Hydrophobic interaction chromatography (HIC) is one of the most widely used methods for separating and purifying proteins in their native state. HIC also proves to be quite useful in isolating protein complexes and in studying protein folding and unfolding. So, how does HIC work? What are the principles behind this technique? To understand the mechanism behind this type of chromatography, here are some things you definitely need to know.
Topics: Protein Purification
It is interesting to note that while detergent-solubilized membrane proteins can be purified and concentrated through phase separation (which is a simple, efficient and cost-effective method that can be used for this purpose), most membrane protein scientists do not use this method or even know that such methods exist. To better understand and appreciate how this method works, here are some things you definitely need to know.
Phase Separation: Understanding the Basics
In a nutshell, phase separation (also known as cloud point extraction) refers to the state wherein the micelles in an aqueous solution become immiscible with water and form large aggregates that will separate from the water phase. This condition is usually achieved upon the addition of detergents or when the temperature or salt concentration of the solution is altered.
Topics: Protein Purification
Researchers have traditionally used agarose beads for immunoprecipitation (IP). However, there has been a growing trend in recent years favoring the use of magnetic beads. According to a recent survey of 1,013 scientists, 60% of the respondents will start using magnetic technology in the next three years.
Topics: Protein Concentration, Antibody Production, Protein Extraction
When it comes to Western blotting, choosing the right membrane for your application usually spells the difference between success and failure. However, this is usually easier said than done since you need to consider the properties of your protein and the downstream detection steps required in your application before you can determine which membrane can give you the results that you need.
Topics: Western Blotting
