Different Buffer preparation for Gel Electrophoresis

By
MN Editors

Acrylamide Stock Solution Preparation

Application: Acrylamide is used along with bisacrylamide to form gels used in SDS-PAGE. Acrylamide forms polyacrylamide polymers, while bisacrylamide crosslinks between polyacrylamide chains.

To prepare 1L of Acrylamide Stock Solution the following components are required;

ComponentAmountConcentration
acrylamide (mw: 71.08 g/mol)389.6 g5.481 M
bisacrylamide (mw: 154.17 g/mol)10.4 g0.0675 M

Acrylamide Stock Solution Preparation Steps

  1. Prepare 800 mL of distilled water in a suitable container.
  2. Add 389.6 g of acrylamide to the solution.
  3. Add 10.4 g of bisacrylamide to the solution.
  4. Add distilled water until volume is 1 L.
  5. Filter the stock solution using Whatman filter paper and store at 4°C. Prepare fresh acrylamide stock solution every few weeks. T denotes the total percentage concentration of the monomers (acrylamide plus N,N’-methylenebisacrylamide) in grams per 100 mL. C denotes the percentage (by weight) of the cross-linker N,N’-methylenebisacrylamide relative to the total amount of monomers.
Word Guess Challenge
Only 1% of users are able to solve this challenge.

Guess the word hidden in this story

Electrode Buffer Preparation

Application: Electrode buffer has been used in studies involving polymer solar cells.

To prepare 1L of Electrode Buffer the following components are required;

ComponentAmountConcentration
Tris base (mw: 121.14 g/mol)11.6 g0.096 M
glycine (mw: 75.07 g/mol)37.5 g0.5 M
SDS (mw: 288.38 g/mol)3.98 g0.0138 M

Electrode Buffer Preparation Steps

  1. Prepare 800 mL of distilled water in a suitable container.
  2. Add 11.6 g of Tris base to the solution.
  3. Add 37.5 g of glycine to the solution.
  4. Add 3.98 g of SDS to the solution.
  5. Add distilled water until volume is 1 L.

Electrophoretic Transfer Buffer (20x) Preparation 

Application: Electrophoretic Transfer Buffers are used in SDS-PAGE to transfer proteins from a gel to a nitrocellulose membrane.

To prepare 1L of Electrophoretic Transfer Buffer (20x) the following components are required;

ComponentAmountConcentration
Trisodium citrate (mw: 258.07 g/mol)35.29 g0.12 M
Sodium phosphate, dibasic (Na2HPO4) (mw: 141.96 g/mol)22.71 g0.16 M

Electrophoretic Transfer Buffer (20x) Preparation Steps

  1. Prepare 800 mL of distilled water in a suitable container.
  2. Add 35.29 g of Trisodium citrate to the solution.
  3. Add 22.71 g of Sodium phosphate, dibasic (Na2HPO4) to the solution.
  4. Add distilled water until volume is 1 L.
  5. Adjust the pH to 8.3, autoclave, and store for up to 1-2 yr at room temperature.

Imidazole-SDS Solution Preparation

 To prepare 1L of Imidazole-SDS Solution the following components are required;

ComponentAmountConcentration
imidazole (mw: 68.08 g/mol)13.6 g0.2 M
SDS (mw: 288.38 g/mol)1 g0.0035 M

Imidazole-SDS Solution Preparation Steps

  1. Prepare 800 mL of distilled water in a suitable container.
  2. Add 13.6 g of imidazole to the solution.
  3. Add 1 g of SDS to the solution.
  4. Add distilled water until volume is 1 L.

SDS-PAGE SDS Running Buffer (10x) Preparation

Application: In SDS-PAGE (sodium dodecyl sulfate-polyacrylamide gel electrophoresis), SDS Running Buffer is used as the electrophoresis buffer during stacking and resolution.

 To prepare 1L of SDS-PAGE SDS Running Buffer (10x) the following components are required;

ComponentAmountConcentration
Tris base (mw: 121.14 g/mol)30.3 g0.2501 M
Glycine (mw: 75.07 g/mol)144.4 g1.924 M
SDS (mw: 288.38 g/mol)10 g0.0347 M

SDS-PAGE SDS Running Buffer (10x) Preparation Steps

  1. Prepare 800 mL of distilled water in a suitable container.
  2. Add 30.3 g of Tris base to the solution.
  3. Add 144.4 g of Glycine to the solution.
  4. Add 10 g of SDS to the solution.
  5. Add distilled water until volume is 1 L.

Sodium Carbonate Transfer Buffer (40x, pH 9.5) Preparation

Application: Sodium Carbonate Transfer Buffer is used to transfer proteins following SDS-PAGE.

To prepare 1L of Sodium Carbonate Transfer Buffer (40x, pH 9.5) the following components are required;

ComponentAmountConcentration
NaHCO3 (mw: 84.01 g/mol)21.1 g0.251 M
Na2CO3 (mw: 105.99 g/mol)18.35 g0.173 M

Sodium Carbonate Transfer Buffer (40x, pH 9.5) Preparation Steps

  1. Prepare 800 mL of distilled water in a suitable container.
  2. Add 21.1 g of NaHCO3 to the solution.
  3. Add 18.35 g of Na2CO3 to the solution.
  4. Adjust the pH to 9.5 with 10 N NaOH.
  5. Add distilled water until volume is 1 L.
  6. Store at room temperature.

Tris-Glycine Transfer Buffer (20x) Preparation 

Application: Tris-Glycine Transfer Buffer is used as a transfer buffer in Western blotting.

To prepare 1L of Tris-Glycine Transfer Buffer (20x) the following components are required;

ComponentAmountConcentration
Tris base (mw: 121.14 g/mol)24.2 g0.2 M
Glycine (mw: 75.07 g/mol)150.1 g2 M

Tris-Glycine Transfer Buffer (20x) Preparation Steps

  1. Prepare 800 mL of distilled water in a suitable container.
  2. Add 24.2 g of Tris base to the solution.
  3. Add 150.1 g of Glycine to the solution.
  4. Add distilled water until volume is 1 L.
  5. pH adjustment is not necessary (it will be ~8.8). Store at room temperature.

Tris/EACA Buffer Preparation

To prepare 1L of Tris/EACA Buffer Preparation the following components are required;

ComponentAmountConcentration
ε-aminocaproic acid (EACA) (mw: 131.17 g/mol)1.31 g0.01 M
Tris base (mw: 121.14 g/mol)10.9 g0.09 M

Tris/EACA Buffer Preparation Steps

  1. Prepare 800 mL of distilled water in a suitable container.
  2. Add 1.31 g of ε-aminocaproic acid (EACA) to the solution.
  3. Add 10.9 g of Tris base to the solution.
  4. Add distilled water until volume is 1 L.

We hope you've enjoyed reading our latest blog article! We're thrilled to see the positive response it's been receiving so far. We understand that sometimes, after going through an interesting piece of content, you might have questions or want to delve deeper into the topic.

To facilitate meaningful discussions and encourage knowledge sharing, we've set up a dedicated QNA Forum page related to this specific article. If you have any questions, comments, or thoughts you'd like to share, we invite you to visit the QNA Forum.

QNA Forum Page

Feel free to ask your questions or participate in ongoing discussions. Our team of experts, as well as fellow readers, will be active on the forum to engage with you and provide insightful answers.Remember, sharing your thoughts not only helps you gain a deeper understanding but also contributes to the community's growth and learning. We look forward to hearing from you and fostering an enriching discussion.Thank you for being a part of our journey!

Leave a Comment