Isolation of DNA is a prerequisite step for many molecular biology techniques. The separation of DNA from the complex mixtures in which they are often found is frequently necessary before other studies and procedure like sequencing, amplification, hybridization, detection etc. The presence of large amounts of cellular or other contaminating material like proteins and RNA, in such complex mixtures often impedes many of the reactions and techniques used in molecular biology 5. The conventional protocol for extracting DNA involves cell lysis followed by removal of contaminating cellular components such as proteins, lipids and carbohydrates; and finally isolating DNA using a series of precipitation and centrifugation steps, which are difficult to automate. Improvement in methods for isolating DNA has been made and more recently, methods that rely on the use of solid phase have been proposed. Adsorbents that provide fast, efficient DNA purification are important for making this procedure amenable to automation. The discovery in late 80's that silica can be used as adsorbent for DNA isolation 6, became the basis for most of the DNA isolation kits currently available. One of these kits involves isolation of DNA using silica coated magnetic particles 7.

A high throughput genome isolation protocol has been developed, which is based on SPRI (Solid Phase Reversible Immobilization) chemistry 89. The SPRI protocol is based on DNA binding to the surface of carboxyl coated paramagnetic particles under the condition of high salt and PEG. The procedure has been developed and optimized for single stranded DNA isolation, such as M13 phage and double stranded plasmid DNA utilizing carboxyl coated magnetic particles. The SPRI protocol has allowed the development of an automated procedure in a microplate format with a throughput of about 200, 000 DNA preparations per day, thus becoming a fastest microplate plate based DNA purification system in use on the human genome sequencing.

A number of methods have been reported for isolation of total RNA from a variety of cells or tissues. Chirgwin et al. 10 developed a method for efficient isolation of total RNA by homogenization in a 4 M solution of guanidium thiocyanate containing 0.1 M 2-mercaptoethanol. Homogenization is followed by extraction of RNA by ethanol or by ultracentrifugation through cesium chloride. This method was further modified by Chomczynski and Sacchi 11 to devise a rapid single step isolation procedure for RNA. It involves extraction of RNA using a mixture of guanidium thiocyanate and phenol-chloroform. Many RNA isolation kits are available based on the above two protocols. All these methods isolate RNA on the basis of its biochemical properties. In contrast, biomagnetic separation of mRNA is based on specific complementary hybridization between poly A sequence of isolated mRNA and oligo (dT)₂₅ sequence covalently linked to the surface of paramagnetic particles. In this method oligo (dT)₂₅ coated magnetic beads are added to crude cell or tissue lysate. During incubation poly (A) mRNA from the lysate are caught onto the surface of oligo (dT)₂₅ coated magnetic beads. The beads/mRNA complex is then washed magnetically. The mRNA thus isolated is either eluted or directly applied for many downstream applications which includes cDNA library construction, Substractive hybridization, Northern hybridization, RT-PCR and in vitro translation 12.

The isolation of specific molecules based on its interaction with complementary binding partner is emerging as important technologies in many field of research. The isolation and characterization of specific transcripts or proteins can be employed to monitor the progression of disease. Several kits are available in the market that works on the principle of magnetic labeling and direct isolation of biotinylated molecules such as DNA, RNA or proteins onto streptavidin coated magnetic beads (μMACS streptavidin Microbeads from Miltenyi Biotec, Germany). These biotinylated molecules can then be used for indirect isolation of non-biotinylated target molecules that may interact with them 13. The procedure involves complex formation between the biotinylated probe (DNA, RNA or proteins) and the target molecule (i.e. interacting biomolecules DNA, RNA or protein). Based on the interaction of biotin-streptavidin, the probe-target complex is then separated from rest of the component by addition of streptavidin coated magnetic beads. The complex is magnetically isolated and washed to remove non-specifically bound molecules. The non-biotinylated target molecules can be either eluted off from the complex with high purity, whereas the magnetically labeled biotinylated probe remains bound to the column (Fig. 1). This technique has potential for rapid and efficient screening of transcriptional and translational regulatory proteins. Alam and co-workers have also used target DNA-conjugated magnetic beads for rapid screening of DNA-binding peptide ligands from solid phase combinatorial library 14.

Magnetic particles are now increasingly used as carriers for binding proteins, enzymes and drugs. Studies have shown that proteins and enzymes can be bound covalently to naked magnetic particles in the presence of carbodiimide 15. Such immobilization procedures for proteins, enzymes or drugs will have a major impact in various areas of medicine and biotechnology. The immobilized biomolecules can be used directly for a bioassay or as affinity ligands to capture or modify target molecules or cells. On this basis, Ni-NTA (nitriloacetic acid) tagged magnetic agarose beads have been used for versatile magnetocapture assays using 6xHis-tagged proteins 16. The procedure involves use of metal chelating nitriloacetic acid (NTA) groups covalently bound to the surface of agarose beads, which contain strong magnetic particles. The beads are precharged with nickel, which is ready to capture 6xHis-tagged proteins for sensitive interaction assays or microscale purification of 6xHis tagged proteins (Fig. 2). Thus, this technique bridge the gap between purification-scale procedures using Ni-NTA metal chelate affinity chromatography resins and microplate-based assays 17. Other procedures for magnetic affinity separations of various proteins have been summarized recently 18.

Magnetically guided drug targeting has been attempted in order to increase the efficacy and reduce the unpleasant side effects associated with chemotherapy. This method of drug delivery involves immobilization of drug or radionuclide in biocompatible magnetic nano- or microspheres 19. This method of delivery makes chemotherapy more effective by increasing the drug concentration at the tumour site, while limiting the systemic drug concentration. FeRx Inc. of San Diego have designed Magnetic Targeted Carriers termed MTCs for site-specific targeting, tissue retention, and sustained release of drugs. These MTCs are composed of elemental iron particles and activated carbon. MTCs (1–2 μm in size) can adsorb and desorb pharmaceutical agents such as doxorubicin (DOX). Vialed MTCs are mixed with an anticancer drug already in solution; the mixture is then introduced into the catheter. The drug delivery using MTCs includes insertion of catheter into an arterial feed to the tumor, followed by application of powerful magnetic field to cause the MTC-DOX to extravasate through the capillary bed into the targeted tissue (Fig. 3). The field is left in place for another 15 minutes; after removal of the magnet, the particles remain trapped in the tumor, where the drug is then released 20. FeRx has already initiated phase III clinical trial for MTC-DOX formulation 21. Like wise, Hafeli and co-workers have carried out preclinical studies in collaboration with FeRx for treatment of liver and brain tumors using MTCs labeled with β-emitters such as Rhenium-188 and Yttrium-90 22. Although the main research concern of MTC-drug therapy is cancer treatment, the technology is not limited to this, other drugs that are deliverable in this manner include antibiotics, thrombolytics, anti-inflammatories, peptides and steroids.

Magnetoliposomes are magnetic derivatives of liposomes and can be prepared by entrapment of ferrofluids within the core of liposomes 23. Several groups have investigated the uses of magnetoliposomes for site specific targeting 24, cell-sorting 25 and as magnetic resonance contrast enhancing agents 26.

Babincova et al. 27 have tried to use magnetoliposomes with encapsulated doxorubicin (DOX) for site specific chemotherapy in response to externally applied AC magnetic field. The result of this study showed that magnetoliposomes can be specifically heated to 42°C in a few minutes and during this, the encapsulated doxorubicin is massively released. Another in vivo studies for site specific targeting using magnetoliposomes incorporated with adriamycin (ADR), showed that administration of magnetic ADR liposomes under an applied external magnetic force produced approximately 4-fold higher maximum ADR concentration in the tumor than did administration of ADR solution alone. These results suggested that systemic chemotherapy could effectively control the primary tumor without significant side effects, due to the targeting of magnetic ADR liposomes 2829.

Magnetotactic bacteria belonging mainly to Magnetospirillum species produces ferromagnetic crystalline particles consisting of magnetite (Fe₃O₄) or greigite (Fe₃S₄). These are called bacterial magnetic particles (BMPs). The BMPs are small in size (50–100 nm) and disperse very well because they are covered with a stable lipid membrane 30. Magnetoliposomes containing cis-diamminedichloroplatinum (II) (CDDP) prepared using BMPs have also been evaluated for targeting and controlled release of drugs at tumor site. It has been observed from such studies that the capture volume of the magnetoliposomes with BMPs was 1.7 times higher than the ones with artificial magnetic particles. Also, when a rotating magnetic field was applied to these magnetoliposomes, the contents of them were released over 2 hours, suggesting that magnetoliposomes containing CDDP shows antitumor activity and that the controlled release of drugs from magnetoliposomes is possible 31.

Magnetic fluids have been investigated as potential hyperthermia causing agents due to their high specific absorption rate (SAR). Hyperthermia is a promising approach for cancer treatment that uses AC magnetic fields to heat target areas (cancer tissue) containing magnetic fluids. To study the biological effects of AC magnetic field excited ferrofluids, both in-vitro and in-vivo studies have been carried out in cancer cell lines and spontaneously induced tumors in animal models. The results of these studies have shown that magnetic fluid hyperthermia is able to reduce the viability of cancer cells, thereby indicating the potential of this therapy 32. There also exist the possibility of combination therapy, which would include hyperthermia treatment followed by chemotherapy or gene therapy. The approach involves use of magnetic nano- or microspheres or magnetoliposomes containing a drug to cause hyperthermia using the standard procedure, followed by the release of encapsulated drug that will act on the injured cells. It is anticipated that the combined treatment might be very efficient for treating solid tumor 19.

Immunomagnetic cell separation methods have become increasingly popular among the cell biologist. The general approach involves use of paramagnetic particles coated with antibodies against the target specific cell surface molecules. The method employs two schemes for isolating the target cell, direct method that involves coupling of affinity ligand (antibody) onto the magnetic particles, which are then directly added to the sample containing the target cells. During the incubation the magnetic particles binds the target cells that can then be recovered using a magnet. While in case of indirect method, a free affinity ligand (an appropriate antibody) is first added to the cell suspension. After incubation excess unbound affinity ligand is removed by washing, and the antibody-target cell complex is then captured by magnetic particles bearing an affinity ligand (secondary antibody) with the affinity for the primary label. Both positive and negative selection can be performed with immunomagnetic separation. In case of positive selection target cellular subsets are magnetically labeled and subsequently separated, while negative selection involves target purification by removing all other contaminating cells 3334.

Using this technology large number of cell types has been isolated to date. Some of the examples where immunomagnetic separation technology has been applied includes, detection and removal of circulating tumor cells from peripheral blood 35, isolation of antigen specific CD 8+ T cells 36, selective separation of CD34+ cells 37, isolation and enrichment of retinal ganglion cells (RGCs) for culturing 38 etc. Immunomagnetic separation is used extensively for detection of pathogenic bacteria in clinical, food and environmental samples 3339. In addition to whole cell isolation, even cell organelles can be selectively separated using magnetic particles. The lysosome fraction was isolated from the amoeba Dictyostelium discoideum after feeding with dextran-based nanoparticles and subsequent homogenization 40. Likewise a variety of cell types can be isolated using automated systems and different surface coated magnetic particles that are available from Miltenyi Biotec, Germany, or StemCell Technologies, USA.

Immunoassays are becoming an important tool in clinical diagnostics and in fundamental investigations, because of its sensitivity, specificity and general applicability. Magnetic particles have been applied increasingly for various immunoassays that include fluoroimmunoassays, enzyme immunoassays or radioimmunoassays. Magnetic particles bound with primary or secondary antibodies are used for separation and quantification of antigens. Use of magnetically bound antibodies eliminates the centrifugation step thus reducing assay time and simplifying operation thereby increasing the efficiency and accuracy of the assay. Matsunaga and co-workers 41 have developed a novel fluoroimmunoassay method, where a fluorescein isothiocyanate (FITC) conjugated monoclonal anti-Escherichia coli antibody was immobilized onto bacterial magnetic particles (BMPs) for detection and removal of E. coli. The same group has also developed a chemiluminescence enzyme immunoassay with BMPs using IgG as a model antigen 42. Likewise a magnetic bead based ELISA has been developed for detection of Staphylococcus species 43.