US 4,868,103 AGranted

Analyte detection by means of energy transfer

AssigneeEnzo Biochem, Inc.

InventorsJannis Stavrianopoulos, Elazar Rabbani, Samuel B. Abrams, James G. Wetmur

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Abstract

A method is disclosed to detect the presence of an analyte. The method involves forming a complex comprising the analyte and a binding entity. The binding entity comprises a first partner of an energy transfer system. The complex is then contacted with a reporting entity to form a unit. The reporting entity comprises a second partner of the energy transfer system. The first partner and the second partner are within Furster's radius of each other in the formed unit. The unit is irradiated with energy which can only be absorbed by one of said partners, namely, the energy donor, which then emits fluorescent energy. Some of this energy is absorbed by the other of said partners, namely, the energy acceptor, which also emits fluorescent energy. However, the fluorescent energy of the energy acceptor is of longer wavelength and in addition may be of substantially greater duration than the fluorescent energy of the energy donor. The detection of fluorescence at the longer wavelength or after a given time interval verifies the presence of the analyte.

Claims34

Dependent claims

An assay for detecting the presence of an analyte comprising the steps of:

a. forming a complex comprising said analyte and binding entity comprising an analyte recognition segment and a first partner of a fluorescent energy transfer system, wherein said first partner is selected from the group consisting of an energy donor and an energy acceptor, wherein said energy donor is a fluorescent aromatic agent or a lanthanide metal and said energy acceptor is a fluorescent, aromatic agent or a lanthanide metal, with the proviso that when said energy donor is a fluorescent, aromatic agent, then said energy acceptor is a fluorescent, aromatic agent or a lanthanide metal and when said energy donor is a lanthanide metal, then said energy acceptor is a fluorescent, aromatic agent;
b. contacting said complex with a reporting entity composition comprising a second partner of said fluorescent energy transfer system and at least one component for rendering said analyte detectable which binds to said second partner to form a unit, wherein:
(i) said second partner is selected from the group consisting of an energy donor and an energy acceptor, with the proviso that when said first partner is an energy donor then said second partner is an energy acceptor and when said first partner is an energy acceptor then said second partner is an energy donor, and (ii) the distance between said first partner and said second partner is 30 Angstroms or less;
c. irradiating said unit with energy that can be absorbed by said energy donor and not by said energy acceptor with the proviso that said energy donor emits fluorescent energy which can excite said energy acceptor;
and
d. detecting the fluorescence emitted by said energy acceptor.

3 more elements

An assay for detecting the presence of an analyte comprising the steps of:

a. forming a complex comprising said analyte and a binding entity comprising an analyte recognition segment and a first partner of a fluorescent energy transfer system, wherein said first partner is selected from the group consisting of an energy donor and an energy acceptor, wherein said energy donor is a fluorescent, aromatic agent and said energy acceptor is selected from the group consisting of fluorescent, aromatic agents, europium and terbium;
b. contacting said complex with a reporting entity comprising a second partner of said fluorescent energy transfer system and a component which binds to said second partner to form a unit, wherein:
(i) said second partner is selected from the group consisting of an energy donor and an energy acceptor, with the proviso that when said first partner is an energy donor then said second partner is an energy acceptor and when said first partner is an energy acceptor then said second partner is an energy donor, and (ii) the distance between said first partner and said second partner is 30 Angstroms or less;
c. irradiating said unit with energy that can be absorbed by said energy donor and not by said energy acceptor with the proviso that said energy donor emits fluorescent energy which can excite said energy acceptor and wherein said irradiation is performed without prior separation of said binding entity which has not formed said complex with said analyte;
and
d. detecting the fluorescence emitted by said energy acceptor.

3 more elements

Dependencies are inferred from the claim text.

Facts

Application: 06/831,250Filed 19 Feb 1986
Granted: 19 Sep 1989
Term ends: Not estimatedThe source records don’t support an estimate.

Classification11 codes

G01N 33/542
With immune complex formed in liquid phase: with steric inhibition or signal modification, e.g. fluorescent quenching
C12Q 1/6818
Characterised by the detection means: involving interaction of two or more labels, e.g. resonant energy transfer
G01N 21/6428
Fluorescence; Phosphorescence: measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g.…
G01N 2021/6441
With indicators, stains, dyes, tags, labels, marks: with two or more labels