Method to use fluorescent magnetic polymer particles as markers in an immunoassay
First Claim
1. A process to determine the presence or concentration of an analyte in a fluid specimen comprising:
- (a) contacting a solution of monodispersed fluorescent magnetic particles having an average diameter from about 1 micron to about 100 microns, a uniform size distribution and magnetic content, comprising an inner fluorescent core polymer particle able to adsorb a monomer, and a coating on said inner fluorescent core polymer particle wherein said coating evenly covers said inner fluorescent core polymer particle, said coating comprising magnetic metal oxide particles of about 1 micron or less in diameter and selected from the group consisting of supermagnetic, paramagnetic and ferromagnetic metal oxide particles and a polymer, wherein said polymer is prepared from monomers which are adsorbable by the inner fluorescent core particle, said fluorescent magnetic particles having a ligand which specifically binds said analyte attached to said fluorescent magnetic particles, with said fluid specimen to form a suspension;
(b) incubating said suspension until sufficient analyte has reacted with said ligand which specifically binds said analyte;
(c) separating said magnetic particles from said suspension;
(d) adding a second labeled ligand which specifically binds said analyte to said separated magnetic particles;
(e) incubating said suspension until sufficient analyte has reacted with said second labeled ligand which specifically binds said analyte;
(f) separating said magnetic particles from said suspension;
(g) detecting or measuring duplex formation on said magnetic particles by means of said label; and
(h) relating the amount of labeled ligand measured with the amount of analyte measured for a control sample also, said control sample having been evaluated by the same process as the fluid specimen, wherein said fluorescent magnetic particles are used to monitor the number of particles present during said process, wherein the number of said particles is monitored by measuring the fluorescence intensity of fluorescent particles prior to contacting said particles with said fluid specimen and by measuring the fluorescent intensity of fluorescent particles after measuring the amount of labeled ligand associated with said fluorescent magnetic particles.
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Abstract
This invention provides a novel process of using magnetically responsive fluorescent polymer particles comprising polymeric core particles coated evenly with a layer of polymer containing magnetically responsive metal oxide as highly sensitive quantitative reagents for biochemical and immunological studies, wherein the fluorescent magnetic particles can serve as a marker for the number of particles. A wide variety of polymeric particles with sizes ranging from 1 to 100 microns can be used a core particles and transformed into magnetically responsive polymer particles. The surface of these magnetically responsive polymer particles can be coated further with another layer of functionalized polymer. These magnetically responsive fluorescent polymer particles can be used for passive or covalent coupling of biological material such as antigens, antibodies, enzymes or DNA/RNA hybridization and used as solid phase for various types of immunoassays, DNA/RNA hybridization probes assays, affinity purification, cell separation and other medical, diagnostic, and industrial applications.
217 Citations
4 Claims
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1. A process to determine the presence or concentration of an analyte in a fluid specimen comprising:
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(a) contacting a solution of monodispersed fluorescent magnetic particles having an average diameter from about 1 micron to about 100 microns, a uniform size distribution and magnetic content, comprising an inner fluorescent core polymer particle able to adsorb a monomer, and a coating on said inner fluorescent core polymer particle wherein said coating evenly covers said inner fluorescent core polymer particle, said coating comprising magnetic metal oxide particles of about 1 micron or less in diameter and selected from the group consisting of supermagnetic, paramagnetic and ferromagnetic metal oxide particles and a polymer, wherein said polymer is prepared from monomers which are adsorbable by the inner fluorescent core particle, said fluorescent magnetic particles having a ligand which specifically binds said analyte attached to said fluorescent magnetic particles, with said fluid specimen to form a suspension; (b) incubating said suspension until sufficient analyte has reacted with said ligand which specifically binds said analyte; (c) separating said magnetic particles from said suspension; (d) adding a second labeled ligand which specifically binds said analyte to said separated magnetic particles; (e) incubating said suspension until sufficient analyte has reacted with said second labeled ligand which specifically binds said analyte; (f) separating said magnetic particles from said suspension; (g) detecting or measuring duplex formation on said magnetic particles by means of said label; and (h) relating the amount of labeled ligand measured with the amount of analyte measured for a control sample also, said control sample having been evaluated by the same process as the fluid specimen, wherein said fluorescent magnetic particles are used to monitor the number of particles present during said process, wherein the number of said particles is monitored by measuring the fluorescence intensity of fluorescent particles prior to contacting said particles with said fluid specimen and by measuring the fluorescent intensity of fluorescent particles after measuring the amount of labeled ligand associated with said fluorescent magnetic particles. - View Dependent Claims (2)
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3. A process to determine the presence or concentration of an analyte in a fluid specimen comprising:
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(a) contacting a solution of monodispersed fluorescent magnetic particles having an average diameter from about 1 micron to about 100 microns, a uniform size distribution and magnetic content, comprising an inner core polymer particle able to adsorb a monomer, and a coating on the inner core polymer particle wherein the coating evenly covers the inner core polymer particles, said coating comprising a fluorescent dye or combination of fluorescent dyes, magnetic metal oxide particles of about 1 micron or less in diameter and selected from the group consisting of paramagnetic, supermagnetic and ferromagnetic metal oxides and a polymer, said polymer being prepared from monomers which are adsorbable by the inner core polymer particle said fluorescent magnetic particles having a ligand which specifically binds said analyte attached to said fluorescent magnetic particles, with a fluid specimen to form a suspension; (b) incubating said suspension until sufficient analyte has reacted with said ligand which specifically binds said analyte; (c) separating said magnetic particles from said suspension; (d) adding a second labeled ligand which specifically binds said analyte to said separated magnetic particles; (e) incubating said suspension until sufficient analyte has reacted with said second labeled ligand which specifically binds said analyte; (f) separating said magnetic particles from said suspension; (g) detecting or measuring duplex formation on said magnetic particles by means of said label; and (h) relating the amount of labeled ligand measured with the amount of analyte measured for a control sample, said control sample also having been evaluated by the same process as the fluid specimen, wherein said fluorescent magnetic particles are used to monitor the number of particles present during said process, wherein the number of said particles is monitored by measuring the fluorescence intensity of fluorescent particles prior to contacting said particles with said fluid specimen and by measuring the fluorescent intensity of fluorescent particles after measuring the amount of labeled ligand associated with said fluorescent magnetic particles. - View Dependent Claims (4)
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Specification