Claims
- 1. A process for selectively isolating IgY antibodies from egg yolk of an anseriform bird, comprising:
(a) absorbing yolk antibodies in a water-miscible fraction obtained from the egg yolk of an anseriform bird with a water insoluble non-charged absorbent selected from the group consisting of silicate, silicon compounds, carbonate, sulfate, phosphate, carbon, cellulose and synthetic fiber, ceramics, and metal oxide, and wherein the water insoluble non-charged absorbent is at an amount effective for separating the yolk antibodies from the water-miscible fraction; and (b) flowing the water insoluble non-charged absorbent with a buffer to obtain an aqueous fraction containing the yolk antibodies, wherein the IgY antibodies comprise antibodies with Fe regions and antibodies that lack Fe regions.
- 2. The process of claim 1, wherein the water insoluble non-charged absorbent is a silicate selected from the group consisting of synthetic or natural clays, kaolin, talc, and calcium silicate.
- 3. The process of claim 1, wherein the water insoluble non-charged absorbent is a silicon compound selected from the group consisting of fumed silica, amorphous silica, silica dioxide, silica gel, silicates, diatomaceous earth, and Fuller's earth.
- 4. The process of claim 1, wherein the water insoluble non-charged absorbent is comprised of calcium carbonate or barium carbonate.
- 5. The process of claim 1, wherein the water insoluble non-charged absorbent is comprised of calcium sulfate.
- 6. The process of claim 1, wherein the water insoluble non-charged absorbent is comprised of calcium phosphate.
- 7. The process of claim 1, wherein the water insoluble non-charged absorbent is comprised of activated carbon or carbon fiber.
- 8. The process of claim 1, wherein the water insoluble non-charged absorbent is comprised of cellulose powder.
- 9. The process of claim 1, wherein the water insoluble non-charged absorbent is comprised of a porous ceramic.
- 10. The process of claim 1, wherein the water insoluble non-charged absorbent is comprised of aluminum oxide or titanium oxide.
- 11. The process of claim 1, wherein the anseriform bird is a duck or a goose.
- 12. A method for isolating avian antibodies from egg yolk, the method comprising:
(a) contacting a preparation that comprises avian antibodies with a water insoluble absorbent that absorbs both avian antibodies and lipid; and (b) eluting the avian antibodies from the water insoluble absorbent with an aqueous buffer that releases the absorbed avian antibodies, but not the lipid, thereby providing an isolated preparation of avian antibodies.
- 13. The process of claim 1, wherein the buffer for flowing the water insoluble non-charged absorbent in step (b) comprises a chaotropic salt.
- 14. The process of claim 13, wherein the chaotropic salt comprises about 3M to about 6M guandine-HCl or about 1 M to about 3 M sodium thiocyanate.
- 15. The process of claim 1, further comprising a purification step by an immunoaffinity chromatography at a pH value ranging from about 4 to about 7 under an ionic strength of lower than about 50 mM.
- 16. The process of claim 15, wherein the pH value is ranging from about 5 to about 6.
- 17. The process of claim 16, wherein the pH value is ranging from about 5.6 to about 5.8.
- 18. The process of claim 1, wherein the aqueous fraction containing the yolk antibodies in step (b) is a flow-through solution which has flowed through the water insoluble non-charged absorbent.
- 19. The process of claim 1, wherein the aqueous fraction containing the yolk antibodies in step (b) is an eluate eluted from the water insoluble non-charged absorbent.
- 20. A process for selectively isolating IgY antibodies from egg yolk of an anseriform bird comprising performing first salting out by salting out an aqueous fraction containing yolk antibodies with (NH4)2SO4 of a first concentration ranging from about 15% (w/v) to about 24% (w/v) based on the volume of the aqueous fraction, and then performing second salting out by salting out the aqueous fraction containing yolk antibodies treated in the first salting out with (NH4)2SO4 of a second concentration ranging from about 25% (w/v) to about 40% (w/v) based on the volume of the aqueous fraction treated in the first salting out; wherein the IgY antibodies comprise antibodies with Fe regions and antibodies that lack Fc regions.
- 21. The process of claim 20, wherein the first concentration is not more than about 21% (w/v) based on the volume of the aqueous fraction.
- 22. The process of claim 20, wherein the second concentration is not more than about 31% (w/v) based on the volume of the aqueous fraction treated in the first salting out.
- 23. The process of claim 20, wherein the anseriform bird is a duck or a goose.
- 24. A method for enriching 5.7 S isoform avian antibodies relative to 7.8 S isoform avian antibodies, the method comprising:
(a) precipitating 7.8 S isoform antibodies using a precipitant salt at an ionic strength less than the ionic strength of 24% (NH4)2SO4 (w/v) to provide a supernatant that includes 5.7 S antibodies; and (b) precipitating the 5.7 S isoform antibodies from the supernatant using a precipitant salt, to thereby provide an isolated preparation enriched for 5.7 S isoform antibodies.
- 25. The process of claim 20, further comprising a purification step by an immunoaffinity chromatography at a pH value ranging from about 4 to about 7 under an ionic strength of lower than about 50 mM.
- 26. The process of claim 25, wherein the pH value is ranging from about 5 to about 6.
- 27. The process of claim 26, wherein the pH value is ranging from about 5.6 to about 5.8.
- 28. A process for selectively isolating IgY antibodies from egg yolk of an anseriform bird comprising:
(a) absorbing yolk antibodies in a water-miscible fraction obtained from the egg yolk of an anseriform bird with a water insoluble non-charged absorbent selected from the group consisting of silicate, silicon compounds, carbonate, sulfate, phosphate, carbon, cellulose and synthetic fiber, ceramics, and metal oxide, and wherein the water insoluble non-charged absorbent is at an amount effective for separating the yolk antibodies from the water-miscible fraction; (b) flowing the water insoluble non-charged absorbent with a buffer to obtain an aqueous fraction containing the yolk antibodies; (c) salting out the aqueous fraction containing yolk antibodies in step (b) with (NH4)2SO4 of a first concentration ranging from about 15% (w/v) to about 24% (w/v) based on the volume of the aqueous fraction; and (d) salting out the aqueous fraction containing yolk antibodies treated in step (c) with (NH4)2SO4 of a second concentration ranging from about 25% (w/v) to about 40% (w/v) based on the volume of the aqueous fraction treated in step (c); and wherein the IgY antibodies comprise antibodies with Fc regions and antibodies lack of Fe regions.
- 29. The process of claim 28, wherein the silicate comprises synthetic or natural clays, kaolin, talc, and calcium silicate.
- 30. The process of claim 28, wherein the silicon compound comprises fumed silica, amorphous silica, silica dioxide, silica gel, silicates, diatomaceous earth, and Fuller's earth.
- 31. The process of claim 28, wherein the carbonate comprises calcium carbonate and barium carbonate.
- 32. The process of claim 28, wherein the sulfate comprises calcium sulfate.
- 33. The process of claim 28, wherein the phosphate comprises calcium phosphate.
- 34. The process of claim 28, wherein the carbon comprises activated carbon and carbon fiber.
- 35. The process of claim 28, wherein the cellulose and synthetic fiber comprise cellulose powder.
- 36. The process of claim 28, wherein the ceramics comprises porosity ceramics.
- 37. The process of claim 28, wherein the metal oxide comprises aluminum oxide and titanium oxide.
- 38. The process of claim 28, wherein the anseriform bird is a duck or a goose.
- 39. The method of claim 12, further comprising separating 5.7 S isoform antibodies in the isolated preparation of avian antibodies from 7.8 S isoform antibodies.
- 40. The process of claim 28, wherein the buffer for flowing the water insoluble non-charged absorbent in step (b) comprises a chaotropic salt.
- 41. The process of claim 40, wherein the chaotropic salt comprises about 3M to about 6M guandine-HCl or about 1 M to about 3 M sodium thiocyanate.
- 42. The process of claim 28, wherein the first concentration is not more than about 21% (w/v) based on the volume of the aqueous fraction.
- 43. The process of claim 28, wherein the second concentration is not more than about 31% (w/v) based on the volume of the aqueous fraction treated in the first salting out.
- 44. The process of claim 28, further comprising a purification step by an immunoaffinity chromatography at a pH value ranging from about 4 to about 7 under an ionic strength of lower than about 50 mM.
- 45. The process of claim 44, wherein the pH value is ranging from about 5 to about 6.
- 46. The process of claim 45, wherein the pH value is ranging from about 5.6 to about 5.8.
- 47. The process of claim 28, wherein the aqueous fraction containing the yolk antibodies in step (b) is a flow-through solution which has flowed through the water insoluble non-charged absorbent.
- 48. The process of claim 28, wherein the aqueous fraction containing the yolk antibodies in step (b) is an eluate eluted from the water insoluble non-charged absorbent.
- 49. An IgY antibody selectively isolated from egg yolk of an anseriform bird, the IgY antibody being prepared according a process comprising:
(a) absorbing yolk antibodies in a water-miscible fraction obtained from the egg yolk of an anseriform bird with a water insoluble non-charged absorbent selected from the group consisting of silicate, silicon compounds, carbonate, sulfate, phosphate, carbon, cellulose and synthetic fiber, ceramics, and metal oxide, and wherein the water insoluble non-charged absorbent is at an amount effective for separating the yolk antibodies from the water-miscible fraction; (b) flowing the water insoluble non-charged absorbent with a buffer to obtain an aqueous fraction containing the yolk antibodies; (c) salting out the aqueous fraction containing yolk antibodies in step (b) with (NH4)2SO4 of a first concentration ranging from about 15% (w/v) to about 24% (w/v) based on the volume of the aqueous fraction; and (d) salting out the aqueous fraction containing yolk antibodies treated in step (c) with (NH4)2SO4 of a second concentration ranging from about 25% (w/v) to about 40% (w/v) based on the volume of the aqueous fraction treated in step (c); and wherein the IgY antibodies comprise antibodies with Fc regions and antibodies lack of Fc regions.
- 50. A preparation comprising 5.7 S isoform avian antibodies, wherein the 5.7 S are present at a concentration at least 10 fold greater than any 7.8 S isoform antibodies, if present, the preparation being prepared by a method comprising:
(a) contacting a preparation that comprises avian antibodies with a water insoluble absorbent that absorbs both avian antibodies and lipid; (b) eluting avian antibodies from the water insoluble absorbent with an aqueous buffer that releases the absorbed antibodies, but not the lipid, thereby providing an eluate; and (c) separating 5.7S isoform antibodies from the eluate, the separating being preferential for 5.7 S isoform antibodies relative to 7.8 S isoform antibodies.
- 51. The IgY antibody of claim 49, wherein the silicon compound comprises fumed silica, amorphous silica, silica dioxide, silica gel, silicates, diatomaceous earth, and Fuller's earth.
- 52. The IgY antibody of claim 49, wherein the carbonate comprises calcium carbonate and barium carbonate.
- 53. The IgY antibody of claim 49, wherein the sulfate comprises calcium sulfate.
- 54. The IgY antibody of claim 49, wherein the phosphate comprises calcium phosphate.
- 55. The IgY antibody of claim 49, wherein the carbon comprises activated carbon and carbon fiber.
- 56. The IgY antibody of claim 49, wherein the cellulose and synthetic fiber comprise cellulose powder.
- 57. The IgY antibody of claim 49, wherein the ceramics comprises porosity ceramics.
- 58. The IgY antibody of claim 49, wherein the metal oxide comprises aluminum oxide and titanium oxide.
- 59. The IgY antibody of claim 49, wherein the anseriform bird is a duck or a goose.
- 60. The IgY antibody of claim 49, wherein the buffer for flowing the water insoluble non-charged absorbent in step (b) comprises a chaotropic salt.
- 61. The IgY antibody of claim 60, wherein the chaotropic salt comprises about 3M to about 6M guandine-HCl or about 1 M to about 3 M sodium thiocyanate.
- 62. The IgY antibody of claim 49, wherein the first concentration is not more than about 21% (w/v) based on the volume of the aqueous fraction.
- 63. A preparation comprising 5.7 S isoform avian antibodies, wherein the 5.7 S isoform avian antibodies are at least 80% pure relative to other protein species and are present at a concentration at least 10 fold greater than any 7.8 S isoform antibodies, if present.
- 64. A pharmaceutical composition comprising an IgY antibody according to claim 49 together with a pharmaceutically acceptable carrier.
- 65. A kit for immunoassay, comprising an IgY antibody according to claim 49 and instructions for use.
RELATED APPLICATIONS
[0001] This application is a continuation-in-part and claims priority to U.S. application Ser. No. 09/733,210, filed Dec. 8, 2000, the content of which is incorporated herein by reference.
Continuation in Parts (1)
|
Number |
Date |
Country |
Parent |
09733210 |
Dec 2000 |
US |
Child |
10209006 |
Jul 2002 |
US |