Magnetic Separation Device Research Articles

Automated extraction of DNA and PCR setup using a Tecan Freedom EVO ® liquid handler

We have implemented and validated automated methods for DNA extraction and PCR setup developed for a Tecan Freedom EVO ® liquid handler mounted with a Te-MagS™ magnetic separation device. The DNA was extracted using the Qiagen MagAttract ® DNA Mini M48 kit. The DNA was amplified using AmpF ℓSTR ® Identifiler ®, Y-filer ® (Applied Biosystems), GenePrint ® FFFL and PowerPlex ® Y (Promega). The methods were validated for fresh whole blood and blood from deceased according to EN/ISO 17025.

Characterization of a Prototype Compact High Gradient Magnetic Separator Device for Blood Detoxification

A prototype compact magnetic separator device for human blood detoxification was characterized using blood-mimicking fluid (ethylene glycol-water solution) as well as whole blood. Magnetic separation at various applied magnetic fields (0.125, 0.33, and 0.44 T) and various flow rates (3.1–29.5 ml/min) showed that the device could efficiently separate magnetic spheres from blood-mimicking fluid at a moderate applied magnetic field (for example, <0.44 T) and relatively high flow rates (for example, ≤29.5 ml/min). The experiments done in flow circulation systems showed that higher flow rates might shorten the sphere recovery time and accelerate the detoxification process. In vitro separation from flowing blood showed that it is possible to use the device to efficiently recover spheres in a reasonably short time (≤60 min). Moreover, it was also demonstrated that the separator had little effect on the occurrence of hemolysis. All the results revealed that the separator could be a clinically applicable device for efficient separation of magnetic spheres from blood flow for human detoxification purpose.

Theoretical Analysis of a Magnetic Separator Device for Ex‐Vivo Blood Detoxification

The feasibility of a magnetic separator device for ex‐vivo blood detoxification was studied. This blood detoxification approach entails administering functionalized magnetic microspheres (FMMSs) into a patient's body by transdermal injection to capture and remove toxins from the blood using highly specific receptors attached to the surface of the FMMSs. These toxin‐loaded FMMSs are then removed from the body using extracorporeal blood circulation through a specially designed magnetic separator, based on high gradient magnetic separation principles. The performance of the magnetic separator, in terms of its collection efficiency (CE) of the FMMSs, was evaluated theoretically using a streamline analysis of a 2‐D model. The effects of blood velocity (1 to 20 cm/s), magnetic field strength (0.1 to 2.0 T), wire size (0.125 to 2.0 mm in radii), separator unit size at a fixed ratio of tube to wire diameter of one, tube length (2.0 to 20 cm), wire material (nickel, SS 430 and wairauite), and magnetic material comprising the FMMSs (iron, typical magnetite and weaker magnetite) on the CE were evaluated. Provided that the blood velocity was below 2 cm/s, CEs >80% could be attained under reasonable conditions, like when using FMMSs 400 nm in diameter and containing 60 wt% magnetite in a magnetic field of 0.5 T using a magnetic separator with 0.5 mm radii wire (at a fixed ratio of tube to wire diameter of close to one) that was 10 cm in length (same as the tube) and made of SS 430. CEs of between 30% and 80% could also be attained at blood velocities up to 20 cm/s without compromising the magnetic separator design. The magnetic separator performance improved by reducing the size of the unit with tubes and wires of equal radii, increasing the applied magnetic field strength, utilizing magnetic materials with the highest magnetizations, and increasing the length of the unit. Overall, the results from this study delineated the physically realistic conditions that make ex vivo blood detoxification possible with this magnetic separator device.

Magnetic nanocarriers: from material design to magnetic manipulation

Magnetic nanocarriers play an increasing role in various biomedical applications such as the separation of magnetically 'tagged' DNA, drug delivery or identification of biological species. Recent developments in nanotechnology allow the fabrication of both artificial nanocarriers and magnetic separation devices which may achieve great performances at an incredibly small-volume sample handling. However, as the sizes of magnetic carriers and separation devices diminish, important theoretical and experimental challenges may occur mainly due to the important variations of the surface-to-volume ratio. Under these circumstances, intrinsic properties of individual carriers and their influence on functionality and performances of the magnetic manipulation have to be investigated with a higher degree of accuracy. In this paper, we present the state-of-the-art techniques for extracting accurate information about individual magnetic entities from magnetic measurements performed on ordered arrays or clusters of magnetic nano-objects of various dimensionalities and geometries. As the mutual magnetic interactions may be responsible for collective effects, both analytical and numerical techniques for evaluating the mutual interactions between magnetic nanoparticles and nanowires are reviewed, special emphasis being put on the dimensionality of their assemblages. As the efficiency of the manipulation and capture of individual magnetic carriers in magnetic confinement devices depend strongly on their size, the theoretical description of the motion of magnetic particles under various conditions of flow and field configurations become very important especially with the transition to the nanoscale regime. The equations of motion of magnetic nanocarriers in continuous-flow microfluidic devices are reviewed and some solutions for superparamagnetic interacting clusters, non-interacting beads and ferromagnetic contiguous or codebar nanowires are presented. The influence of both carrier size and transition toward the nanoscale regime on the trappability of some magnetic nanocarriers is evaluated in terms of finite-length Navier boundary conditions and applied in order to compare the motion of superparamagnetic beads and ferromagnetic nanowires in conventional continuous-flow magnetic confinement devices.

A comprehensive in vitro investigation of a portable magnetic separator device for human blood detoxification

A portable magnetic separator device is being developed for a proposed magnetically based detoxification system. In this paper, the performance of this device was evaluated via preliminary in vitro flow experiments using simple fluids and a separator unit consisting of one tube and two metal wires, each at the top and bottom of the tube. The effects of the following factors were observed: mean flow velocity Uo (0.14–45 cm s−1), magnetic field strength μoHo (0.125–0.50 T), wire size Rw (0.125, 0.250 and 0.500 mm), wire length Lw (2, 5 and 10 cm), wire materials (nickel, stainless steel 304 and 430) and tube size (outer radius Ro = 0.30 mm and inner radius Ri = 0.25 mm; Ro = 0.50 mm and Ri = 0.375 mm; and Ro = 2.0 mm and Ri = 1.0 mm). Our observations showed that the experimental results fit well with the corresponding theoretical results from the model we previously developed at a low flow velocity area (for example, Uo ⩽ 20 cm s−1), strong external magnetic field (for example, ⩾0.30 T) and long wire length (for example, Lw = 10 cm). The experimental results also showed that more than 90% capture efficiency is indeed achievable under moderate systemic and operational conditions. Pressure drop measurements revealed that the device could work well under human physiological and clinical conditions, and sphere buildup would not have any considerable effect on the pressure drop of the device. The breakthrough experiments demonstrated that a lower flow rate V, higher applied magnetic field μoHo and diluted sphere suspension, i.e. lower Co, would delay the breakthrough. All the results indicate the promise of this portable magnetic separator device to efficiently in vivo sequestrate nano-/micro-spheres from blood flow in the future magnetically based detoxification system.

Improvement of Immunomagnetic Separation for Escherichia coli O157:H7 Detection by the PickPen Magnetic Particle Separation Device

Conventional immunomagnetic separation (IMS) procedures, which use an external magnetic source to capture magnetic particles against the side of a test tube, are labor-intensive and can have poor sensitivity for the target organism because of high background microflora that is not effectively washed away during the IMS process. This report compares the conventional IMS procedure to a new IMS procedure with an intrasolution magnetic particle transfer device, the PickPen. The IMS target for the majority of these studies is Escherichia coli O157:H7 in various types of samples, including cattle feces, hides, carcasses, and ground beef. Comparison of the two IMS methods showed a significant difference (P < 0.05) in the efficiency of detecting E. coli O157:H7 from cattle carcass surface, cattle hide, and cattle fecal samples. No significant improvement (P > 0.05) in E. coli O157:H7 detection was observed when the PickPen IMS procedure was used to isolate this pathogen from ground beef samples. Use of the PickPen IMS greatly increases the throughput of the IMS procedure and may be more compatible with various emerging technologies for pathogen detection. In addition, the efficacy of sequential IMS for multiple pathogens is reported herein.

A novel human detoxification system based on nanoscale bioengineering and magnetic separation techniques

We describe the conceptual approach, theoretical background and preliminary experimental data of a proposed platform technology for specific and rapid decorporation of blood-borne toxins from humans. The technology is designed for future emergent in-field or in-hospital detoxification of large numbers of biohazard-exposed victims; for example, after radiological attacks. The proposed systems is based on nanoscale technology employing biocompatible, superparamagnetic nanospheres, which are functionalized with target-specific antitoxin receptors, and freely circulate within the human blood stream after simple intravenous injection. Sequestration of the blood-borne toxins onto the nanosphere receptors generates circulating nanosphere-toxin complexes within a short time interval; mathematical modeling indicates prevailing of unbound nanosphere receptors over target toxin concentrations at most therapeutic injection dosages. After a toxin-specific time interval nanosphere-toxin complexes are generated within the blood stream and, after simple arterial or venous access, the blood is subsequently circulated via a small catheter through a portable high gradient magnetic separator device. In this device, the magnetic toxin complexes are retained by a high gradient magnetic field and the detoxified blood is then returned back to the blood circulation (extracorporeal circulation). Our preliminary in vitro experiments demonstrate >95% first pass capture efficiency of magnetic spheres within a prototype high gradient magnetic separation device. Further, based on the synthesis of novel hydrophobic magnetite nanophases with high magnetization ( approximately 55 emu/g), the first biodegradable magnetic nanospheres at a size range of approximately 280 nm and functionalized with PEG-maleimide surface groups for specific antibody attachment are described here. In future applications, we envision this technology to be suitable for emergent, in-field usage for acutely biohazard exposed victims as both the injectable toxin-binding magnetic spheres and the separator device are made to be portable, light-weight, zero-power, and self- or helper-employed. Details of the technology are presented and the state-of-knowledge and research is discussed.

Removal of As(V) and Cr(VI) Ions from Aqueous Solution using a Continuous, Hybrid Field‐Gradient Magnetic Separation Device

A continuous flow colloidal affinity magnetic separation device is used for the removal of As(V) and Cr(VI) from aqueous solutions. Langmuir isotherms fit the adsorption behavior of the individual ions on Orica MIEX® ion exchange particles. In a mixture of equal weight percent As(V) and Cr(VI), the adsorption of As(V) begins only above a critical cut‐off concentration, implying preferential adsorption of the higher valence ion at the available sites. Cr(VI) is removed selectively from the mixture in the continuous flow device, consistent with the presence of a higher concentration of the higher valence ion in the proximity of a charged (anion‐exchange) surface.

Continuous sorting of magnetic cells via on-chip free-flow magnetophoresis

The ability to separate living cells is an essential aspect of cell research. Magnetic cell separation methods are among some of the most efficient methods for bulk cell separation. With the development of microfluidic platforms within the biotechnology sector, the design of miniaturised magnetic cell sorters is desirable. Here, we report the continuous sorting of cells loaded with magnetic nanoparticles in a microfluidic magnetic separation device. Cells were passed through a microfluidic chamber and were deflected from the direction of flow by means of a magnetic field. Two types of cells were studied, mouse macrophages and human ovarian cancer cells (HeLa cells). The deflection was dependent on the magnetic moment and size of the cells as well as on the applied flow rate. The experimentally observed deflection matched well with calculations. Furthermore, the separation of magnetic and non-magnetic cells was demonstrated using the same microfluidic device.

Dual Functionality of the Anti-β1 Integrin Antibody, 12G10, Exemplifies Agonistic Signalling from the Ligand Binding Pocket of Integrin Adhesion Receptors

Although integrins are known to mediate connections between extracellular adhesion molecules and the intracellular actin cytoskeleton, the mechanisms that are responsible for coupling ligand binding to intracellular signaling, for generating diversity in signaling, and for determining the efficacy of integrin signaling in response to ligand engagement are largely unknown. By characterizing the class of anti-integrin monoclonal antibodies (mAbs) that stimulate integrin activation and ligand binding, we have identified integrin-ligand-mAb complexes that exhibit differential signaling properties. Specifically, addition of 12G10 mAb to cells adhering via integrin alpha4beta1 was found to trigger disruption of the actin cytoskeleton and prevent cell attachment and spreading, whereas mAb addition to cells adhering via alpha5beta1 stimulated all of these processes. In contrast, soluble ligand binding to either alpha4beta1 or alpha5beta1 was augmented or unaffected by 12G10. The regions of the integrin responsible for differential signaling were then mapped using chimeras. Surprisingly, a chimeric alpha5 integrin containing the beta-propeller domain from the ligand binding pocket of alpha4 exhibited the same signaling properties as the full-length alpha4 integrin, whereas exchanging or removing cytoplasmic domains had no effect. Thus the mAb 12G10 demonstrates dual functionality, inhibiting cell adhesion and spreading while augmenting soluble ligand binding, via a mechanism that is determined by the extracellular beta-propeller domain of the associating alpha-subunit. These findings therefore demonstrate a direct and variable agonistic link between the ligand binding pocket of integrins and the cell interior that is independent of the alpha cytoplasmic domains. We propose that either ligand-specific transmembrane conformational changes or ligand-specific differences in the kinetics of transmembrane domain separation underlie integrin agonism.

Immune Function Assay Enters the Clinical Laboratory

Methods for assessing a patient’s state of immune responsiveness has traditionally involved the use of radioactive markers to purified populations of lymphocytes after they have been stimulated. Briefly, a lymphocyte stimulant or mitogen such as phytohemagglutinin (PHA) is added to isolated lymphocytes. Lymphocytes in the peripheral blood express receptors on the cell surface, which bind to mitogens or specific antigens presented in conjunction with the major histocompatibility complex (MHC). Exposure to the mitogen or antigen results in activation and expansion of the lymphocyte subpopulation reactive to the antigen or mitogen. The suspension can then be labeled with a radioactive marker such as 3 H thymidine where DNA synthesis is measured on a scintillation counter. The drawbacks of this assay include a long turnaround time (72 hours) and the use of radioactive materials. A new assay, Immuknow by Cylex (Columbia, MD) was FDA approved in 2002 for measuring global cell mediated immunity. Heparinized whole blood is diluted in sample diluent and stimulated with the mitogen PHA. After an overnight incubation period at 37°C in 5% CO 2 , the CD4 lymphocyte population is isolated using a magnetic separation device and monoclonal antibodies to CD4. After a series of washes, the target cell population is lysed releasing cellular adenosine triphosphate (ATP), which is measured using a microplate luminometer and the luciferase-luciferin enzyme system. In the presence of luciferin and Mg-ATP, luciferase catalyzes a reaction producing oxyluciferin and light which is directly proportional to the ATP concentration 1 in the patient sample. An unstimulated control of patient sample is run in parallel, and the results are expressed as the change or delta between the stimulated and non-stimulated wells. A standard curve utilizing sequential ATP concentrations is incorporated in the test system. This assay has a 24-hour turnaround time, utilizes whole blood, and requires no radioactive materials.

Magnetic separation of iron and heavy metals from water

A magnetic separation device is being developed for removal of iron and heavy metals from water. The device consists of a column of supported magnetite surrounded by a movable permanent magnet. The mineral magnetite, or synthetically prepared iron ferrite (FeO x Fe2O3), is typically supported on various materials to permit adequate water passage through the column. In the presence of an external magnetic field, enhanced capacity was observed in using supported magnetite for removal of actinides and heavy metals from wastewater. The enhanced capacity is primarily due to magnetic filtration of colloidal and nanoscale particles along with some complex and ion exchange sorption mechanisms. This paper will review some previous work on the use of magnetite for wastewater treatment and discuss the development and potential of the magnetic nanoscale filtration/sorption process for water treatment. Recent research results are also presented on preliminary experimental studies using the process with water samples containing iron.

Continuous magnetic separation of blood components from whole blood

Direct magnetic separations of red blood cells from whole blood have been carried out using a continuous magnetic separation method based on high gradient magnetic separation (HGMS) and a gas-permeable membrane with nitrogen gas. The experimental results have shown good agreements with the theoretical model taking into account the gravitational force. Based on the analysis, the feasibility of a direct magnetic separation device for white blood cells and plasma from whole blood is discussed.

A Flow-Through, Hybrid Magnetic-Field-Gradient, Rotating Wall Device for Magnetic Colloidal Separations

A 2.0 cm internal diameter 1 m long, axially-rotating horizontal glass tube, with four axially located repeating hybrid magnetic units, is used as part of a flow-through, colloidal magnetic affinity separation device. Each magnetic unit consists of an alternating current solenoid surrounding the chamber followed by four azimuthally distributed permanent magnets that rotate with the chamber. Experiments were carried out on a model feed system consisting of a mixture of 1.0 μm diameter biotinylated latex beads (targets) and 9.7 μm diameter nonfunctionalized latex beads (nontargets) at a 1:1 number ratio. Streptavidin labeled magnetic particles (2.8 μm diameter polystyrene—Dynabeads) at a number concentration of ∼3 × 106 beads/mL were used as the separation vehicles. Two feed flow rates of 12 and 35 mL/min were used until a total of 600 mL of sample were processed for each. At the low rate we achieved a capture efficiency of 60 ± 4% and a separation factor of 18.2 ± 1.2 with 95 ± 4% purity. For the higher flow rate the capture efficiency was 40 ± 4% and the separation factor was 18.6 ± 1.5 with 87 ± 4% purity. A single stage batch process, where a total of 10 mL of feed was processed at identical feed and magnetic bead concentrations required ∼2 hours, produced a capture efficiency of 42 ± 4% and a separation factor of 3.8 ± 1 with 79 ± 4% purity. Three batch stages (2 hours processing time per stage) were required to surpass the capture efficiency of the flow-through device at the smaller flow rate. Thus, this flow-through separation device can lead to a large increase in processing volume and reduced “down” time, compared to current batch processes, without loss of either efficiency or purity, potentially opening up magnetic colloidal separations for large scale applications.

Organelle isolation by magnetic immunoabsorption.

Currently, most organelle isolation procedures rely on physical parameters and centrifugation for separation. Here, we report the rapid and gentle isolation of a variety of organelles by immunolabeling whole cell lysates with organelle-specific antibodies and streptavidin magnetic particles followed by separation in a magnetic field. Using magnetic immunoabsorption, we have been able to specifically label mouse metaphase chromosomes and a variety of plant organelles, including: amyloplasts, choroplasts and nuclei from whole cell lysates of various plant tissues. We find that the distinct magnetic properties, surface characteristics and mean diameter-size ranges of different particle preparations significantly influence their specific utility for organelle isolations. By using an internal-field magnetic separation device, we have developed a method for quantitative recovery of labeled organelles in microarrays and tested a variety of antibodies to chloroplast outer envelope proteins for their ability to immune-isolate chloroplasts.

Recent advances in the recovery of tungsten values in the fine and ultrafine size range

Recent advances in the design of fine gravity concentrators and high gradient magnetic separation devices have led to significant improvements in the recovery of tungsten values in fine and ultrafine size range. Amongst the more promising techniques of fine particles processing, encouraging results are reported on froth flotation, shear flocculation and spherical agglomeration of tungsten ore slimes. Development of highly selective reagents such as alkyl hydroxamates, phosphonic acid derivatives and alkylated nitrosonapthols is vital for enhancing separation efficiencies in the slimes size range. Froth flotation of Indian tungsten ore slimes (particularly at Degana) using appropriate reagent combinations appears promising and needs to be examined in greater detail.

An experimental study with a vortex magnetic separation (VMS) device

A vortex magnetic separation (VMS) system was designed and applied to the separation of a number of materials. An important feature is that the slurry passageways and the vortex magnetic capture regions are separately arranged in the device. The vortex traps are formed in the vortex magnetic capture regions under certain conditions so that magnetic particles can be driven into the vortex traps from the slurry and captured there. The VMS was used to treat an artificial mixture of wolframite and quartz and some real industrial ores. This paper presents some experimental results obtained. The results showed that the VMS can largely solve the mechanical entrapment problem encountered by the current practice of HGMS. VMS can be run at higher slurry velocities and so produce a higher output. The preliminary experiments carried out on industrial ores showed that this device can treat samples with a wide size range without blockage.

The traction force in magnetic separators

The theoretical expression for the traction force in magnetic separation devices is re-examined. It is argued that the formula usually used is correct only to lowest order in chi . A more accurate expression is presented, and its structure and order of magnitude are discussed. With the help of this more precise force formulation, the reliability of future modelling should increase considerably. In addition, the anisotropic features of the traction force open up the possibility of new separation methods.

Design of large-scale separation systems for positive and negative immunomagnetic selection of cells using superparamagnetic microspheres.

The ex vivo selective separation of cells from bone marrow and peripheral blood stem cell preparations is increasingly used as an adjunct to hematopoietic rescue following high-dose therapy for refractory cancer. Immunomagnetic separation, in which the target cells are identified using monoclonal antibodies and separated by attachment to paramagnetic particles and passage through a magnetic field, is widely used for both negative and positive cell selection. In this paper, we discuss the factors that should be considered when developing a magnetic separation device for purging tumor cells and selecting stem cells from bone marrow using superparamagnetic microspheres.

4710472 Magnetic separation device: Joseph W Saur, Charles P Reynolds, Alfred Black assigned to The United States of America as represented by the Secretary of the Navy

Abstract : This invention pertains to magnetic separation devices and more particularly to; magnetic separation devices used to selectively remove magnetic bead-coated cells from tissues such as bone marrow or blood. Depletion of cell populations from boner marrow has been mainly approached using antibodies conjugated to toxins such as ricin or with antibodies complement to effect lysis of the target cells. These techniques have severals disadvantages including difficulty in measuring the selective cell kill in the marrow, non-specific toxicity of either toxins or complement, and the necessity to prepare large amounts of complement. In addition, many antibodies are neither cytotoxic with complement nor toxin conjugates. Polymeric microspheres conjugated to antibodies have been used to probe the cell surface for receptor sites using scanning electron microscopy. Iron-containing polymeric microspheres tagged with fluroescent dyes conjugated to antibodies were used to separate red blood cells and lymphoid cells by binding the antibody-microsphere to selected cells and exposing the cell population to a magnetic field. Over 99% of the bound cells were attracted by the magnet.