Homogeneous enzyme immunoassay for simultaneous detection of multiple analytes
First Claim
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1. A method for detecting the presence of one or more non-serologically cross-reactive analyte types in a sample using a competitive homogeneous assay:
- where the assay detects a plurality of different analyte types that are non-serologically cross-reactive and, where the assay involves analyte and receptor binding pairs such that the presence of one or more different analyte types is determined by enzyme activity reflecting the concentration of analyte when present in excess of a predetermined concentration of the cutoff said method comprising the steps of;
(I) combining in an aqueous medium;
(a) glucose-6-phosphate dehydrogenase (G6PDH) analyte binding pair member conjugates, the conjugates comprised of G6PDH covalently linked to a plurality of known analyte binding pair members of which at least two are non-serologically cross-reactive;
(b) receptors able to bind to each analyte type to be detected and to the G6PDH-analyte binding pair member conjugates; and
,(c) a sample to be tested for the presence of any of the plurality of analyte types; and
,(II) detecting increased G6PDH activity in the aqueous medium due to competitive binding of the receptors with the analyte types in the sample where analyte types bind receptors permitting the G6PDH-analyte binding pair member conjugates to exhibit maximal enzyme activity;
wherein;
(i) concentrations of G6PDH-analyte binding pair member conjugates and of the receptors are adjusted in the aqueous mixture so that the enzyme rate at the predetermined cutoff concentrations is approximately the same for the different analyte types whose presence is to be detected;
(ii) wherein the G6PDH is deactivated by from about 20% to about 85% resulting from the covalent linkage to the analyte binding pair member; and
(iii)wherein the deactivated G6PDH is inhibited by from about 20% to about 85% when bound to the receptors.
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Abstract
Methods and kits are provided for the simultaneous detection of multiple analytes, i.e. to the determination of whether one or more of a plurality of analytes is present in a sample, particularly the plurality of analytes are structurally unrelated or significantly different, particularly to whether one or more of such analytes is present in a sample in a concentration that is above a predetermined minimum or cutoff value, and particularly for such analytes with different cutoffs. The methods and kits are particularly useful for-screening for the presence of a plurality of drugs (licit and/or illicit), performance-enhancing substances, and other chemicals, and involve a competitive enzyme immunoassay employing one or more conjugates of G6PDH with a plurality of the analytes.
28 Citations
16 Claims
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1. A method for detecting the presence of one or more non-serologically cross-reactive analyte types in a sample using a competitive homogeneous assay:
- where the assay detects a plurality of different analyte types that are non-serologically cross-reactive and, where the assay involves analyte and receptor binding pairs such that the presence of one or more different analyte types is determined by enzyme activity reflecting the concentration of analyte when present in excess of a predetermined concentration of the cutoff said method comprising the steps of;
(I) combining in an aqueous medium; (a) glucose-6-phosphate dehydrogenase (G6PDH) analyte binding pair member conjugates, the conjugates comprised of G6PDH covalently linked to a plurality of known analyte binding pair members of which at least two are non-serologically cross-reactive; (b) receptors able to bind to each analyte type to be detected and to the G6PDH-analyte binding pair member conjugates; and
,(c) a sample to be tested for the presence of any of the plurality of analyte types; and
,(II) detecting increased G6PDH activity in the aqueous medium due to competitive binding of the receptors with the analyte types in the sample where analyte types bind receptors permitting the G6PDH-analyte binding pair member conjugates to exhibit maximal enzyme activity; wherein; (i) concentrations of G6PDH-analyte binding pair member conjugates and of the receptors are adjusted in the aqueous mixture so that the enzyme rate at the predetermined cutoff concentrations is approximately the same for the different analyte types whose presence is to be detected; (ii) wherein the G6PDH is deactivated by from about 20% to about 85% resulting from the covalent linkage to the analyte binding pair member; and (iii)wherein the deactivated G6PDH is inhibited by from about 20% to about 85% when bound to the receptors. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16)
- where the assay detects a plurality of different analyte types that are non-serologically cross-reactive and, where the assay involves analyte and receptor binding pairs such that the presence of one or more different analyte types is determined by enzyme activity reflecting the concentration of analyte when present in excess of a predetermined concentration of the cutoff said method comprising the steps of;
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Current AssigneeLin-Zhi International Incorporated
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Original AssigneeLin-Zhi International Incorporated
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InventorsChang, Chiu Chin, Chia, Tom, Lin, Marie
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Primary Examiner(s)Cook; Lisa V
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Application NumberUS10/163,018Publication NumberTime in Patent Office2,597 DaysField of Search435/7.1, 435/7.4, 435/7.9, 435/7.92, 435/7.93, 435/26, 436/501, 436/518, 436/536, 424/130.1US Class Current435/7.1CPC Class CodesG01N 33/581 with enzyme label (includin...G01N 33/946 CNS-stimulants, e.g. cocain...G01N 33/948 Sedatives, e.g. cannabinoid...G01N 33/9486 Analgesics, e.g. opiates, a...
