US5328603A
Expired / Lapsed
US
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Lignocellulosic and cellulosic beads for use in affinity and immunoaffinity chromatography of high molecular weight proteins
Assignee: Virginia Tech Intellectual Properties Inc
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Filing: 1992-08-19
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Granted: 1994-07-12
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Expires: 2012-08-19
Abstract
Abstract Improved cellulosic beads for use as supports in bioaffinity chromatography are produced by dissolution of cellulose in a chaotropic cellulose solvent, formation of the dissolved cellulose into droplets, and immersion of the droplets into a non-solvent capable of solvent interchange with the cellulose solvent to form generally spherical porous cellulose beads of narrow particle size distribution. The beads formed are preferably made with cellulose having a degree of polymerization between 100 and 200, and the resulting beads when saturated with water without drying contain between 1% and 7% cellulose by weight and have a particle size of at least about 0.3 mm. The beads can be activated by a suitable activation method, and specific bioaffinity ligands are bound to the active sites in the beads. The beads reacted ligands, the beads then used in bioaffinity chromatography to isolate specific bioaffinity molecules having molecular weights between 5,000 and 500,000 from complex solutions. The beads are particularly useful in bioaffinity chromatography of antibodies, therapeutic proteins, enzymes, and other high molecular weight proteins. The cellulose beads have similar properties to agarose beads used in the prior art for bioaffinity chromatography of high molecular weight proteins, but the cellulosic beads of the present invention have much greater mechanical strength and resist crushing under higher column flow rates without chemical cross linking are much cheaper to produce than prior art agarose and dextran beads.
Inventors
- 👤 William H. Velander
- 👤 Jeffrey A. Kaster
- 👤 Wolfgang G. Glasser
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-11-09 03:03
CPC Classifications
B; B01; B01J; B01J20/00; B01J20/281; B01J20/291; B01J20/30; B01J20/32; B01J20/3202; B01J20/3206; B01J20/3208; B01J20/3212; B01J20/3214; B01J20/3217; B01J20/3219; B01J20/3225; B01J20/3227; B01J20/3231; B01J20/3242; B01J20/3268; B01J20/3272; B01J20/3274; B01J20/3287; B01J20/3291; B01J20/3293; B01J2220/00; B01J2220/50; B01J2220/54
Claims Analysis
Total Claims: 10
Claim 1
Independent Claim
163 words
Beads for use in affinity chromatography, comprising: from about 1% to about 7% of said beads being cellulose, said beads being sufficiently porous to allow molecules with molecular weights ranging from 5,000 to 5,000,000 to have access to their interiors, having a particle size greater than about 0.3 mm, and having sufficient strength to avoid crushing when used in chromatographic separation at linear flow rates above 20 cm/min in a 15 cm column bed; active sites on said cellulose; bioaffinity ligands attached to said active sites, said ligands having molecular weights up to about 500,000; and specific molecular recognition sites on said ligands, said molecular recognition sites capable of selectively attracting and binding specific bioaffinants, said bioaffinants having molecular weights ranging up to about 500,000; wherein, said pores are of sufficient size to allow the attachment of said ligands to said active sites on said cellulose without altering the capability of said specific molecular recognition sites to selectively attract and bind specific bioaffinants.
Claim 2
Dependent Claim
163 words
Beads for use in affinity chromatography, comprising: from about 1% to about 7% of said beads being cellulose, said beads being sufficiently porous to allow molecules with molecular weights ranging from 5,000 to 5,000,000 to have access to their interiors, having a particle size greater than about 0.3 mm, and having sufficient strength to avoid crushing when used in chromatographic separation at linear flow rates above 20 cm/min in a 15 cm column bed; active sites on said cellulose; bioaffinity ligands attached to said active sites, said ligands having molecular weights up to about 500,000; and specific molecular recognition sites on said ligands, said molecular recognition sites capable of selectively attracting and binding specific bioaffinants, said bioaffinants having molecular weights ranging up to about 500,000; wherein, said pores are of sufficient size to allow the attachment of said ligands to said active sites on said cellulose without altering the capability of said specific molecular recognition sites to selectively attract and bind specific bioaffinants.
Claim 3
Dependent Claim
24 words
Beads according to claim 1, wherein: said ligands and said bioaffinants are proteins selected from the group consisting of antibodies, therapeutic proteins, and enzymes.
Claim 4
Dependent Claim
26 words
Beads according to claim 2, wherein: said beads are of substantially spherical shape, and said cellulose has a degree of polymerization ranging less than about 200.
Claim 5
Dependent Claim
22 words
Beads according to claim 1, wherein: said beads are never dried, and are combined with an aqueous medium to form a gel.
Claim 6
Dependent Claim
18 words
Beads according to claim 1, wherein: said beads having a particle size of at least about 0.5 mm.
Claim 7
Dependent Claim
181 words
A chromatography support, comprising: highly porous cellulose beads which contain when saturated with water a content of cellulose between about 1% to 7% by weight, and a particle size greater than about 0.3 mm; said cellulose imparting to said beads sufficient strength to avoid crushing when used in chromatographic separation at linear flow rates above 20 cm/min in a 15 cm high bed, wherein said support is a bioaffinity support, active sites on said cellulose; bioaffinity ligands attached to said active sites, said bioaffinity ligands having molecular weights ranging up to about 500,000 said ligands having specific molecular recognition sites thereon to selectively attract and bind specific bioaffinants having molecular weights ranging up to about 500,000; and the pores of said porous beads being of sufficient size to allow the attachment of said ligands to said active sites on said cellulose without altering the capability of said molecular recognition sites in said ligands to selectively attract and bind specific bioaffinants and being sufficiently porous to allow molecules with molecular weights ranging from 5,000 to 5,000,000 to have access to their interiors.
Claim 8
Dependent Claim
181 words
A chromatography support, comprising: highly porous cellulose beads which contain when saturated with water a content of cellulose between about 1% to 7% by weight, and a particle size greater than about 0.3 mm; said cellulose imparting to said beads sufficient strength to avoid crushing when used in chromatographic separation at linear flow rates above 20 cm/min in a 15 cm high bed, wherein said support is a bioaffinity support, active sites on said cellulose; bioaffinity ligands attached to said active sites, said bioaffinity ligands having molecular weights ranging up to about 500,000 said ligands having specific molecular recognition sites thereon to selectively attract and bind specific bioaffinants having molecular weights ranging up to about 500,000; and the pores of said porous beads being of sufficient size to allow the attachment of said ligands to said active sites on said cellulose without altering the capability of said molecular recognition sites in said ligands to selectively attract and bind specific bioaffinants and being sufficiently porous to allow molecules with molecular weights ranging from 5,000 to 5,000,000 to have access to their interiors.
Claim 9
Dependent Claim
18 words
The chromatography support, of claim 6, wherein said beads have a particle size greater than about 0.5 mm.
Claim 10
Dependent Claim
22 words
The chromatorgraphy support of claim 6, wherein said bioaffiants are proteins selected from the group consisting of antibodies, therapeutic proteins, and enzymes.
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