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.
PVDF or Nitrocellulose - Which Membrane is Best?
Topics: Western Blotting
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.
Topics: Protein Purification
Which agarose (sepharose) to choose? 2, 4 or 6%? Crosslinked?
Agarose beads are small spherical beads of agarose gel which are commonly used in gel filtration or molecular size exclusion chromatography and biomolecular purification and immobilization. These beads act as porous gel to filter mixtures of molecules based on their individual sizes. And since these beads are easy to activate, they can also be used to bind biomolecules in a reversible or irreversible manner. In addition, their inert nature and unique internal surface area can also be activated for ligand attachment, making them the ideal basis for various affinity chromatography beads such as protein A and G, and glutathione.
Topics: Protein Purification, Sample Clean Up
Spectrophotometry and Its Application in Protein Estimation
Basically, spectrophotometry is one of the most widely used analytical procedures in biochemistry. It is commonly used to estimate the level of an analyte in solution and is ideal for simple routine determination of small quantities of materials. This method is based on the two laws of light absorption by solutions, namely Lambert's Law and Beer's Law.
Topics: Protein Estimation