Method for Selecting Desired level of Dye Loading and Controlling Loading of Polymer Microparticles

Abstract

Solute-loaded polymer microparticles are obtained by immersing microparticles in a bath comprising a selected solute dissolved in a ternary solvent system. A first solvent of the ternary system is a strong solvent for both the solute and the polymer from which the microparticle was formed. A second solvent is a weak solvent or non-solvent for the solute and the polymer (tuning solvent). A third solvent is a weak solvent or non-solvent for the solute and polymer, but serves as a co-solvent with respect to the first and second solvents in that it is miscible with both the first and second solvents. The amount of solute incorporated into the microparticles is controlled by adjusting the ratio of solute with respect to the microparticle polymer, and by adjusting the composition of the ternary solvent system, principally the amount of tuning solvent. The method is particularly useful for providing libraries of combinatorially encoded microparticles containing distinguishable dye loadings, particularly distinguishable fluorescent dye loadings.

1 - 39 . (canceled) 
     
     
         40 . A method of staining a population of preformed polymeric microparticles using a dye bath solution, wherein the dye bath solution contains known amounts of at least two fluorescent dyes solubilized in a homogeneous, ternary solvent mixture consisting of (i) at least one first solvent in which the dye and the microparticle polymer are soluble; (ii) at least one second solvent in which the dye and the polymer are not or only weakly soluble, said first and second solvents being immiscible or at most partially miscible; (iii) at least one third solvent in which the dye and the polymer are not or only weakly soluble, said third solvent being miscible with the first and second solvents and the concentration of the second solvent controlling the uptake of the dye into the microparticles by decreasing the solvency of the mixture of the first, second and third solvents for the dye;
 wherein the microparticles upon excitation emit at least two distinct fluorescent signals corresponding to the at least two fluorescent dyes, wherein an intensity of each of the at least two emitted signal is proportional to an amount of its corresponding dye in microparticle, and wherein the amount of second solvent in the dye bath solution has been determined by targeting the amount of dye needed to be loaded into the microparticles to achieve a target fluorescence intensity of said stained polymeric microparticles.   
     
     
         41 . The method according to  claim 40  wherein the dye is a fluorescent dye. 
     
     
         42 . The method according to  claim 40  wherein the dye is a hydrophobic dye. 
     
     
         43 . The method according to  claim 40  wherein the dye is selected from the group consisting of styryl dyes, pyrromethane dyes, coumarin dyes, and combinations thereof. 
     
     
         44 . The method according to  claim 40  wherein the microparticles comprise a hydrophobic polymer. 
     
     
         45 . The method according to  claim 44  wherein the hydrophobic polymer is a homopolymer or copolymer comprising a vinyl-containing monomer. 
     
     
         46 . The method according to  claim 45  selected from the group consisting of homopolymers or copolymers of styrene, methyl methacrylate, acrylamide, ethylene glycol, hydroxyethylmethacrylate, vinyltoluene, divinylbenzene, and combinations thereof. 
     
     
         47 . The method according to  claim 46  wherein the polymer is polystyrene or copolymer thereof containing at least 50% by weight styrene monomer units. 
     
     
         48 . The method according to  claim 47  wherein the polymer is a styrene/methacrylic acid copolymer. 
     
     
         49 . The method according to  claim 47  wherein the polymer is cross-linked. 
     
     
         50 . The method according to  claim 40  wherein the microparticles have a diameter of from about 0.1 to about 100 microns. 
     
     
         51 . The method according to  claim 50  wherein the particles are monodisperse. 
     
     
         52 . The method according to  claim 40  wherein the concentration of dye present in the microparticle suspension formed by contacting microparticles with said dye solution is from about 10 μg/g to about 100 μg/g, based upon the weight of the microparticle suspension. 
     
     
         53 . The method according to  claim 40  comprising a solvent wherein the first solvent is selected from the group consisting of methylene chloride, chloroform, tetrahydrofuran, dioxane, cyclohexane, benzene, toluene, butylacetate, lower chlorinated aliphatic hydrocarbons, and combinations thereof; the second solvent is water; and the third solvent is selected from the group consisting of acetone, lower alcohols, and combinations thereof. 
     
     
         54 . The method according to  claim 53  wherein the first solvent is methylene chloride or dichloromethane, and the second solvent is water. 
     
     
         55 . The method according to  claim 53  wherein the third solvent is alcohol. 
     
     
         56 . The method according to  claim 40  wherein the microparticle is a core-shell microparticle comprising a central core comprising one or more core polymers surrounded by a shell comprising one or more shell polymers. 
     
     
         57 . The method according to  claim 56  wherein the core polymer comprises a copolymer of styrene and a monomer more hydrophilic than styrene 
     
     
         58 . The method according to  claim 56  wherein the core polymer comprises methacrylic acid. 
     
     
         59 . The method according to  claim 40  wherein at least a portion of the microparticles are magnetically responsive. 
     
     
         60 . The method of  claim 40  wherein an emulsion is not formed in said polymer microparticle suspension.