The Role of Follicle-Stimulating Hormone Receptor in the Treatment of Ovarian Cancer.Targeting FLT3 to Treat Acute Myeloid Leukemia.MiRNAs: the Next Generation Drugs in Human Cancer.Converting Adherent Cells to Suspension Cells (Serum Dilution).Recombinant Lentivirus Production Protocol.Generation of Stable Cell Lines Using Lentivirus.Generation and Amplification of Recombinant Adenovirus.Determining Antibiotic Killing Curve for Stable Cell Line Generation.Adenovirus Construction by Homologous Recombination in Bacteria.Synthetic mRNA – Emerging New Class of Gene-based Drugs.CoV-S Pseudotyped Lentivirus - Efficient Tool for Anti-CoV Antibody Screening.STAT Reporter Cell Line - Tool for Screening of Drugs Targeting JAK/STAT Pathway.BaF3-BCR-ABL & BaF3-BCR-ABL-T315I Stable Cell Lines - In Vitro Screening Tool for CML Drugs.CRISPR/Cas9: From Genome Engineering to Drug Discovery and Therapy.Adenovirus: The Effective In Vivo Gene Delivery Vector.Oncolytic Virotherapy - Sharpening the Sword for Improved Cancer Treatment Strategies.LAM PCR - Approach for Tracking Lentiviral Integration Sites.
RNA Interference Technology-Development and Applications.Lentiviral Vectors - the Application for CAR-T Therapies.Knockout Cell Lines in Antibody Screening and Validation.Baculovirus-Efficient Tool for Protein Expression.Genome Editing - Generation of Reporter Stem Cells.Phagemid Vector - Efficient Tool for Phage Display.Robust Tn5 Transposase - Efficient Tool for Sequencing.Antibody-Producing Cell Lines Development.Endotoxin Detection of Pharmaceutical and Medical Devices.Baculovirus Expression Vector Systems Guide.The concentration occurs due to the difference in the rate of migration of glycine ion, chloride ion, and proteins, as illustrated below. Proteins are highly concentrated when they migrate through a stacking gel prior to entering a separating gel. Pour acrylamide solution for a stacking gel insert a comb and allow the acrylamide to polymerize. Gels with an acrylamide concentration gradient (gradient gels) are also used. In general, an acrylamide concentration between 6 and 15% is used. Using a higher acrylamide concentration produces a gel with a smaller mesh size suitable for the separation of small proteins. Use an appropriate gel concentration for your target protein. Proteins migrate at different rate depending on the concentration of the separating gel. Allow acrylamide to polymerize for 20-30 minutes to form a gel. Overlay with water to prevent contact with air (oxygen), which inhibits polymerization. Pour acrylamide solution for a separating gel. Thoroughly clean the glass plates with ethanol, and assemble the gel casting mold. Use an appropriate comb depending on the sample size.Įxample: Use an 8-lane comb for 7 samples and molecular weight markers. Gather combs, glass plates, spacer (silicone tubing), and binder clips.Ī comb is used to make wells (lanes) to load samples. Preparation of polyacrylamide gel ※An example performed at MBL Polyacrylamide gel electrophoresis of SDS-treated proteins allows researchers to separate proteins based on their length in an easy, inexpensive, and relatively accurate manner. The strength of the gel allows easy handling. Polymerized acrylamide (polyacrylamide) forms a mesh-like matrix suitable for the separation of proteins of typical size. In the presence of SDS and a reducing agent that cleaves disulfide bonds critical for proper folding, proteins unfold into linear chains with negative charge proportional to the polypeptide chain length. SDS is a detergent with a strong protein-denaturing effect and binds to the protein backbone at a constant molar ratio. In SDS-PAGE, the use of sodium dodecyl sulfate (SDS, also known as sodium lauryl sulfate) and polyacrylamide gel largely eliminates the influence of the structure and charge, and proteins are separated solely based on polypeptide chain length. Other influences on the rate of migration through the gel matrix include the structure and charge of the proteins.
When proteins are separated by electrophoresis through a gel matrix, smaller proteins migrate faster due to less resistance from the gel matrix.