Method of attaching a ligand to a solid support
First Claim
1. A lateral-flow test method for detecting one or more analytes in a test sample, the method comprising:
- (a) adding the test sample to a lateral flow test strip, the test strip including a test zone binder with affinity for an unbound labeled analyte receptor, the test strip comprising a test zone and a control zone in a spaced apart configuration on the test strip;
(b) allowing the sample to contact the unbound labeled receptor, the unbound labeled receptor having affinity to an analyte to be detected so that the unbound labeled receptor reacts with the analyte to form analyte bound labeled receptor;
(c) subsequent to allowing the sample to contact the unbound labeled receptor, allowing the unbound labeled receptor to come in contact with the test zone binder, the test zone binder having affinity to unbound labeled receptor but not the analyte bound labeled receptor, wherein when the unbound labeled receptor reacts with the test zone binder a visible signal can be detected in the test zone; and
(d) detecting the visible signal in the test zone,wherein the test is negative for the detection of the one or more analytes when the signal from the test zone is more intense than the signal in the control zone and the test is positive for the one or more analytes when the signal from the control zone is more intense than the signal from the test zone,and characterized in that the test zone binder is attached to the test strip through a spacer linkage to a protein and wherein the protein is attached to both the test strip and the spacer linkage.
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Abstract
The invention features a method of attaching a ligand that has a free carboxyl group to a solid support by adding an amino group to the ligand to form a ligand-amino derivative, converting the ligand amino derivative to a ligand sulfhydryl derivative, attaching the ligand sulfhydryl derivative to a protein to form a ligand-linker-protein conjugate, and applying the ligand-linker-protein conjugate to the solid support. The method is particularly useful for immobilizing small molecule ligands having a free carboxyl group, such as cloxicillin, to a lateral-flow test strip, in order to make a detection zone on the test strip that exhibits a clear signal and enhanced sensitivity.
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Citations
39 Claims
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1. A lateral-flow test method for detecting one or more analytes in a test sample, the method comprising:
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(a) adding the test sample to a lateral flow test strip, the test strip including a test zone binder with affinity for an unbound labeled analyte receptor, the test strip comprising a test zone and a control zone in a spaced apart configuration on the test strip; (b) allowing the sample to contact the unbound labeled receptor, the unbound labeled receptor having affinity to an analyte to be detected so that the unbound labeled receptor reacts with the analyte to form analyte bound labeled receptor; (c) subsequent to allowing the sample to contact the unbound labeled receptor, allowing the unbound labeled receptor to come in contact with the test zone binder, the test zone binder having affinity to unbound labeled receptor but not the analyte bound labeled receptor, wherein when the unbound labeled receptor reacts with the test zone binder a visible signal can be detected in the test zone; and (d) detecting the visible signal in the test zone, wherein the test is negative for the detection of the one or more analytes when the signal from the test zone is more intense than the signal in the control zone and the test is positive for the one or more analytes when the signal from the control zone is more intense than the signal from the test zone, and characterized in that the test zone binder is attached to the test strip through a spacer linkage to a protein and wherein the protein is attached to both the test strip and the spacer linkage. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11)
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12. A lateral-flow test method for detecting one or more analytes in a test sample, the method comprising:
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(a) attaching to a lateral flow test strip, through a spacer linkage to a protein a test zone binder with affinity for unbound labeled analyte receptor, said attaching through a spacer linkage comprising;
(i) providing the test zone binder, said test zone binder having a free carboxyl group;
(ii) adding an amino group to said test zone binder to form a test zone binder-amino derivative;
(iii) converting said test zone binder-amino derivative to a test zone binder-sulfhydryl derivative;
(iv) joining said test zone binder-sulfhydryl derivative to the protein to form a test zone binder-spacer linkage-protein conjugate;(b) applying said test zone binder-spacer linkage-protein conjugate to said test strip; (c) contacting the sample with the unbound labeled analyte receptor, the unbound labeled analyte receptor having affinity to at least one analyte; (d) allowing the unbound labeled receptor to react with the analyte to form a bound labeled receptor; (e) subsequent to contacting the sample with the unbound labeled receptor, contacting the unbound labeled receptor with the test zone binder, the test zone binder having affinity to the unbound labeled receptor but not the bound labeled receptor, wherein at least a portion of the unbound labeled receptor will react with the test zone binder generating a visible signal in the test zone that can be detected; and (f) detecting the visible signal in the test zone, wherein the test is negative for the detection of the one or more analytes when the signal from test zone is more intense than the signal in a control zone, the control zone located on the test strip in a spaced apart relationship with the test zone, and the test is positive for one or more analytes when the signal from a control zone is more intense than the signal from the test zone. - View Dependent Claims (13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35)
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36. A lateral-flow test method for detecting one or more analytes in a test sample, the method comprising:
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(a) attaching a test zone binder with affinity for unbound labeled analyte receptor to a test zone on a lateral flow test strip, said attaching comprising;
(i) providing a test zone binder having a free carboxyl group;
(ii) adding an amino group to said test zone binder to form a binder-amino derivative;
(iii) joining said test zone binder amino derivative to a protein having a free amino group, through an amino group directed homobifunctional cross-linker, to form a test zone binder-spacer linkage-protein conjugate; and
(iv) applying said test zone binder-spacer linkage-protein conjugate to said test strip;(b) contacting the sample with the unbound labeled receptor, the unbound labeled receptor having affinity to at least one analyte to be detected; (c) allowing the unbound labeled receptor to react with the analyte; (d) contacting the unbound labeled receptor, previously allowed to react with the analyte, with the test zone binder, wherein a portion of the receptor, unbound to analyte from the sample, will react with the test zone binder in the test zone generating a visible result that can be detected; (e) detecting the visible signal in the test zone, wherein the test is negative for the detection of the one or more analytes when the signal from the test zone is more intense than the signal in a control zone, the control zone located on the test strip in a spaced apart relationship with the test zone, and the test is positive for one or more analytes when the signal from the control zone is more intense than the signal from the test zone. - View Dependent Claims (37, 38, 39)
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Specification