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Efficient methods for DNA detectionin clinical, environmental, and experimental samples are constantly in demand. In a clinical sample, DNA capture and identification can be essential to the diagnosis of disease. In public heath and environmental situations it can be used to identify contamination of food or water. DNA collection and characterization is constantly expanding our ability to answer experimental questions. DNA capture is a mainstay of modern biotechnology. Traditional techniques rely on affinity columns, centrifugation, and multiple washing steps. Newer methods are based on magnetic separation. In the beginning, magnetic separation was limitedto packing a column with magnetic material and running a solution through it. As nanotechnology evolved it became possible to use mobile solid support systems such as magnetic nanoparticles.

 

Free guide: The basic guide to scale-up biomagnetic separation processes

Benefits of using mobile magnetic nanoparticles for DNA capture

Magnetic DNA capture is an improvement over affinity columns because it is faster, uses fewer washing reagents, and can often be gentler on the nucleic acids because a centrifuge is unnecessary. In traditional methods the centrifuge introduced high shear forces that potentiallydegraded the DNA. This is avoided in modern DNA capture using magnetic nanoparticles.

Additionally, the nanoparticles are an improvement over solid magnetic columns because they can move freely in solution. It is possible to incubate the beads with the sample for as long as necessary instead of pouring the sample multiple times through a column. The incubation time provides a greater opportunity for DNA to adsorb to the nanoparticle surface, which can lead to improved capture efficiency.

The magnetic particles are easily isolated by magnetic separation. The magnetic particles are manipulated by an external magnet, which sets up a magnetic field gradient and pulls the DNA-bound particles to the walls of the container. The particles are held there while the contaminating solution is washed away. To recover the DNA-bound particles one only needs to remove the magnet to release them into a collection buffer.

 

Methods of binding DNA to magnetic nanoparticles

Uncoated iron oxide (Fe3O4) nanoparticles are capable of adsorbing DNA, but the particles are susceptible to aggregating together and limiting the surface area available for DNA binding. To solve this problem the nanoparticles are coated with polymer to make them biocompatible and to reduce aggregation. Another benefit ofpolymer coatings is the ability to functionalize the surfaces to adsorb DNA with greater specificity. The surface chemistry of the polymer can be functionalized. A few common strategies include:  

  1. Streptavidin coating: single strand probe DNA with a biotin tag can be purchased with the exact sequence of interest. The streptavidin-biotin affinity is specific and strong.
  2. Silica surface: the positively charged silica attracts the negatively charged DNA through electrostatic interactions
  3. Amine modification:introduces a pH responsive reversible DNA binding component based on electrostatic interactions
  4. Imidazol functionalization: improves the reversibility of DNA adsorption link to recent article (DNA isolation from bacteria using charge-reversible magnetic nanoparticles)
  5. Sandwich hybridization (article from this month): hybridization of DNA to two oligonucleotide probes attached to magnetic particles and oftenalso attached to fluorescent probes for quick detection of DNA as an in vitro diagnostic device (IVD)
 
 

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Lluis M. Martínez | SEPMAG Chief Scientific Officer

Founder of SEPMAG, Lluis holds a PhD in Magnetic Materials by the UAB. He has conducted research at German and Spanish academic institutions. Having worked in companies in Ireland, USA and Spain, he has more than 20 years of experience applying magnetic materials and sensors to industrial products and processes. He has filed several international patents on the field and co-authored more than 20 scientific papers, most of them on the subject of magnetic particle movement.

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