The Protein Man's Blog | A Discussion of Protein Research

Due to the fact that nucleic acids carry genetic information and that proteins regulate various life processes, they are considered to be two of the most important biomolecules in any living organism. In addition, their interactions play a crucial role in most biological processes, which include everything from replication,transcription and recombination to enzymatic eventsusing nucleic acids as substrates. Taking all of these things into consideration, it is not surprising why protein-nucleic acids interactions have been the subject of intensive research for the past few years.

Since 1975, immobilized metal affinity chromatography (IMAC) has been popularly used in purifying proteins, especially those that are fused to a polyhistidine tag, typically a 6X His tag. This process gained immense popularity since it allows for the efficient purification of proteins, even those from crude lysates. In addition, its robust nature makes it ideal for methods that require protein-specific conditions. Its functional simplicity, affordability and compatibility with a wide range of reagents also add to its popularity.

Protein crosslinking reagents or crosslinkers can be accurately defined as molecules containing two or more reactive ends that are capable of chemically attaching to specific functional groups on proteins or other molecules. These reagents are generally used in creating detectable scientific probes to facilitate a number of proteomics methods, including Western blotting, ELISA and other strategies for studying protein-protein interactions.

Most biological samples contain non-protein substances and/or contaminants that may interfere with the resolution of the electrophoretic separation. Considering the fact that cell lysates usually contain macromolecules and small ionic molecules which may interfere with the electrophoretic process, these agents should be eliminated prior to electrophoresis, especially if their amount exceeds the critical interference threshold.