After scientists disrupt cell membranes through the process of cell lysis and remove the biological material within, they carry out various downstream applications to fulfill their research goals and objectives. This article looks at 4 common downstream applications following cell lysis: protein purification, proteomics, Western blotting and immunoprecipitation.
Protein Purification
Prevalent in the biotech and pharmaceutical industries, protein purification involves several processes and specific tools designed to isolate one or a small number of proteins from the cell, so that the protein(s) function, structure and interaction can be studied.
Proteomics
Often used in disease treatment research, drug discovery and proteogenomics (studies based on proteomic data in order to improve gene annotations), proteomics is the large scale study of the full set of proteins produced by a system or organism; and in particular, their functions and structures. During proteomics, researchers purify and analyze proteins using various tools (e.g. electrophoresis systems, mass spectrophotometry systems, etc.). Proper tool selection depends on various factors, including the type of cell, the biological system of interest, and time, size and budget limitations faced by the laboratory.
Western blotting
Western blotting (a.k.a. immunoblotting) is a common downstream application that uses antibodies to help researchers identify specific proteins from a sample. There are three aspects of the western blotting process: 1) separating the extracted proteins based on size; 2) transferring the proteins to a solid support such as an enzyme or fluorescent dye; 3) visualizing the target protein via chemiluminescent or chemifluorescent detection reagents, or by using the fluorescent tag.
Immunoprecipitation
Immunoprecipitation – often referred to in research literature by the acronym IP – is a process that leverages the antigen-antibody reaction principle in order to enable the purification or a protein (or a protein complex), so that researchers can examine physical characteristics or quantity. There are typically four steps in the process: 1) the proteins extracted via cell lysis are suitably precipitated; 2) the immune complex is captured on a solid support upon which Protein G or Protein B has been immobilized; 3) elements that bind to the immune complex are removed via elution from the support; 4) researchers further analyze the immunoprecipited proteins as per their plans.
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