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Membrane Protein Isolation and Purification With Phase Separation

Posted by The Protein Man on Dec 22, 2014 11:00:00 AM

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.

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Topics: Protein Purification

Magnetic Beads for Immunoprecipitation

Posted by The Protein Man on Nov 18, 2014 9:00:00 AM

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.

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Topics: Protein Concentration, Antibody Production, Protein Extraction

PVDF or Nitrocellulose - Which Membrane is Best?

Posted by The Protein Man on Nov 12, 2014 9:00:00 AM

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.

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Topics: Western Blotting

A Look into the 6XHis Tag and its Uses

Posted by The Protein Man on Oct 20, 2014 9:03:00 AM

The 6xHis tag, also known as polyhistidine tag, His6 tag and/or hexa histidine tag, is an amino acid motif consisting of at least 6 histidine residues fused to the carboxyl (C-) or amino (N-) terminus of a target protein in transfected cells. This tag is most commonly used in the production of recombinant proteins since the string of histidine residues binds to several types of immobilized ions (such as nickel, cobalt and copper) under specific buffer conditions to allow for the simple detection and purification of His-tagged proteins.

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Topics: Protein Purification

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