Claims
- 1. A method for identifying whether a target agent is present in a biological sample, the method comprising the steps of:
preparing a plurality of capture beads each having at least one transport probe affixed thereto; preparing a plurality of reporter beads each having at least one signal probe and one anchor agent affixed thereto; mixing said capture beads, said reporter beads, and said sample under binding conditions to permit formation of a dual bead complex, so that when said target agent is present in the sample, the reporter bead and capture bead each are bound to the target agent; isolating the dual bead complex from the mixture; selectively breaking up dual bead complexes bound by the target agent employing a digestion agent thereby dissociating the reporter beads and capture beads; isolating the dissociated reporter beads from the mixture to obtain an isolate; exposing the isolate to a target zone on an optical bio-disc, the target zone having a capture agent that binds to the anchor agent on the reporter beads thereby maintaining the reporter beads within the target zone; and detecting the presence of the reporter beads in the disc to indicate that the target agent is present in the sample.
- 2. The method according to claim 1 wherein said selective breaking up of non-specific binding between capture beads and reporter beads is performed prior to target quantification.
- 3. A method for identifying whether a target agent is present in a biological sample, the method comprising the steps of:
preparing a plurality of capture beads, each of said capture bead having at least one transport probe affixed thereto; preparing a plurality of reporter beads, each of said reporter beads having at least one signal probe affixed thereto; mixing said capture beads, said reporter beads, and said sample under binding conditions to permit formation of a dual bead complex, so that when said target agent is present in the sample, the reporter bead and capture bead each are bound to the target agent; selectively breaking up non-specific binding between capture beads and reporter beads by employing a wash buffer containing a dissociation agent during formation of said dual bead complex; isolating the dual bead complex from the mixture to obtain an isolate; exposing the isolate to a target zone on an optical bio-disc, the target zone having a capture agent that binds to the dual bead complex; and detecting the presence of the dual bead complex in the disc to indicate that the target agent is present in the sample.
- 4. The method according to claim 3 wherein said selective breaking up of non-specific binding between capture beads and reporter beads is performed prior to target quantification.
- 5. A method for identifying whether a target agent is present in a biological sample, the method comprising the steps of:
preparing a plurality of capture beads, each of said capture beads having at least one transport probe affixed thereto; preparing a plurality of reporter beads, each of said reporter beads having at least one signal probe affixed thereto; denaturing said target agent and maintaining the target agent in the denatured form by employing a hybridization buffer including a denaturing agent; mixing said capture beads, said reporter beads, and said sample under binding conditions to permit formation of a dual bead complex, so that when said target agent is present in the sample, the reporter bead and capture bead each are bound to the target agent; isolating the dual bead complex from the mixture to obtain an isolate; exposing the isolate to a target zone on an optical bio-disc, the target zone having a capture agent that binds to the dual bead complex; and detecting the presence of the dual bead complex in the disc to indicate that the target agent is present in the sample.
- 6. The method according to claim 5 wherein said target agent is a medical target agent.
- 7. A method of preparing a dual bead assay for use in an optical bio-disc, said method comprising the steps of:
providing a mixture of capture beads that have transport probes bound thereto; providing a mixture of reporter beads that have signal probes bound thereto; suspending said mixture of capture beads in a hybridization solution; adding to said mixture a target agent that hybridizes with said transport probes; adding to said mixture said reporter beads; allowing said signal probes to hybridize with said target agent to thereby form a dual bead complex including at least one capture bead and one reporter bead; separating non-specifically bound capture beads and reporter beads employing a wash buffer containing a dissociation agent during formation of said dual bead complex; separating said dual bead complex from unbound reporter beads; removing from said mixture said unbound reporter beads; and loading said mixture including said dual bead complex into an optical bio-disc for analysis.
- 8. The method according to claim 7 wherein said step of adding said target agent is performed after said step of adding said reporter beads.
- 9. A method of testing for the presence of a target-DNA in a DNA sample by use of an optical bio-disc, said method comprising the steps of:
preparing a DNA sample to be tested for the presence of a target-DNA; preparing a plurality of reporter beads each having attached thereto a plurality of strands of signal-DNA and an anchor agent, the target-DNA and the signal-DNA being complementary; preparing a plurality of capture beads each having attached thereto a plurality of transport-DNA, the target-DNA and transport-DNA being complimentary; mixing said DNA sample, said plurality of reporter beads, and said plurality of capture beads to thereby form a test sample, the transport-DNA and the signal-DNA being non-complimentary; allowing hybridization between said signal-DNA, any target-DNA, and transport-DNA existing in the DNA sample to thereby form a dual bead complex including at least one capture bead and one reporter bead; separating non-specifically bound capture beads and reporter beads employing a wash buffer containing a dissociation agent during formation of said dual bead complex; removing from the test sample reporter beads that are not associated with the dual bead complex; depositing said test sample in a flow channel of an optical bio-disc which is in fluid communication with a target zone, the target zone including a plurality of capture agents each including an amino group that attaches to an active layer to immobilize the capture agents within the target zone; allowing any anchor agent to bind with the capture agents so that reporter beads associated with the dual bead complex are maintained within the target zone; and detecting any dual bead complexes in the target zone to thereby determine whether target-DNA is present in the DNA sample.
- 10. A method of testing for the presence of a target-DNA in a test sample by use of an optical bio-disc, said method comprising the steps of:
preparing a test sample to be tested for the presence of a target-DNA; preparing a plurality of reporter beads each having attached thereto a plurality of strands of signal-DNA, the target-DNA and the signal-DNA being complementary; preparing a plurality of capture beads each having attached thereto a plurality of transport-DNA and an anchor agent, the target-DNA and transport-DNA being complimentary; depositing a plurality of capture beads and reporter beads in a mixing chamber, each of said reporter beads and said capture beads including said signal-DNA and said transport-DNA, respectively, being non-complimentary to each other; depositing said test sample in the mixing chamber of an optical bio-disc which is linked to a target zone by a connecting flow channel allowing any target-DNA existing in the test sample to bind to the signal-DNA and the transport-DNA on the reporter and the capture bead, respectively, to thereby form a dual bead complex; separating non-specifically bound capture beads and reporter beads employing a buffer containing a dissociation agent during formation of said dual bead complex; rotating the optical bio-disc to cause the dual bead complex to move from the mixing chamber through the flow channel and into the target zone, the target zone including a plurality of capture agents each including an amino group that attaches to an active layer to immobilize the capture agents within the target zone, said capture agent having affinity for the anchor agent; allowing any anchor agent to bind with the capture agent so that capture beads associated with dual bead complex are maintained within the capture zone; removing from the target zone reporter beads that are free of any dual bead complex; and detecting any dual bead complex in the target zone to thereby determine whether target-DNA is present in the test sample.
- 11. A method of testing for the presence of a target-RNA in a test sample by use of an optical bio-disc, said method comprising the steps of:
preparing a test sample to be tested for the presence of a target-RNA; preparing a plurality of reporter beads each having attached thereto a plurality of strands of signal-DNA, the target-RNA and the signal-DNA being complementary; preparing a plurality of capture beads each having attached thereto a plurality of transport-DNA and an anchor agent, the target-RNA and transport-DNA being complimentary; depositing a plurality of capture beads and reporter beads in a mixing chamber, each of said reporter beads and capture beads including the signal-DNA and the transport-DNA, respectively, being non-complimentary to each other; depositing said test sample in the mixing chamber of an optical bio-disc which is linked to a target zone by a connecting flow channel allowing any target-RNA existing in the test sample to hybridize with the signal-DNA and the transport-DNA on the reporter and the capture bead, respectively, to thereby form a dual bead complex; separating non-specifically bound capture beads and reporter beads employing a buffer containing a dissociation agent during formation of said dual bead complex; rotating the optical bio-disc to cause the dual bead complex to move from the mixing chamber through the flow channel and into the target zone, the target zone including a plurality of capture agents each including an amino group that attaches to an active layer to immobilize the capture agents within the target zone, said capture agent and said anchor agent having affinity to each other; allowing any anchor agent to bind with the capture agent so that capture beads associated with dual bead complex are maintained within the capture zone; removing from the target zone reporter beads that are free of any dual bead complex; and detecting any dual bead complex in the target zone to thereby determine whether target-RNA is present in the test sample.
- 12. A method of testing for the presence of a target-antigen in a test sample by use of an optical bio-disc, said method comprising the steps of:
preparing a test sample to be tested for the presence of a target-antigen; preparing a plurality of reporter beads each having attached thereto a plurality of signal-antibody, the signal-antibody having an affinity to epitopes on the target-antigen; preparing a plurality of capture beads each having attached thereto a plurality of transport-antibody and an anchor agent, the transport-antibody having affinity to epitopes on the target-antigen; depositing the capture beads and the reporter beads in a mixing chamber of an optical bio-disc, each of said reporter beads and capture beads including the signal-antibody and the transport-antibody, respectively, having no affinity to each other; depositing said test sample in the mixing chamber of said optical bio-disc which is linked to a target zone by a connecting flow channel allowing any target-antigen existing in the test sample to bind to the signal-antibody and the transport-antibody on the reporter and the capture bead, respectively, to thereby form a dual bead complex; separating non-specifically bound capture beads and reporter beads employing a buffer containing a dissociation agent during formation of said dual bead complex; rotating the optical bio-disc to cause the dual bead complex to move from the mixing chamber through the flow channel and into the target zone, the target zone including a plurality of capture agents each including an amino group that attaches to an active layer to immobilize the capture agents within the target zone; allowing any anchor agent to bind with the capture agent so that capture beads associated with dual bead complex are maintained within the capture zone; removing from the target zone reporter beads that are free of any dual bead complex; and detecting any dual bead complex in the target zone to thereby determine whether target-antigen is present in the test sample.
- 13. The method according to claim 12 wherein said dual bead complex is detected by directing a beam of electromagnetic energy from a disc drive assembly toward said target zone and analyzing electromagnetic energy returned from said target zones.
- 14. A method of making an optical bio-disc to test for the presence of a target agent in a test sample, the method comprising the steps of:
providing a substrate having a center and an outer edge; encoding information on an information layer associated with the substrate, the encoded information being readable by a disc drive assembly to control rotation of the disc; forming a target zone in association with the substrate, the target zone disposed at a predetermined location relative to the center of the substrate; depositing an active layer in the target zone; depositing a plurality of capture agents in the target zone, each capture agent including an amino group that covalently attaches to the active layer to immobilize the capture agent within the target zone; forming a flow channel in fluid communication with the target zone; forming a mixing chamber in fluid communication with the flow channel; depositing a plurality of reporter beads in the mixing chamber, each of the reporter beads having attached thereto a plurality of signal probes, each of the signal probes having affinity to the target agent; depositing a plurality of capture beads in the mixing chamber, each of the capture beads having attached thereto a plurality of transport probes and an anchor agent, each of the transport probes having affinity to the target agent, the transport probes and signal probes having no affinity toward each other, and the capture agents and the anchor agents having specific affinity to each other; separating non-specifically bound capture beads and reporter beads employing a buffer containing a dissociation agent; and adding a pre-determined amount of blocking agent to the mixing chamber to prevent non-specific binding of the beads to each other and the walls of the mixing chamber.
- 15. An optical bio-disc, comprising:
a substrate having encoded information associated therewith, said encoded information being readable by a disc drive assembly to control rotation of the disc; a target zone associated with said substrate, said target zone disposed at a predetermined location relative to said substrate; an active layer associated with said target zone; and a plurality of capture agents attached to said active layer so that when said substrate is rotated, said capture agents remain attached to said active layer to thereby maintain a number of said capture agents within said target zone so that when a dual bead complex that has been pre-washed in a buffer containing a dissociation agent is introduced into said target zone, said capture agent sequesters said dual bead complex therein to thereby allow detection of captured dual bead complex.
- 16. The optical bio-disc according to claim 15 wherein said capture agent is a single stranded oligonucleotide sequence, a double stranded oligonucleotide sequence, an antibody, an antigen, biotin, or streptavidin.
- 17. A method of making an optical bio-disc for testing for the presence of a target-DNA in a DNA sample, said method comprising the steps of:
providing a substrate having a center and an outer edge; encoding information on an information layer associated with the substrate, said encoded information being readable by a disc drive assembly to control rotation of the disc; forming a target zone in association with said substrate, said target zone disposed at a predetermined location relative to said center of said substrate; applying an active layer in said target zone; depositing within said target zone, a plurality of strands of capture-DNA each including an amino group that covalently attaches to said active layer to immobilize said strands of capture-DNA within said target zone; forming a flow channel in fluid communication with said target zone; forming a mixing chamber in fluid communication with the flow channel; depositing a plurality of reporter beads in the mixing chamber, each of said reporters including a signal-DNA that has an affinity for the target-DNA; depositing a plurality of capture beads in the mixing chamber, each of said capture bead including a transport-DNA that hybridizes with a portion of the target-DNA and is complementary to said capture-DNA, the transport-DNA and signal-DNA being non-complimentary; depositing a pre-determined amount of dissociation agent to reduce non-specific binding between said capture beads and said reporter beads; and designating an input site associated with the mixing chamber, the input site implemented to receive a DNA sample to be tested for the presence of any target-DNA.
- 18. The method according to claim 17 wherein when the DNA sample is deposited in the mixing chamber, hybridization occurs between the signal-DNA, the target-DNA, and the transport-DNA to thereby form a dual bead complex including at least one reporter bead and one capture bead.
- 19. The method according to claim 18 wherein when the disc is rotated, the dual bead complex move into the target zone and hybridization occurs between the anchor-DNA and the capture-DNA to thereby place the dual bead complex in the target zone.
- 20. A method of performing a genetic dual bead assay in association with a magneto-optical bio-disc, said method comprising the steps of:
providing a plurality of magnetic capture beads having covalently attached transport probes; providing a plurality of reporter beads having covalently attached specific sequences of DNA; preparing a sample containing target DNA molecules to be tested for DNA sequences complementary to said specific DNA sequences; loading said capture beads into a magneto-optical bio-disc via an inlet port provided therein, said magneto-optical bio-disc having a magnetic capture layer; loading said sample and said plurality of reporter beads into the bio-disc; rotating the bio-disc to facilitate hybridization of any target DNA present in the sample to said specific sequences of DNA on said reporter beads and to said transport probes to form dual bead complexes; interrogating a number of said magnetic capture beads with an incident beam of radiant energy to determine whether each of said number of magnetic capture beads has formed a dual bead complex; magnetizing specific regions of said magnetic capture layer to bind thereto a plurality of said dual bead complexes; and quantitating said plurality of said dual bead complexes.
- 21. The method according to claim 20 including the further step of rotating the disc to direct any unbound beads into a waste chamber.
- 22. The method according to claim 21 including the further step of de-magnetizing said specific regions of said magnetic capture layer to thereby release a number of said plurality of said dual bead complexes.
- 23. The method according to claim 22 including the further step of rotating the disc to direct the released number of dual bead complexes to an analysis area for further processing so that said released number of dual bead complexes are sequestered in said analysis area.
- 24. A method of performing a dual bead assay in association with a magneto-optical bio-disc, said method comprising the steps of:
providing a plurality of magnetic capture beads having attached transport probes; providing a plurality of reporter beads having attached signal probes; loading said capture beads into a magneto-optical bio-disc via an inlet port provided therein, said magneto-optical bio-disc having a magnetic capture layer; loading a sample containing a target and said plurality of reporter beads into the bio-disc; rotating the bio-disc to facilitate binding of said target and said reporter beads to said magnetic capture beads to form dual bead complexes; interrogating a number of said magnetic capture beads with an incident beam of radiant energy to determine whether each of said number of magnetic capture beads has formed a dual bead complex; magnetizing specific regions of said magnetic capture layer to bind thereto a plurality of said dual bead complexes; and quantitating said plurality of said dual bead complexes.
- 25. The method according to claim 24 including the further step of rotating the disc to direct any unbound beads into a waste chamber.
- 26. The method according to claim 25 including the further step of de-magnetizing said specific regions of said magnetic capture layer to thereby release a number of said plurality of said dual bead complexes.
- 27. The method according to claim 26 including the further step of rotating the disc to direct the released number of dual bead complexes to an analysis area for further processing so that said released number of dual bead complexes are sequestered in said analysis area.
- 28. The method according to claim 27 wherein said analysis area includes a reaction chamber having agents that react with the sequestered dual bead complexes.
- 29. A method of performing a multiplexed dual bead assay in association with a magneto-optical bio-disc, said method comprising the steps of:
providing at least two groups of differently sized magnetic capture beads, each group having magnetic capture beads of the same size and having a different specific type of transport probe associated with each group; providing a plurality of reporter beads having attached at least two different types of signal probes; loading said capture beads into a magneto-optical bio-disc via an inlet port provided therein, said magneto-optical bio-disc having a magnetic capture layer; loading a sample containing at least one target and said plurality of reporter beads into the bio-disc; rotating the bio-disc to facilitate binding of said target and said reporter beads to said magnetic capture beads to form dual bead complexes; interrogating a number of said magnetic capture beads with an incident beam of radiant energy to determine whether each of said number of magnetic capture beads has formed a dual bead complex; determining the size of the magnetic bead in the dual bead complex; magnetizing specific regions of said magnetic capture layer to bind thereto a plurality of said dual bead complexes; and quantitating said plurality of said dual bead complexes.
- 30. The method according to claim 29 wherein said step of quantitating includes quantitating said plurality of said dual bead complexes according to the size of the magnetic capture bead.
- 31. The method according to claim 29 including the further step of rotating the disc to direct any unbound beads into a waste chamber.
- 32. The method according to claim 31 including the further step of de-magnetizing said specific regions of said magnetic capture layer to thereby release a number of said plurality of said dual bead complexes containing same-sized magnetic capture beads.
- 33. The method according to claim 32 including the further step of rotating the disc to direct the released number of same-sized dual bead complexes to an analysis area for further processing so that said released number of same-sized dual bead complexes are sequestered in said analysis area.
- 34. The method according to claim 33 wherein said analysis area includes a reaction chamber having agents that react with the sequestered same-sized dual bead complexes.
- 35. A method of performing a multiplexed dual bead assay in association with a magneto-optical bio-disc, said method comprising the steps of:
providing at least two groups of different types of reporter beads, each group having reporter beads of the same type and having a different specific type of signal probe associated with each group; providing a plurality of magnetic capture beads having different types of transport probes attached thereto; loading said capture beads into a magneto-optical bio-disc via an inlet port provided therein, said magneto-optical bio-disc having a magnetic capture layer; loading a sample to be tested for at least one target and said plurality of reporter beads into the bio-disc; rotating the bio-disc to facilitate binding of any target present in said sample to said reporter beads and to said magnetic capture beads to form dual bead complexes; interrogating a number of said reporter beads with an incident beam of radiant energy to determine whether each of said number of reporter beads has formed a dual bead complex; determining the type of the reporter bead in the dual bead complex; magnetizing specific regions of said magnetic capture layer to bind thereto a plurality of said dual bead complexes; and quantitating said plurality of said dual bead complexes.
- 36. The method according to claim 35 wherein said step of quantitating includes quantitating said plurality of said dual bead complexes according to the type of reporter bead.
- 37. The method according to claim 35 including the further step of rotating the disc to direct any unbound beads into a waste chamber.
- 38. The method according to claim 37 including the further step of de-magnetizing said specific regions of said magnetic capture layer to thereby release a number of said plurality of said dual bead complexes containing same-type reporter beads.
- 39. The method according to claim 38 including the further step of rotating the disc to direct the released number of same-type dual bead complexes to an analysis area for further processing so that said released number of same-type dual bead complexes are sequestered in said analysis area.
- 40. The method according to claim 39 wherein said analysis area includes a reaction chamber having agents that react with the sequestered same-type dual bead complexes.
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application is a continuation-in-part of U.S. application Ser. No. 09/997,741 filed Nov. 27, 2001 which claimed the benefit of priority from U.S. Provisional Application Serial No. 60/253,283 filed Nov. 27, 2000; U.S. Provisional Application Serial No. 60/253,958 filed Nov. 28, 2000; and U.S. Provisional Application Serial No. 60/272,525 filed Mar. 1, 2001.
[0002] This application also claims the benefit of priority from U.S. Provisional Application Serial No. 60/275,643 filed Mar. 14, 2001; U.S. Provisional Application Serial No. 60/278,106 filed Mar. 23, 2001; U.S. Provisional Application Serial No. 60/278,110 also filed Mar. 23, 2001; U.S. Provisional Application Serial No. 60/278,697 filed Mar. 26, 2001; U.S. Provisional Application Serial No. 60/314,906 filed Aug. 24, 2001; and U.S. Provisional Application Serial No. 60/352,270 filed Jan. 30, 2002. Each of the above utility and provisional applications is herein incorporated by reference in its entirety.
Provisional Applications (9)
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Number |
Date |
Country |
|
60275643 |
Mar 2001 |
US |
|
60278106 |
Mar 2001 |
US |
|
60278110 |
Mar 2001 |
US |
|
60278697 |
Mar 2001 |
US |
|
60314906 |
Aug 2001 |
US |
|
60352270 |
Jan 2002 |
US |
|
60253283 |
Nov 2000 |
US |
|
60253958 |
Nov 2000 |
US |
|
60272525 |
Mar 2001 |
US |
Continuation in Parts (1)
|
Number |
Date |
Country |
Parent |
09997741 |
Nov 2001 |
US |
Child |
10099266 |
Mar 2002 |
US |