Biomolecule purification methods

The ability to control the interaction between a wide diversity of biomolecules with surfaces can be also exploited as an effective way to develop reagentless, sensitive, reusable, and real-time biosensors [51-56]. Such sophisticated biosensors are expected to impact a wide range of applications, from clinical diagnosis[57] and environmental monitoring [58] to forensic analysis [59]. Another significant potential application of dynamic surfaces is in bioseparation of proteins and other biomolecules for basic life science research, as well as industrial applications [60-63]. With the rapid development of recombinant proteins in the treatment of various human diseases, the dynamic surface-based bioseparation systems could meet the demand for more reliable and efficient protein purification methods [64]. Stimuli-responsive surfaces are also expected to play a crucial role in the search for more controllable and precise drug delivery systems [65]. 

Countercurrent chromatography is based on the distribution of substances in two liquid phases [128,129]. The liquid is fed into a coiled tube that is moved along an orbital trajectory. Due to centrifugal power, the liquids move in a counter-current. For proteins and many other biomolecules, this method is not practical because of denaturation in a nonaqueous phase. In aqueous two-phase systems, at least one phase exhibits high viscosity and, therefore, mass transfer between the two phases is limited. Similar problems occur with reversed micelle extraction as were observed with the aqueous two-phase extraction [130]. CCC has not been used for large-scale purification of proteins and other biopolymers. 

The application and preparation of afEnity resins was first described in a paper by Cuatrecasas in 1968 [1]. In this work, nearly aU features of this technique were explored. Affinity chromatography is the youngest of the four major purification methods used in biochromatography. In 1968, ion exchange (lEX), hydrophobic interaction (HIC), and gel filtration (GF) were already well established. What new opportunities in the purification of biomolecules were then added by affinity chromatography ... 

Most of these methods are suitable for analyzing the components present in cell homogenates and other biochemical preparations. The sequential use of several techniques will generally permit purification of most biomolecules. The reader is referred to texts on methods of biochemical research for details. 

In 1994, thiols were firstly used as stabilizers of gold nanoparticles [6a]. Thiols form monolayer on gold surface [18] and highly stable nanoparticles could be obtained. Purification of nanoparticles can be carried out, which makes chemical method of metal nanoparticles a real process for nanomaterial preparation. Various thiol derivatives have been used to functionalize metal nanoparticles [6b, 19]. Cationic and anionic thiol compounds were used to obtain hydrosols of metal nanoparticles. Quaternary ammonium-thiol compounds make the nanoparticle surface highly positively charged [20]. In such cases, cationic nanoparticles were densely adsorbed onto oppositely charged surfaces. DNA or other biomolecule-attached gold nanoparticles have been proposed for biosensors [21]. 

Nucleic acids may also be removed by treatment with nucleases, which catalyse the enzymatic degradation of these biomolecules. Indeed, nuclease treatment is quickly becoming the most popular method of nucleic acid removal during protein purification. This treatment is efficient, inexpensive and, unlike many of the chemical precipitants used, nuclease preparations themselves are innocuous and do not compromise the final protein product. 

Chromatography Separation method that uses columns filled with gels that separate a mixture of molecules into individual components by size or charge Purification of biomolecules... 

Electrophoresis makes use of differences in the electrophoretic mobility of electrically charged particles (biomolecules, micro-organisms etc.) as a means to separate them. For this purpose, a homogeneous, rectified electrical field is used. Thanks to the excellent resolution and mild operating conditions, this is currently the best analytical method for protein separation, purification and characterization. It is also used as a preparative separation method which allows a few grams per hour to be purified. 

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