Effects of different quantities of antibody conjugated with magnetic nanoparticles on cell separation efficiency

Muscle precursor cells may act not only as a means of inserting normal genes into diseased muscle fibres, in order to correct or alleviate a genetically inherited myopathy, but recent demonstrations have shown they may prove an invaluable tool for the expression of, and systemic dissemination of, non-muscle gene products. If muscle precursor cells are proved to act as such widespread vectors in terms of gene therapy, then it is imperative that methods are properly elucidated to produce large populations of pure viable myogenic cells for such purposes. In the past, many methods of cell separation have been investigated but carry with them the problems of either a lack of myogenic purity of the population or poor percentage recovery of the original cell population. In the present work we have investigated two methods for segregating myogenic from non-myogenic cells and have critically reviewed the efficiency of separation of the two techniques used. To obtain a quantitative measure of separation efficiency, segregation was carried out on a 1:1 mixture of murine C2 myogenic and murine 3T3 fibroblastic cells. To distinguish between C2 and 3T3 cells, the latter were prelabelled with the fluorescent strain carboxyfluorescein diacetate succinimyl ester (CFSE). Once incorporated into the cell, CFSE remains there, thus preventing transfer of the label to C2 cells. Both methods of separation used depend on the affinity of myogenic cells for the monoclonal antibody Mab H28, which specifically binds to the mouse neuronal cell adhesion molecule N-CAM, but differ in that one method, “panning”, completes segregation by adherence of N-CAM positive cells to a dish precoated with secondary IgG antibody whereas in the other separation proceeds by the use of commercially available IgG-coated magnetic beads. Results indicate magnetic bead separation to be more efficient than panning if the beads are precoated with 0.1% gelatin.

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Design and Simulation of an Integrated Centrifugal Microfluidic Device for CTCs Separation and Cell Lysis

Micromachines ◽

10.3390/mi11070699 ◽

2020 ◽

Vol 11 (7) ◽

pp. 699

Author(s):

Rohollah Nasiri ◽

Amir Shamloo ◽

Javad Akbari ◽

Peyton Tebon ◽

Mehmet R. Dokmeci ◽

...

Keyword(s):

Numerical Simulation ◽

Angular Velocity ◽

Microfluidic Device ◽

Cell Separation ◽

High Efficiency ◽

Separation Efficiency ◽

White Blood Cells ◽

Cell Lysis ◽

Target Cells ◽

Separation Unit

Separation of circulating tumor cells (CTCs) from blood samples and subsequent DNA extraction from these cells play a crucial role in cancer research and drug discovery. Microfluidics is a versatile technology that has been applied to create niche solutions to biomedical applications, such as cell separation and mixing, droplet generation, bioprinting, and organs on a chip. Centrifugal microfluidic biochips created on compact disks show great potential in processing biological samples for point of care diagnostics. This study investigates the design and numerical simulation of an integrated microfluidic device, including a cell separation unit for isolating CTCs from a blood sample and a micromixer unit for cell lysis on a rotating disk platform. For this purpose, an inertial microfluidic device was designed for the separation of target cells by using contraction–expansion microchannel arrays. Additionally, a micromixer was incorporated to mix separated target cells with the cell lysis chemical reagent to dissolve their membranes to facilitate further assays. Our numerical simulation approach was validated for both cell separation and micromixer units and corroborates existing experimental results. In the first compartment of the proposed device (cell separation unit), several simulations were performed at different angular velocities from 500 rpm to 3000 rpm to find the optimum angular velocity for maximum separation efficiency. By using the proposed inertial separation approach, CTCs, were successfully separated from white blood cells (WBCs) with high efficiency (~90%) at an angular velocity of 2000 rpm. Furthermore, a serpentine channel with rectangular obstacles was designed to achieve a highly efficient micromixer unit with high mixing quality (~98%) for isolated CTCs lysis at 2000 rpm.

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Functionalized Magnetic Nanoparticles for the Detection and Quantitative Analysis of Cell Surface Antigen

BioMed Research International ◽

10.1155/2013/349408 ◽

2013 ◽

Vol 2013 ◽

pp. 1-9 ◽

Cited By ~ 14

Author(s):

Daryoush Shahbazi-Gahrouei ◽

Mohammad Abdolahi ◽

Sayyed Hamid Zarkesh-Esfahani ◽

Sophie Laurent ◽

Corine Sermeus ◽

...

Keyword(s):

Magnetic Nanoparticles ◽

Cell Surface ◽

Surface Antigen ◽

Cell Separation ◽

Cell Surface Antigen ◽

Cell Surface Antigens ◽

Vitro Assay ◽

Gold Standard Method ◽

Magnetic Cell Separation

Cell surface antigens as biomarkers offer tremendous potential for early diagnosis, prognosis, and therapeutic response in a variety of diseases such as cancers. In this research, a simple, rapid, accurate, inexpensive, and easily available in vitro assay based on magnetic nanoparticles and magnetic cell separation principle was applied to identify and quantitatively analyze the cell surface antigen expression in the case of prostate cancer cells. Comparing the capability of the assay with flow cytometry as a gold standard method showed similar results. The results showed that the antigen-specific magnetic cell separation with antibody-coated magnetic nanoparticles has high potential for quantitative cell surface antigen detection and analysis.

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Droplet-based magnetically activated cell separation: analysis of separation efficiency based on the variation of flow-induced circulation in a pendent drop

Analytical and Bioanalytical Chemistry ◽

10.1007/s00216-009-3131-z ◽

2009 ◽

Vol 395 (7) ◽

pp. 2415-2421 ◽

Cited By ~ 2

Author(s):

Youngho Kim ◽

Sang Ho Lee ◽

Byungkyu Kim

Keyword(s):

Cell Separation ◽

Separation Efficiency ◽

Pendent Drop

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3D Printed Microfluidic Spiral Separation Device for Continuous, Pulsation-Free and Controllable CHO Cell Retention

Micromachines ◽

10.3390/mi12091060 ◽

2021 ◽

Vol 12 (9) ◽

pp. 1060

Author(s):

Anton Enders ◽

John-Alexander Preuss ◽

Janina Bahnemann

Keyword(s):

Cell Separation ◽

Separation Efficiency ◽

Chinese Hamster Ovary Cells ◽

Chinese Hamster ◽

Optimal Operation ◽

Dead Volume ◽

Cell Retention ◽

Separation Unit ◽

Peristaltic Pumps ◽

3D Printed

The development of continuous bioprocesses—which require cell retention systems in order to enable longer cultivation durations—is a primary focus in the field of modern process development. The flow environment of microfluidic systems enables the granular manipulation of particles (to allow for greater focusing in specific channel regions), which in turn facilitates the development of small continuous cell separation systems. However, previously published systems did not allow for separation control. Additionally, the focusing effect of these systems requires constant, pulsation-free flow for optimal operation, which cannot be achieved using ordinary peristaltic pumps. As described in this paper, a 3D printed cell separation spiral for CHO-K1 (Chinese hamster ovary) cells was developed and evaluated optically and with cell experiments. It demonstrated a high separation efficiency of over 95% at up to 20 × 106 cells mL−1. Control over inlet and outlet flow rates allowed the operator to adjust the separation efficiency of the device while in use—thereby enabling fine control over cell concentration in the attached bioreactors. In addition, miniaturized 3D printed buffer devices were developed that can be easily attached directly to the separation unit for usage with peristaltic pumps while simultaneously almost eradicating pump pulsations. These custom pulsation dampeners were closely integrated with the separator spiral lowering the overall dead volume of the system. The entire device can be flexibly connected directly to bioreactors, allowing continuous, pulsation-free cell retention and process operation.

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Sheathless High-Throughput Circulating Tumor Cell Separation Using Viscoelastic non-Newtonian Fluid

Micromachines ◽

10.3390/mi10070462 ◽

2019 ◽

Vol 10 (7) ◽

pp. 462 ◽

Cited By ~ 8

Author(s):

Hyunjung Lim ◽

Seung Min Back ◽

Min Ho Hwang ◽

Dae-Hee Lee ◽

Hyuk Choi ◽

...

Keyword(s):

High Throughput ◽

Blood Cells ◽

Cell Separation ◽

Separation Efficiency ◽

White Blood Cells ◽

Flow Characteristics ◽

Circulating Tumor Cell ◽

Size Difference ◽

Biological Applications ◽

Mcf 7

Circulating tumor cells (CTCs) have attracted increasing attention as important biomarkers for clinical and biological applications. Several microfluidic approaches have been demonstrated to separate CTCs using immunoaffinity or size difference from other blood cells. This study demonstrates a sheathless, high-throughput separation of CTCs from white blood cells (WBCs) using a viscoelastic fluid. To determine the fluid viscoelasticity and the flow rate for CTC separation, and to validate the device performance, flow characteristics of 6, 13, and 27 μm particles in viscoelastic fluids with various concentrations were estimated at different flow rates. Using 0.2% hyaluronic acid (HA) solution, MCF-7 (Michigan Cancer Foundation-7) cells mimicking CTCs in this study were successfully separated from WBCs at 500 μL/min with a separation efficiency of 94.8%. Small amounts of MCF-7 cells (~5.2%) were found at the center outlet due to the size overlap with WBCs.

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Analysis of cell separation efficiency in dielectrophoresis-activated cell sorter

2008 3rd IEEE International Conference on Nano/Micro Engineered and Molecular Systems ◽

10.1109/nems.2008.4484482 ◽

2008 ◽

Author(s):

Jaemin An ◽

Jangwon Lee ◽

Youngho Kim ◽

Byungkyu Kim ◽

Sangho Lee

Keyword(s):

Cell Separation ◽

Separation Efficiency ◽

Cell Sorter

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Cationized Magnetoferritin Enables Rapid Labeling and Concentration of Gram-Positive and Gram-Negative Bacteria in Magnetic Cell Separation Columns

Applied and Environmental Microbiology ◽

10.1128/aem.00720-16 ◽

2016 ◽

Vol 82 (12) ◽

pp. 3599-3604 ◽

Cited By ~ 3

Author(s):

S. Correia Carreira ◽

J. Spencer ◽

W. Schwarzacher ◽

A. M. Seddon

Keyword(s):

Magnetic Nanoparticles ◽

Cell Separation ◽

Gram Negative Bacteria ◽

Gram Positive ◽

Gram Negative ◽

Content Type ◽

E Coli ◽

Magnetic Cell Separation ◽

Magnetic Labeling ◽

Separation Columns

ABSTRACTIn order to identify pathogens rapidly and reliably, bacterial capture and concentration from large sample volumes into smaller ones are often required. Magnetic labeling and capture of bacteria using a magnetic field hold great promise for achieving this goal, but the current protocols have poor capture efficiency. Here, we present a rapid and highly efficient approach to magnetic labeling and capture of both Gram-negative (Escherichia coli) and Gram-positive (Staphylococcus aureus) bacteria using cationized magnetoferritin (cat-MF). Magnetic labeling was achieved within a 1-min incubation period with cat-MF, and 99.97% of the labeled bacteria were immobilized in commercially available magnetic cell separation (MACS) columns. Longer incubation times led to more efficient capture, withS. aureusbeing immobilized to a greater extent thanE. coli. Finally, low numbers of magnetically labeledE. colibacteria (<100 CFU per ml) were immobilized with 100% efficiency and concentrated 7-fold within 15 min. Therefore, our study provides a novel protocol for rapid and highly efficient magnetic labeling, capture, and concentration of both Gram-positive and Gram-negative bacteria.IMPORTANCEAntimicrobial resistance (AMR) is a significant global challenge. Rapid identification of pathogens will retard the spread of AMR by enabling targeted treatment with suitable agents and by reducing inappropriate antimicrobial use. Rapid detection methods based on microfluidic devices require that bacteria are concentrated from large volumes into much smaller ones. Concentration of bacteria is also important to detect low numbers of pathogens with confidence. Here, we demonstrate that magnetic separation columns capture small amounts of bacteria with 100% efficiency. Rapid magnetization was achieved by exposing bacteria to cationic magnetic nanoparticles, and magnetized bacteria were concentrated 7-fold inside the column. Thus, bacterial capture and concentration were achieved within 15 min. This approach could be extended to encompass the capture and concentration of specific pathogens, for example, by functionalizing magnetic nanoparticles with antibodies or small molecule probes.

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Synthesis of orientedly bioconjugated core/shell Fe3O4@Au magnetic nanoparticles for cell separation

Talanta ◽

10.1016/j.talanta.2011.05.010 ◽

2011 ◽

Vol 85 (3) ◽

pp. 1246-1252 ◽

Cited By ~ 67

Author(s):

Yi-Ran Cui ◽

Chao Hong ◽

Ying-Lin Zhou ◽

Yue Li ◽

Xiao-Ming Gao ◽

...

Keyword(s):

Magnetic Nanoparticles ◽

Cell Separation ◽

Core Shell

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The Selective Protein Separations with Polyaminofunctionality on Controlled Silica Coating-Layers of Magnetic Nanoparticles

Solid State Phenomena ◽

10.4028/www.scientific.net/ssp.124-126.903 ◽

2007 ◽

Vol 124-126 ◽

pp. 903-906 ◽

Cited By ~ 1

Author(s):

Moo Eon Park ◽

Ki Ho Kang ◽

Kyung Ja Kim ◽

Jeong Ho Chang

Keyword(s):

Magnetic Nanoparticles ◽

Protein Separation ◽

Separation Efficiency ◽

Sol Gel ◽

Silica Coating ◽

Chemical Precipitation ◽

Molar Ratio ◽

Basic Solution ◽

Particle Sizes ◽

Protein Separations

This work reported the development of the high throughput protein separation process with molecularly assembled silica-coated magnetic nanoparticles as a function of amino group numbers such as mono-, di-, and tri-aminofunctionality, in which the range of silica coating thicknesses were optimized to be interacted with protein. The protein separation efficiency was demonstrated as a function of each aminofunctional group and the particle sizes of the silica coated magnetic nanoparticles. The particles were prepared by the chemical precipitation of Fe2+ and Fe3+ salts with a molar ratio of 1:2 under basic solution. The silica coated magnetic nanoparticles were directly produced by the sol-gel reaction of a tetraethyl orthosilicate (TEOS) precursor, in which the coating layer serves as a biocompatible and versatile group for further biomolecular functionalization. To effectively capture the proteins, silica coated magnetic nanoparticles need to be functionalized reproducibly on the silica surface, and three kinds of amino functional groups were adapted as a function of number of amine using the mono-, di-, and tri-aminopropylalkoxysilanes.

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