Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving antigen-antibody binding – specific binding protein...
Patent
1996-08-06
1998-06-02
Saunders, David
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving antigen-antibody binding, specific binding protein...
210660, 210661, 210695, 435 2, 435 30, 435325, 435369, 435370, 435372, 436526, C12N 500, C12Q 124
Patent
active
057597936
DESCRIPTION:
BRIEF SUMMARY
TECHNICAL FIELD
This invention relates to the use of a magnetically stabilized fluidized bed (MSFB) for selective separation and purification of mammalian cell populations.
BACKGROUND ART
Separation of mixtures of chemicals, biomolecules and cell types is often effected by immunoaffinity chromatography. Packed beds, such as those used in column chromatography, are often used in affinity separation. However, problems such as nonspecific trapping or filtration of cells and clogging make the use of a packed bed undesirable for cell separation. In addition, when fine particles are used to increase the mass transfer efficiency of packed beds, a large pressure drop across the bed often results. These problems require significant washing of the packed bed in order to flush contaminants and other cellular debris from the column.
One device that has been developed for reducing the pressure drop across a column of particles is the fluidized bed. A fluidized bed consists of solid particles and a gas or liquid which is passed upwardly through the particle bed with velocity sufficient to ensure that the drag forces of the fluid counterbalance the gravitational forces on the particle and cause random motion of the particles. The bed of particles will become fluidized and expand, resulting in a lower pressure drop across the fluidized bed as compared to the pressure drop across a packed bed of the same height. The fluidization of the bed also provides more surface contact between the particle and the fluid passing through the bed.
One disadvantage associated with fluidized beds is the radial and axial movement of the particles which result in significant intermixing of the particles. An advancement in fluidized bed technology is the magnetically stabilized fluidized bed (MSFB) which involves the use of a magnetic field and magnetizable particles to stabilize the bed. It has been found that, by supplying a magnetic field parallel to the path of fluid flow, the magnetizable particles can be locked in place, thus eliminating the intermixing of the particles.
In general, the MSFB combines some of the best characteristics of the fluidized bed with those of a fixed bed. More particularly, the MSFB provides a low pressure drop, the ability to transport solids through a system and good mass transfer driving force even as the fluid is depleted of its source. See Burns, Structural Studies of a Liquid-Fluidized Magnetically Stabilized Bed, Chem. Eng. Comm., 67:315-330 (1988). All documents cited herein are hereby incorporated by reference.
Because of these advantages, MSFBs have been used to separate various chemical species and proteins, and to filter yeast. For example, for the use of a MSFB to separate proteins see Burns, et al., Application of Magnetically Stabilized Fluidized Beds to Bioseparations, Reactive Polymers, 6:45-50 (1987) (human serum albumin); Lochmuller et al., Affinity Separations in Magnetically Stabilized Fluidized Beds: Synthesis and Performance of Packing Materials, Separation Science and Technology; 22:2111-2125 (1987) (trypsin); and U.S. Pat. No. 5,130,027, issued to Noble, Jul. 14, 1992, (Cytochrome-C). The use of a MSFB to separate various organic and inorganic compounds is discussed in U.S. Pat. No. 5,084,184 issued to Burns on Jan. 28, 1992. Finally, the use of an MSFB as a filter to collect yeast cells was reported by Terranova et al. Continuous Cell Suspension Processing using Magnetically Stabilized Fluidized Beds, Biotechnology and Bioengineering, 37:110-120 (1991). In this latter reference, the filtration was not based on immunoaffinity but rather on electrostatic interaction between the positively charged nickel particles contained in the MSFB and the negatively charged yeast cells.
However, none of these references discuss the use of the MSFB for the affinity separation of mammalian cell population from a mixture of cell populations. The separation of a particular mammalian cell population from a mixture of cell populations is quite different from the separation of chemical species such as pr
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Elkalay Mohammed A.
Schwartz Richard M.
Saunders David
Systemix, Inc.
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