Techniques Involved in Proteomics Study

Some of the very basic analytical techniques are used as major proteomic tools for studying the proteome of an organism. We shall study most of these techniques as we progress in the course. The initial step in all proteomic studies is the separation of a mixture of proteins. This can be carried out using Two Dimensional Gel Electrophoresis technique in which proteins are first of all separated based on their individual charges in 1D. The gel is then turned 90 degrees from its initial position to separate proteins based on the difference in their size. This separation occurs in 2 nd dimension hence the name 2D. The spots obtained in 2D electrophoresis are excised and further subjected to mass spectrometric analysis of each protein present in the mixture.

Apart from charge and size of proteins there are number of other intrinsic properties of proteins that can be employed for their separation and detection. One of these techniques is Field Flow Fractionation (FFF) which separates proteins based on their mobility in presence of applied field. The difference in mobility may be attributed to different size and mass of proteins. The applied field can be of many types such as electrical, gravitational, centrifugal etc. This technique helps in determining different components in a protein mixture, different conformations of protein, their interaction with other proteins as well as some organic molecules such as drugs (Fig. 2).

Figure 2. Techniques involved in protein identification during proteomic analysis.