Magnetic cell separation has become a popular technique to enrich or deplete cells of interest from a heterogeneous cell population. One important aspect of magnetic cell separation is the degree to which a cell binds paramagnetic material. It is this paramagnetic material that imparts a positive magnetophoretic mobility to the target cell, thus allowing effective cell separation. A mathematical relationship has been developed to correlate magnetic labeling to the magnetophoretic mobility of an immunomagnetically labeled cell. Four parameters have been identified that significantly affect magnetophoretic mobility of an immunomagnetically labeled cell: the antibody binding capacity (ABC) of a cell population, the secondary antibody amplification (psi), the particle-magnetic field interaction parameter (DeltachiV(m)), and the cell diameter (D(c)). The ranges of these parameters are calculated and presented along with how the parameters affect the minimum and maximum range of magnetophoretic mobility. A detailed understanding of these parameters allows predictions of cellular magnetophoretic mobilities and provides control of cell mobility through selection of antibodies and magnetic particle conjugates.

in Wiley InterScience (www.interscience.wiley.com).Mechanical forces are important signals in the development and function of the heart and lung, growth of skin and muscle, and maintenance of cartilage and bone. The specific mechanical force ''shear stress'' has been implicated as playing a critical role in the physiological responses of blood vessels through endothelial cell signaling. More recently, studies have shown that shear stress can induce differentiation of stem cells toward both endothelial and bone-producing cell phenotypes. This review will highlight current data supporting the role of shear stress in stem cell fate and will propose potential mechanisms and signaling cascades for transducing shear stress into a biological signal. V

Methods:A methodology and a mathematical theory have been developed, which allow quantitation of the expression levels of cellular surface antigens using immunomagnetic labels and cell tracking velocimetry (CTV) technology. Results: Quantum Simply Cellular (QSC) microbeads were immunomagnetically labeled with anti-CD2 fluorescein isothiocyanate (FITC) antibodies and anti-FITC MACS paramagnetic nanoparticles. Magnetophoretic mobility has been defined as the magnetically induced velocity of the labeled cell or microbead divided by the magnetophoretic driving force, proportional to the magnetic energy density gradient. Discussion: Using computer imaging and processing technology, the mobility measurements were accomplished by microscopically recording and calculating the velocity of immunomagnetically labeled QSC microbeads in a nearly constant magnetic energy gradient. A calibration curve correlating the measured magnetophoretic mobility of the immunomagnetically labeled microbeads to their antibody binding capacities (ABC) has been obtained. Conclusion:The results, in agreement with theory, indicate a linear relationship between magnetophoretic mobility and ABC for microbeads with less than 30,000 ABC. The mathematical relationships and QSC standardization curve obtained allow determination of the number of surface antigens on similarly immunomagnetically labeled cells. Cytometry 40:307-315, 2000.

Nano-and microscale topographical cues play critical roles in the induction and maintenance of various cellular functions, including morphology, adhesion, gene regulation, and communication. Recent studies indicate that structure and function at the heart tissue level is exquisitely sensitive to mechanical cues at the nano-scale as well as at the microscale level. Although fabrication methods exist for generating topographical features for cell culture, current techniques, especially those with nanoscale resolution, are typically complex, prohibitively expensive, and not accessible to most biology laboratories. Here, we present a tunable culture platform comprised of biomimetic wrinkles that simulate the heart's complex anisotropic and multiscale architecture for facile and robust cardiac cell alignment. We demonstrate the cellular and subcellular alignment of both neonatal mouse cardiomyocytes as well as those derived from human embryonic stem cells. By mimicking the fibrillar network of the extracellular matrix, this system enables monitoring of protein localization in real time and therefore the high-resolution study of phenotypic and physiologic responses to in-vivo like topographical cues.

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Antibody binding capacity (ABC) is a term representing a cell's ability to bind antibodies, correlating with the number of specific cellular antigens expressed on that cell. ABC allows magnetically conjugated antibodies to bind to the targeted cells, imparting a magnetophoretic mobility on each targeted cell. This enables sorting based on differences in the cell magnetophoretic mobility and, potentially, a magnetic separation based on the differences in the cell ABC values. A cell's ABC value is a particularly important factor in continuous magnetic cell separation. This work investigates the relationship between ABC and magnetic cell separation efficiency by injection of a suspension of immunomagnetically labeled quantum simply cellular calibration microbeads of known ABC values into fluid flowing through a quadrupole magnetic sorter. The elution profiles of the outlet streams were evaluated using UV detectors. Optimal separation flow rate was shown to correlate with the ABC of these microbeads. Comparing experimental and theoretical results, the theory correctly predicted maximum separation flow rates but overestimated the separation fractional recoveries.

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Embryonic stem (ES) cells serve as an excellent in vitro system for studying differentiation events and for developing methods of generating various specialized cells for future regenerative therapeutic applications. Two obstacles associated with using embryonic stem cells include (a) isolating homogeneous populations of differentiated cells and (b) obtaining terminally differentiated cell populations that are capable of proliferating further. Here, the authors describe methods in which they have overcome these two obstacles by generating highly purified populations (>96%) of actively proliferating endothelial cells from mouse ES cells. Briefly, 60,000 ES cells progress through three different stages of cell induction/expansion and two cell isolation procedures, generating over 300 million endothelial cells. These ES-derived endothelial cells display characteristics similar to vascular endothelial cells in that they express several common endothelial markers, they form two-dimensional (2D) tubelike structures as well as complex microvessels in three-dimensional (3D) collagen type I gels, and they retain the ability to reorganize their cytoskeleton in response to mechanical forces. Our findings indicate that it is possible to obtain proliferating populations of homogeneous endothelial cells from mouse ES cells without genetically manipulating the ES cells or coculturing with feeder cells.