Erprobung eines neuen Verfahrens zur Selektion von Stammzellen aus peripherem Blut
LicenseStandard (Fassung vom 03.05.2003)
The characteristics of Polyacrolein Microspheres (PAMS) are examined for their applicability to cell selection. Material: Polyacrolein is a polymer which binds covalently to protein. It was found that antibody covalently bound to PAMS was able to bind secondary antibody by means of a reversible antigen antibody binding. In the past PAMS was clinically applied for purging of blood in antibody mediated diseases. There was reason to assume that the method is also applicable to cell selection. PAMS were provided in a spheric shape of 0,7 µm diameter covalently bound to glass capillaries. Methods: A considerable number of experiments was performed to institute optimal conditions for cell selection. Thereby four main lines of examination were followed: 1. minimization of unspecific binding by incubation of capillaries in protein solution (BSA 0,5 % for 12 h); 2. optimizing the constitutes of the cell suspension (10 mio/ml of mononuclear cells , thrombocyte reducing preparation, adding of EDTA); 3. constructing an incubation system with laminar flow conditions and visualizing the result; 4. testing for the best method to release the adherent cells from the capillaries (mechanic means - by inducing turbulent flow along the capillary surface, chemical means - by reducing pH to 3 or by using 0,1 % of detergents solution). Results: 1. Unspecific binding of antibodies and cells was satisfactorily blocked. - 2. Cluster formation and clotting in the cell suspensions and on the capillary surface could be eliminated. - 3. Adherence of target cells to PAMS capillaries reached sufficient numbers. Cell adhesion follows a gravitational pattern. - 4. None of the tested releasing methods yielded satisfactory cell numbers in suspension. Conclusion: Specifically bound target cells are more adherent to the PAMS capillaries than explained by an antigen antibody binding. Positive selection could not be shown. PAMS are not suitable for cell selection. The reasons are discussed in detail.
Subject HeadingsKnochenmarktransplantation [GND]
Bone marrow transplantation [MeSH]