Stem cells are defined as cells that are capable of indefinite proliferation as well as differentiation into specialized tissue under suitable conditions. Due to their unlimited potential for tissue regeneration and disease treatment, stem cell research has secured great prominence over the past decade. Identification of specific markers defining neural and hematopoietic stem cells has led to the development of products to isolate distinct stem cell populations.
CD133, a five trans-membrane domain glycoprotein is considered to be a marker for hematopoietic and neural stem cells. Besides, CD133 is also known to be expressed by cancer stem cells capable of initiating tumors in various host species. Multipotent CD133+ cells have been isolated from neural tissue and differentiated into neurons and glia by several laboratories. Isolation of pure populations of stem cells from tissues or cell lines is important for research and development in the stem cell arena.
Miltenyi biotec have developed the CD133 MicroBead Kit for positive isolation of CD133 expressing cells from a variety of tissues and cell populations, including peripheral mononuclear cells, cord blood cells, bone marrow cells and cells from fetal brain tissue. The kit contains magnetic microbeads labeled with anti-CD133 antibodies and an FcR blocking agent to block non-specific binding. The protocol for isolation is simple and is not time consuming. The cells are first magnetically labeled with CD133 microbeads by simply incubating the cells with the magnetic beads in the presence of the blocking agent for 30 min at 4°C. Subsequently, after a few washes, the CD133+ cells are separated by passing the cell suspension through a column placed in the magnetic field of a MACS cell separator. Once the unlabelled CD133 negative cells flow through, the column is removed from the magnetic field and the CD133+ cells are flushed out. Nearly 85-95% of the cells isolated by this method express CD133. An additional advantage of using this method of isolation is that relatively pure and viable populations of both CD133+ and CD133- cells can be isolated and used for comparative studies.
The CD133 MicroBead Kit has been used in our laboratory for over 4 years, for isolation of neural stem cells from fetal human brains. We have also been able to isolate CD133 expressing stem-like cells from neuroblastoma cell lines SK-N-SH and SK-N-BE. The attractive feature of this kit is the relative ease with which a reasonably pure population of CD133 expressing cells can be isolated from a heterogeneous cell suspension. Despite the trauma of flushing out the CD133+ cells, cell viability does not appear to be compromised. The kit does have certain limitations. To use this kit effectively, it is necessary to purchase magnetic cell separators MiniMACS, MidiMACS or SuperMACS and positive selections columns, both of which are supplied by Miltenyi Biotec. The company also offers a PE conjugated CD133 antibody for flow cytometric or fluorescence microscopic verification of the isolated cells.
In summary, the CD133 MicroBead Kit from Miltenyi Biotec is an excellent tool for isolation of CD133 expressing stem cells from various tissues. An initial expense towards purchase of cell separators is well justified. Besides, the company also has excellent technical and sales support.