Applications and Uses of Functionalised Magnetic Silica Nanoparticles

Application Area 3: Magnetic Protein Separation

Isolation, separation and purification of various types of proteins and peptides, as well as of other specific molecules, is used in almost all branches of biosciences and biotechnologies. Separation science and technology is thus very important area necessary for further developments in bio-oriented research and technology. New separation techniques, capable of treating dilute solutions or solutions containing only minute amounts of target molecules in the presence of vast amounts of accompanying compounds in both small and large-scale processes, even in the presence of particulate matter, are necessary.

In the area of biosciences and biotechnology the isolation of proteins and peptides is usually performed using variety of chromatography, electrophoretic, ultrafiltration, precipitation and other procedures, affinity chromatography being one of the most important techniques. Affinity ligand techniques represent currently the most powerful tool available to the downstream processing both in term of their selectivity and recovery. The strength of column affinity chromatography has been shown in thousands of successful applications, especially in the laboratory scale.

However, the disadvantage of all standard column liquid chromatography procedures is the impossibility of the standard column systems to cope with the samples containing particulate material so they are not suitable for work in early stages of the isolation/purification process where suspended solid and fouling components are present in the sample. In this case magnetic affinity, ion-exchange, hydrophobic or adsorption batch separation processes, applications of magnetically stabilized fluidized beds or magnetically modified two-phase systems have shown their usefulness.

Magnetic carriers bearing an immobilized affinity or hydrophobic ligand or ion-exchange groups, or magnetic biopolymer particles having affinity to the isolated structure, are mixed with a sample containing target compound(s). Samples may be crude cell lysates, whole blood, plasma, ascites fluid, milk, whey, urine, cultivation media, wastes from food and fermentation industry and many others.

Following an incubation period when the target compound(s) bind to the magnetic particles the whole magnetic complex is easily and rapidly removed from the sample using an appropriate magnetic separator. After washing out the contaminants, the isolated target compound(s) can be eluted and used for further work.  The magnetic separation techniques are also the basis of various automated procedures, especially magnetic-particle based immunoassay systems for the determination of a variety of analytes, among them proteins and peptides. Several automated systems for the separation of proteins or nucleic acids have become available recently.

Magnetic separation is usually very gentle to the target proteins or peptides. Even large protein complexes that tend to be broken up by traditional column chromatography techniques may remain intact when using the very gentle magnetic separation procedure [2]. Both the reduced shearing forces and the higher protein concentration throughout the isolation process positively influence the separation process.

Separation of target proteins using standard chromatography techniques often leads to the large volume of diluted protein solution. In this case appropriate magnetic particles can be used for their concentration instead of ultrafiltration, precipitation etc.

Application 4: Magnetic Cell Isolation

The separation and sorting of biological cells is critical to a variety of biomedical applications including diagnostics, therapeutics, and fundamental cell biology. As samples of interest are often heterogeneous populations of cells that are in culture or that comprise a tissue, techniques to isolate specific cells are essential for understanding how cells function and respond to various stimuli.

Blood, for example, is an extremely information-rich and easily accessible tissue that is a complex blend of cells; accurate analysis of blood character and condition requires isolation of a few desired cells. Effective cell sorting to support numerous biomedical pursuits relies upon optimal matching between the target cell attributes, desired outcomes, and the parameters of the sorting technique. Numerous cell isolation and sorting techniques have been developed for benchtop and clinical settings that are based on either physical properties of the cell, such as density or size, or on cell affinity that describes electric, magnetic or adhesive properties specific to each cell type.

Standard techniques for the separation of cells include processing steps of filtration, centrifugation and sedimentation, which are carried out either in a batch or in a continuous manner and can be easily translated to large-scale operation. However, in situations where cell size or density differences are not significant, effective cell separation is impeded in these techniques and other methods must be employed, including fluorescence activated cell sorting (FACS) and magnetic activated cell sorting (MACS).

In this context, magnetic particles —nanoparticles (mean diameter 10 – 100 nm), sub-micron particles (0.1 – 1 microns), and microparticles (mean diameter 1 – 50 microns) — have been an important component of cell separation techniques in both biomedical research and in clinical medicine.