87.6g Acrylamide + 2.4g Bis-Acrylamide in 300mls final water. Just remember that acrylamide is a cumulative neurotoxin and is very dangerous to inhale the powder.
Cheers
Ron
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Tq Prem for the answer and Ron for the explanation...really appreciated that.
Mark, i intend to analyzes p53 and bax protein...
Tq Prem for the answer and Ron for the explanation...really appreciated that.
Mark, i intend to analyzes p53 and bax protein...
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A agree with all comments.
Both reagents are necessary for your gel.
At this moment I work with 2 proteins (25 kDa and 27 kDa), and I use a 30% Acrylamide and 0,8% Bis Acrylamide solution (30 g Acrylamide + 0,8 gramas Bis).
I recommend: Filter the solution and always using a fresh solution (3 months are good to use).
Check this article:
https://nationaldiagnostics.com/electrophoresis/article/polyacrylamide-matrix
I use this protocol in my lab.
A agree with all comments.
Both reagents are necessary for your gel.
At this moment I work with 2 proteins (25 kDa and 27 kDa), and I use a 30% Acrylamide and 0,8% Bis Acrylamide solution (30 g Acrylamide + 0,8 gramas Bis).
I recommend: Filter the solution and always using a fresh solution (3 months are good to use).
Check this article:
https://nationaldiagnostics.com/electrophoresis/article/polyacrylamide-matrix
I use this protocol in my lab.
More
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By the way, acrylamide is very toxic.
By the way, acrylamide is very toxic.
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Hi Rasdin,
Prem is correct, you need both (points for you Prem). Just to help out a little more, we generally use a 30% Bis-Acrylamide solution for our standard SDS-PAGE. so 87.6g Acrylamide + 2.4g Bis-Acrylamide in 300mls final water. Just remember that acrylamide is a cumulative neurotoxin and is very dangerous to inhale the powder.
Cheers
Ron
Hi Rasdin,
Prem is correct, you need both (points for you Prem). Just to help out a little more, we generally use a 30% Bis-Acrylamide solution for our standard SDS-PAGE. so 87.6g Acrylamide + 2.4g Bis-Acrylamide in 300mls final water. Just remember that acrylamide is a cumulative neurotoxin and is very dangerous to inhale the powder.
Cheers
Ron
More
VOTE
Relevant paragraphs from the National Diagnostics article:
Control of the pore size of a polyacrylamide gel is accomplished by changing the T and C values. With increasing T, the pore size decreases in a nearly linear relationship. Higher percentage gels (higher T), with smaller pores, are used to separate smaller molecules. The relationship of C to pore size is more complex. Generally, the minimum pore size occurs when C is about 5% (a 19:1 gel). Decreasing C results in a more open pore structure because there are fewer crosslinker molecules. Increasing C beyond 5% also increases the pore size. This appears to be because of nonhomogeneous bundling of strands in the gel.
Researchers have settled on C values of 5.0% (19:1 acrylamide/bis) for most forms of denaturing DNA and RNA electrophoresis and 3.3% (29:1) for most native DNA and RNA gels. For SDS-PAGE electrophoresis of proteins, the standard C value that has been adopted is 2.6% (37.5:1)."
Relevant paragraphs from the National Diagnostics article:
Control of the pore size of a polyacrylamide gel is accomplished by changing the T and C values. With increasing T, the pore size decreases in a nearly linear relationship. Higher percentage gels (higher T), with smaller pores, are used to separate smaller molecules. The relationship of C to pore size is more complex. Generally, the minimum pore size occurs when C is about 5% (a 19:1 gel). Decreasing C results in a more open pore structure because there are fewer crosslinker molecules. Increasing C beyond 5% also increases the pore size. This appears to be because of nonhomogeneous bundling of strands in the gel.
Researchers have settled on C values of 5.0% (19:1 acrylamide/bis) for most forms of denaturing DNA and RNA electrophoresis and 3.3% (29:1) for most native DNA and RNA gels. For SDS-PAGE electrophoresis of proteins, the standard C value that has been adopted is 2.6% (37.5:1)."
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Rasdin, do you intend to analyze proteins or nucleic acids on your polyacrylamide gel?
Rasdin, do you intend to analyze proteins or nucleic acids on your polyacrylamide gel?
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Here is an excellent article from National Diagnostics on this topic: https://nationaldiagnostics.com/electrophoresis/article/polyacrylamide-matrix
"For discussions of the composition of polyacrylamide gels, a standard nomenclature has been widely adopted. In this nomenclature, T represents the total percentage concentration (w/v) of monomer (acrylamide plus crosslinker) in the gel. The term C refers to the percentage of the total monomer represented by the crosslinker. For example, an 8%, 19:1 (acrylamide/bisacrylamide) gel would have a T value of 8% and a C value of 5%."
Here is an excellent article from National Diagnostics on this topic: https://nationaldiagnostics.com/electrophoresis/article/polyacrylamide-matrix
"For discussions of the composition of polyacrylamide gels, a standard nomenclature has been widely adopted. In this nomenclature, T represents the total percentage concentration (w/v) of monomer (acrylamide plus crosslinker) in the gel. The term C refers to the percentage of the total monomer represented by the crosslinker. For example, an 8%, 19:1 (acrylamide/bisacrylamide) gel would have a T value of 8% and a C value of 5%."
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Acrylamide and Bisacrylamide, both are very important for proper polymerization. Acrylamide forms linear polymers whereas Bisacrylamide cross links these linear polymer. More numbers of cross links means smaller pore size. So, ratio of Acrylamide and Bisacrylamide determines pore size.
Acrylamide and Bisacrylamide, both are very important for proper polymerization. Acrylamide forms linear polymers whereas Bisacrylamide cross links these linear polymer. More numbers of cross links means smaller pore size. So, ratio of Acrylamide and Bisacrylamide determines pore size.
More
VOTE
Acrylamide and Bisacrylamide, both are very important for proper polymerization. Acrylamide forms linear polymers whereas Bisacrylamide cross links these linear polymer. More numbers of cross links means smaller pore size. So, ratio of Acrylamide and Bisacrylamide determines pore size.
Acrylamide and Bisacrylamide, both are very important for proper polymerization. Acrylamide forms linear polymers whereas Bisacrylamide cross links these linear polymer. More numbers of cross links means smaller pore size. So, ratio of Acrylamide and Bisacrylamide determines pore size.
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I'm assuming that you will also be analyzing a loading control protein within the size range of your two proteins of interest. Bax should be about 20 kDa, am I correct? You can try starting with a 12.5%, 37.5:1 (acrylamide/bisacrylamide) gel in that case to get good separation in that molecular weight range (20-50 kDa). I hope that helps.
I'm assuming that you will also be analyzing a loading control protein within the size range of your two proteins of interest. Bax should be about 20 kDa, am I correct? You can try starting with a 12.5%, 37.5:1 (acrylamide/bisacrylamide) gel in that case to get good separation in that molecular weight range (20-50 kDa). I hope that helps.
More
VOTE