Fluorescent microspheres and methods of using them
First Claim
1. A fluorescent marker system comprising a plurality of fluorescent microspheres, each fluorescent microsphere comprising an acrylic latex bead having a diameter of less than 500 angstroms, said bead having conjugated, by transacylation of ester terminations on the surface of said bead, (a) at least 5,000 fluorescent molecules, and (b) spacer arms and specific binding means attached to said spacer arms for attaching said microspheres to biological analytes.
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Abstract
Highly fluorescent latex microspheres have a diameter of less than five hundred angstroms and have more than five thousand fluorescent markers per sphere. The microspheres are prepared by reacting an acrylic latex bead with a diamine and a fluorescent amine at elevated pH to attach a spacer arm and a marker, respectively, to the bead by transacylation of ester terminations on the surface of the bead. A protein such as avidin or an immunoglobulin may then be conjugated to the spacer arm. A single fluorescent microsphere is visible using standard fluorescent microscopy. Therefore the microspheres may be utilitzed not only to visualize cell surface antigens but also DNA encoding for single genes, by means of a biotinylated DNA probe.
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Citations
15 Claims
- 1. A fluorescent marker system comprising a plurality of fluorescent microspheres, each fluorescent microsphere comprising an acrylic latex bead having a diameter of less than 500 angstroms, said bead having conjugated, by transacylation of ester terminations on the surface of said bead, (a) at least 5,000 fluorescent molecules, and (b) spacer arms and specific binding means attached to said spacer arms for attaching said microspheres to biological analytes.
- 7. A dyed microsphere comprising an acrylic latex bead having a diameter of less than 500 angstroms, at least 5,000 dye marker means conjugated to the surface of said bead by transacylation of ester terminations on the surface of said bead for rendering said bead individually visible by microscopy, a plurality of spacer arms attached to said bead by transacylation of ester terminations on the surface of said bead, and specific binding means on said spacer arms for binding specifically with a biological analyte to label said biological analyte.
- 10. A method of identifying a site on a biological analyte, said method comprising a step of reacting with said site a fluorescent acrylic latex microsphere having a diameter of less than 500 angstroms and at least 5,000 fluorescent markers conjugated to the microsphere by transcylation of ester terminations on the surface of said microsphere, and at least one species conjugated to the microsphere through a spacer arm by transacylation of ester terminations on the surface of said microsphere, said species having a specific affinity for said binding site, and a step of detecting said individual microsphere by fluorescence microscopy.
Specification