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Driver Bess Um100 Para Windows 7.Cell separation methods, such as positive or negative selection and fluorescence-activated cell sorting (FACS), can be used to purify a desired subset of cells from a mixed population of cells. For example, a cell sample may be stained to allow separation of different cell populations based on the presence or absence of a particular antigen, or binding of a particular monoclonal antibody. Separation of desired cells is often based on the combination of a physical characteristic of the cells (e.g., size or density) and a marker antigen or antibody bound to a second physical property (e.g., fluorescence or biotin).
A sorting machine typically consists of a container, or well, in which the cells are loaded, a feed tube or tube through which the cells are allowed to flow, and a nozzle from which the cells are ejected. Before the cells enter the machine, they may be treated with a sample of a fluorochrome-conjugated antibody that binds to a cell surface antigen of interest.
The cell suspension is fed into the container (also known as a sorter well) by pumping or aspirating. When the cells enter the well, they are separated based on one or more physical characteristics. Common characteristics used are density and size (FACS) and immunofluorescence.
Using negative selection, cells are removed from a well using a magnetic force, leaving the desired cells in the well. By contrast, in positive selection, the fluorescently labeled cells are removed from the well, leaving the desired cells behind.
In negative selection, magnetic particles are mixed with the cell suspension, and the mixture is loaded into the sorter well. The cells adhere to the magnetic particles, which are magnetically pulled out of the well.
The unwanted cells are removed from the well by pipetting them out. After removing the unwanted cells, the desired cells remain in the well for further processing. Positive selection is similar except that the magnetic particles and the unwanted cells are removed from the well.
In FACS, cells are sorted based on the presence or absence of a specific marker. These cells are labeled with
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