Which method is used to separate proteins on the basis of their sizes?
- lon exchange chromatography
- Thin layer chromatography
- Adsorption chromatography
- Gel filtration chromatography
Chromatography is a laboratory technique that separates a mixture of compounds based on their physical and chemical properties. There are several different chromatography techniques, including:
- Gas Chromatography (GC): This technique separates volatile compounds based on their volatility, or their ability to evaporate at room temperature. GC uses a gas as the mobile phase, and a stationary phase, typically a solid or liquid, to separate the compounds.
- High Performance Liquid Chromatography (HPLC): This technique separates compounds based on their interaction with a stationary phase, typically a solid or liquid, and a mobile phase, typically a liquid. HPLC is used to separate compounds in complex mixtures, such as biological samples.
- Thin Layer Chromatography (TLC): This technique separates compounds based on their interaction with a stationary phase, typically a thin layer of solid material on a glass or plastic plate, and a mobile phase, typically a liquid solvent. TLC is used to identify and quantify compounds in a mixture. This technique is mostly used for non-volatile or low-volatility compounds.
- Capillary Electrophoresis (CE): This technique separates compounds based on their interaction with a stationary phase, typically a capillary tube filled with a solid or liquid, and a mobile phase, typically an electrolyte solution. CE uses an electric field to separate the compounds.
- Affinity Chromatography: This technique separates compounds based on their specific interaction with a particular molecule, called a ligand, attached to the stationary phase. Affinity chromatography is often used to purify proteins and other biomolecules.
lon exchange chromatography
Lon Exchange Chromatography is a type of chromatography that is used to purify or separate proteins or other biomolecules based on their charge.
It is based on the principle of charge-based separation, where the molecules are separated based on their interactions with a charged stationary phase. The stationary phase in anion exchange chromatography is a matrix that carries a negative charge, and the molecules to be separated are passed through it in a liquid solution.
The molecules will bind to the stationary phase based on their charge, with positively charged molecules binding more strongly to the negatively charged stationary phase. The purified molecules can then be eluted, or removed, from the stationary phase by changing the pH of the solution or by using a higher salt concentration.
Anion exchange chromatography is commonly used in biochemistry and molecular biology laboratories for purifying proteins, enzymes, and other biomolecules.
Adsorption chromatography is a type of chromatography that uses adsorption to separate mixtures of molecules. In adsorption chromatography, a stationary phase is used to adsorb molecules from a mobile phase. The molecules in the mixture are separated based on their affinity for the stationary phase.
There are several types of adsorption chromatography, including normal phase chromatography, reverse phase chromatography, and affinity chromatography. Normal phase chromatography uses a polar stationary phase and a nonpolar mobile phase, while reverse phase chromatography uses a nonpolar stationary phase and a polar mobile phase. Affinity chromatography uses a stationary phase that is specifically designed to adsorb a particular type of molecule, based on its affinity for the stationary phase.
Adsorption chromatography is often used in biochemistry and molecular biology to purify proteins and other biomolecules. It is also used in the food and beverage industry to purify and concentrate food ingredients.
• The separation is done through the solid stationary phase (Silica gel or Alumina) and the liquid mobile phase single or mixture solvent.
• The separation is mainly due to the adsorption force that existed to hold the molecules based on the interaction with the adsorbate at different rates.
Gel filtration chromatography
Gel filtration chromatography is a separation technique that uses a column filled with a porous, gel-like substance to separate molecules based on their size. The column is packed with beads that have tiny pores, and as the sample mixture is passed through the column, the molecules are separated based on their ability to pass through the pores. Smaller molecules pass through the pores more easily and elute (come out of the column) first, while larger molecules are retained in the pores and elute later.
Gel filtration chromatography is commonly used to purify proteins and other biomolecules, and can also be used to determine the size of molecules or to separate molecules based on their charge or hydration properties. It is a useful technique for separating molecules that are too large or complex to be separated by other methods, such as size exclusion chromatography or affinity chromatography.
• It separates the proteins based on molecular size or weight differences.
• The microporous packing material made of gel is used to separate the molecules pumped through it where the small-sized molecules access the interior of the pores partly or wholly.