An Overview to Cell Selection
What is Cell Selection?
Cell Selection, also known as cell separation, cell isolation, or cell sorting, is the process of selectively separating a desired cell species from a collection of cells: usually from blood, tissue, or from a processed sample such as Apheresis or a Leukopak. Any cell species can be targeted, from T cells (CD3, CD4, CD8) to B cells, monocytes, stem cells and more, depending on the desired product and downstream plans for the cells.
Applications for Cell Selection
Cell Biology Research
The study of the inner mechanics, kinetics, signaling and the immune response cells can give, among many other research topics.
Cell & Gene Therapy (CGT)
The clinical process of transferring live cells into a patient in the hope that the healthy, genetic engineered cells will help the patient's to restore a normal function or fight in a disease. These cells can be cells previously taken and expanded from the patient (autologous cells), or can be from a donor who has an HLA (human leukocyte antigen) match to the patient (allogeneic cells). Common cells isolated for use in CGT are stem cells (embryonic, hematopoietic, epithelial, etc) or immune cells (T cells, B cells, natural killer cells, etc).1
CAR-T Cell Therapy
The process of taking the patient's own T-cells and modifying them to express a surface receptor (CAR: chimeric antigen receptor) to recognize antigens on the surface of malignant cells in the patient. Once the modified T cell's receptor binds to a tumor antigen, the immune response begins, and the T-cell will attack the malignant cells.1
The process of isolating cells at high purity and expanding them to produce a high number of purified cells of the same species.
The practice of manufacturing drugs (reagent) that can be approved to use in clinical conditions on the human body. For example: CellSearch CTC©, which identifies and selects tumor cells in the blood for early detection of cancer.
Principal Methods of Cell Selection
Immunomagnetic Cell Separation
Immunomagnetic Cell Separation uses a magnetic or magnetically coated particle or bead to bind to cells, and then uses a magnet to separate out the magnetically labeled cells.
Fluorescence-Activated Cell Sorting (FACS)
Fluorescence activated Cell Sorting uses fluorophore-conjugated antibodies to label target cells. A laser in a flow cytometer is used to excite the fluorophores on the individual cell in each drop labeling it with a charge. Deflection plates are then used to separate cells of similar charges into collection tubes.
Buoyancy Activated Cell Sorting (BACS)
Buoyancy Activated Cell Sorting binds microbubbles with a density < 1 g/mL to target cells. The solution is then centrifuged and targeted cells float to the top with the microbubbles, while non-targeted cells pellet at the bottom of the tube. The targeted cells can then be aspirated off the solution.
Filtration uses filters with specific pore sizes to isolate cells by size. Sequential filtration with filters of decreasing size can increase the specificity of isolation.
Aptamer Based Cell Isolation
Aptamer Based Cell Sorting uses aptamers (single-stranded DNA/RNA molecules that fold into a functional form) to bind to the target cell species instead of antibodies. These aptamers are produced by an exponential enrichment procedure known as SELEX (Systematic Evolution of Ligands by Exponential enrichment). These aptamers can then be bound to a separation body such as a bead or a bubble for target cell isolation.
Density Gradient Centrifugation
Density Gradient Centrifugation involves carefully layering your biological sample over a dense substance, centrifuging the sample, and isolating cells by density. Typically used to remove dead cells and to separate RBCs from plasma
Immunodensity Cell Separation
Immunodensity Cell Separation (Erythrocyte Rosetting) is a negative selection technique with whole blood where antibodies specific to unwanted cells are added to the blood. These antibody labeled cells then cross-link with RBCs, forming immunorosettes of high density. The whole blood is layered over a dense solution as in Density Gradient Centrifugation, and after centrifuging the unwanted cells are pelleted at the bottom of the tube with the RBCs, while the desired cells are suspended above the density medium2.
Sedimentation uses centrifugation to separate cells by their sedimentation rate, which is determined by their size and their density. Successive centrifugations increasing in centrifugation force or time are used to isolate the desired cells from the total population.
Adhesion uses the properties of certain cell species to adhere to a specific surface suitable for those properties. If the adhesion surface is a petri dish coated with serum for example, the cell mixture is poured into the petri dish, and only adherent cells will stick to the dish, while non-adherent cells will be poured out.
Microfluidic Cell Separation
Microfluidic Cell Separation encompasses a vast variety of technologies that work at the microfluidic level to isolate cells, even to the scale that several of the technologies work on a single-cell isolation. Many of the technologies build their isolation systems on a microchip, which allows the sample size to be minuscule and the device to be portable.
Methods of Immunomagnetic Separation
The target cell of the isolation is labeled with an antibody specific to the receptors on the cell. That antibody is then targeted by the separating body (i.e. FF), and the target cell is isolated.
● Inexpensive – only need a target antibody
● High purity – Targeting one type of cell
● Premature activation of immune cells reduces downstream applications
The target cell remains unlabeled (untouched), while other cell types are labeled with antibodies specific to the receptors on their cell surfaces. Those antibodies are then targeted by a separating body and removed, leaving the target cell behind.
● Produces untouched target cells
● Quicker protocol
● Typically more expensive – need more antibodies
● Lower purities – More cells to remove
Why should you choose XpresSepTM Ferrofluid?
● Colloidal sized Ferrofluids mean no continuous mixing, and no need for columns!
● Short incubation times –> quick separations!
● Separation leaves cells with high viability!
● Target cell purities above 90%!
1. For more information on Gene and Cell Therapy, read https://www.asgct.org/education/more-resources/gene-and-cell-therapy-faqs
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