The History Of Electrophoresis

The movement of electrically-charged particles or ions in liquids caused by an applied electric field is known as electrophoresis. The capacity to separate highly similar compounds, including distinct proteins, for analytical and preparative applications has risen, particularly after 1950, as a result of the advent of zone electrophoresis in paper and, later, in polyacrylamide or agarose gels. After 1960, disc and displacement electrophoresis (isotachophoresis), as well as isoelectric focusing, provided much-improved resolution.

Electrophoretic methods currently promote advances in biochemistry and molecular biology and will proceed to be quite important in science and for various applications in genetics, gene technology, nucleic acid and protein sequencing, studies of diseases and malfunctions, including cancer, and in the identification of species and individuals, for example, in forensic medicine.

History of electrophoresis

Arne Tiselius, a Swedish scientist who published his first study on electrophoresis, in 1937, in the publication, “A New Apparatus for Electrophoretic Analysis of Colloidal Mixtures”, and was given the Nobel Prize in 1948 for his work, was a pioneer in the creation of gel electrophoresis. This was recognised as Tiselius’ Movable Boundary Electrophoresis (MBE), which employed a U-shaped tank with electrodes at each end to effectively separate molecules in free solution. After its creation, Arne Tiselius’ moving boundary electrophoresis method was widely used. However, it was quickly overtaken by zone electrophoresis (ZE) in the 1950s, which divided the molecules in multiple solid supporting media including cellulose, starch grains and filter paper.

The name “electrophoresis” was derived from the Greek word “phoresis,” which means “being carried,” and originally referred to the mobility of charged particles inside an electrical field. Since the 1980s, these related approaches have swiftly grown to become crucial bioanalytical tools, serving as the foundation for a wide range of biochemical procedures such as DNA fingerprinting, Western blot, Southern blot, and others.

Furthermore, it is a critical preparative technique for fractionally purifying the necessary biomolecules (DNA) prior to further characterisation and identification using other advanced molecular techniques and technologies like DNA sequencing or polymerase chain reaction (PCR).

Emergence of electrophoresis

Protein separation at a high resolution based on charge differences is feasible by utilising disc electrophoresis, displacement electrophoresis (isotachophoresis), and, most significantly, isoelectric focusing (IEF). Electrophoresis of sodium dodecyl sulphate-polyacrylamide gel yields size separation (SDS-PAGE). The combination of gel IEF accompanied by SDS-PAGE in a second-dimensional slab gel, i.e. two-dimensional gel electrophoresis, provides the maximum resolution with thousands of spots per gel. Protein staining produces high-resolution patterns that may be scanned and saved in large databases.

Throughout the last ten years, electrophoretic separation in gels and subsequent imaging of nucleic acids (DNA, RNA), genes, and nucleotides have greatly improved, allowing for effective gene mapping in humans and all other animals. This has resulted in the largest coordinated scientific undertaking in history, namely the mapping of the human genome, which will be important for the rest of humanity’s existence.

Electrophoresis in capillaries has gained popularity in recent years because it provides great resolution on the analytical scale for a variety of compounds such as proteins, nucleic acids, medicines, metabolites, and peptides. Electrophoretic approaches have had an unparalleled influence on life sciences, laying the groundwork for breakthroughs in biochemistry, molecular biology, genetics, gene technology, and medicine.

Basic principle of electrophoresis

Gel electrophoresis is a typical molecular biology laboratory technique for identifying, quantifying, and purifying nucleic acids. The approach is frequently used for nucleic acid isolation and analysis because of its rapidity, simplicity, and adaptability. Nucleic acids in the range of 0.1–25 kbp may be separated for examination using gel electrophoresis within a few hours or minutes, and separated nucleic acids could be retrieved from the gels with reasonably high purity and efficiency. 

The approach includes applying an electrical field to a mixture of the molecules that are charged to force them to migrate across a gel matrix based on charge, size, and structure. The phosphate groups of nucleic acid ribose-phosphate backbones are charged negatively at basic pH to neutral.

As a result, each nucleotide has a net negative charge, which commonly includes charge of a sequence of nucleotides that is dependent on the total number of nucleotides or mass. In other words, the charge-to-mass ratio of DNA or RNA molecules is constant. As a result, while they have equivalent structure, their mobility in gel electrophoresis is dictated mostly by size. When exposed in front of a field of electric, the nucleic acids might migrate from the end of negative electrodes to the positive electrode (anode), with shorter fragments migrating faster compared to the ones that are longer which results in the separation of sizes.

Moreover, the migration lengths of nucleic acids in gel electrophoresis frequently show a predictable relationship with their sizes, allowing the size of nucleic acids in a specific sample to be estimated. Migration length for linear double-stranded DNA fragments is inversely proportional to the log of the molecular weight within a certain range.

Conclusion

Generally, gel electrophoresis became a ubiquitous approach for isolating nucleic acids in molecular biology. This analytical and preparative approach is not only essential in typical workflows such as molecular cloning and PCR, but also plays a critical role in nucleic acid separation and analysis in upcoming technologies such as genome editing and next-generation sequencing.