Method and apparatus for flow cytometry linked with elemental analysis

Abstract

An apparatus ( 100 ) for sequentially analyzing particles such as single cells or single beads, by spectrometry. The apparatus, an elemental flow cytometer, includes means ( 102 ) for sequential particle introduction, means ( 104 ) to vaporize, atomize and excite or ionize the particles, or an elemental tag associated with an analyte on the particles, and means ( 106 ) to analyze the elemental composition of the vaporized, atomized and excited or ionized particles, or an elemental tag associated with the particles. Methods for sequentially analyzing particles such as singe cells or single beads by spectrometry are also described.

1 . An elemental flow cytometer, comprising:
 means for introducing particles sequentially into a device to vaporize, atomize and excite or ionize the particles or an elemental tag associated with the particles;   a device to vaporize, atomize and excite or ionize the particles or an elemental tag associated with the particles, downstream of the means for introducing particles sequentially; and   a spectrometer to analyze the vaporized, atomized and excited or ionized particles, or the elemental tag associated with the particles.   
     
     
         2 . The elemental flow cytometer of  claim 1  wherein the means for introducing particles sequentially introduces particles one at a time. 
     
     
         3 . The elemental flow cytometer of  claim 1  wherein the means for introducing particles sequentially comprises a cell injector. 
     
     
         4 . A mass-spectrometer-based flow cytometer, comprising:
 means for introducing particles sequentially into a device to vaporize, atomize and ionize the particles or an elemental tag associated with the particles;   a device to vaporize, atomize and ionize the particles or an elemental tag associated with the particles downstream of the means for introducing particles sequentially; and   a mass spectrometer operatively connected and downstream of the ion pretreatment device.   
     
     
         5 . The mass spectrometer-based flow cytometer of  claim 4  wherein the device to vaporize, atomize and ionize comprises an inductively coupled plasma. 
     
     
         6 . The mass spectrometer-based flow cytometer of  claim 4  wherein the means for introducing particles sequentially comprises a cell injector. 
     
     
         7 . The mass spectrometer-based flow cytometer of  claim 4  further comprising an in-line lysis between the means for introducing particles sequentially and the device to vaporize, atomize and ionize. 
     
     
         8 . The mass spectrometer-based flow cytometer of  claim 4  further comprising an in-line desolvation between the means for introducing particles sequentially and the device to vaporize, atomize and ionize. 
     
     
         9 . The mass spectrometer-based flow cytometer of  claim 4  wherein the mass spectrometer comprises a time-of-flight mass spectrometer. 
     
     
         10 . The mass spectrometer-based flow cytometer of  claims 4  wherein the mass spectrometer comprises a magnetic sector array detector mass spectrometer. 
     
     
         11 . The mass spectrometer-based flow cytometer of  claim 4  wherein the mass spectrometer comprises a 3D ion trap mass spectrometer. 
     
     
         12 . The mass spectrometer-based flow cytometer of  claims 4  wherein the mass spectrometer comprises a linear ion trap mass spectrometer. 
     
     
         13 . A mass-spectrometer-based flow cytometer, comprising:
 means for introducing particles sequentially into a device to vaporize, atomize and ionize the particles or an elemental tag associated with the particles;   a device to vaporize, atomize and ionize the particles or an elemental tag associated with the particles downstream of the means for introducing particles sequentially;   an ion pretreatment device operatively connected and downstream of the device to vaporize, atomize and ionize; and   a mass analyzer operatively connected and downstream of the ion prtreatment device.   
     
     
         14 . The mass spectrometer-based flow cytometer of  claim 13  wherein the ion pretreatment device comprises at least one of:
 a vacuum interface;   a high-pass mass filter downstream of the vacuum interface; and   a gas filled ion cooler cell downstream of the vacuum interface.   
     
     
         15 . The mass spectrometer-based flow cytometer of  claim 14  wherein the high-pass mass filter is operated as a bandpass mass filter with both high and low mass cutoffs. 
     
     
         16 . The mass spectrometer-based flow cytometer of  claim 14  comprising a high-pass mass filter and a gas filled ion cooler cell in a shared housing. 
     
     
         17 . A mass-spectrometer-based flow cytometer, comprising:
 means for introducing particles sequentially into a device to vaporize, atomize and ionize the particles or an elemental tag associated with the particles;   a device to vaporize, atomize and ionize the particles or an elemental tag associated with the particles downstream of the means for introducing particles sequentially; and   a mass spectrometer operatively connected and downstream of the ion pretreatment device.   
     
     
         18 . The mass spectrometer-based flow cytometer of  claim 17  wherein the device to vaporize, atomize and ionize comprises an inductively coupled plasma. 
     
     
         19 . The mass spectrometer-based flow cytometer of  claim 17  wherein the means for introducing particles sequentially comprises a cell injector. 
     
     
         20 . The mass spectrometer-based flow cytometer of  claim 17  further comprising an in-line lysis between the means for introducing particles sequentially and the device to vaporize, atomize and ionize. 
     
     
         21 . The mass spectrometer-based flow cytometer of  claim 17  further comprising an in-line desolvation between the means for introducing particles sequentially and the device to vaporize, atomize and ionize. 
     
     
         22 . The mass spectrometer-based flow cytometer of  claim 17  wherein the mass spectrometer comprises a time-of-flight mass spectrometer. 
     
     
         23 . The mass spectrometer-based flow cytometer of  claims 17  wherein the mass spectrometer comprises a magnetic sector array detector mass spectrometer. 
     
     
         24 . The mass spectrometer-based flow cytometer of  claim 17  wherein the mass spectrometer comprises a 3D ion trap mass spectrometer. 
     
     
         25 . The mass spectrometer-based flow cytometer of  claims 17  wherein the mass spectrometer comprises a linear ion trap mass spectrometer. 
     
     
         26 . An optical emission spectrometer based flow cytometer, comprising:
 means for introducing particles sequentially into a device to vaporize, atomize and excite or ionize the particles or an elemental tag associated with the particles;   a device to vaporize, atomize and excite or ionize the particles or an elemental tag associated with the particles downstream of the means for introducing particles sequentially; and   an optical emission spectrometer to analyze the vaporized, atomized and excited or ionized particles, or the elemental tag associated with the particles.   
     
     
         27 . The optical emission flow cytometer of  claim 26  wherein the device to vaporize, atomize and ionize comprises an inductively coupled plasma. 
     
     
         28 . The optical emission flow cytometer of  claim 26  wherein the means for introducing particles sequentially comprises a cell injector. 
     
     
         29 . The optical emission flow cytometer of  claim 26  further comprising an in-line lysis between the means for introducing particles sequentially and the device to vaporize, atomize and excite or ionize. 
     
     
         30 . The optical emission flow cytometer of  claim 26  further comprising an in-line desolvation between the means for introducing particles sequentially and the device to vaporize, atomize and excite or ionize. 
     
     
         31 . A method of analyzing a particle by spectrometxy, comprising:
 sequentially introducing particles or particles associated with an elemental tag, into a device to vaporize, atomize and excite or ionize the particles or the elemental tag associated with the particles; and   introducing the vaporized, atomized and excited or ionized particles or, the elemental tag associated with the particles into a spectrometer.   
     
     
         32 . The method of  claim 31  further comprising a step of labeling the particle with an elemental tag. 
     
     
         33 . The method of  claim 32  where the particle is a bead and in which at least one element tag is attached to the surface of the bead. 
     
     
         34 . The method of  claim 32  where the particle is a bead and in which at least one element tag is incorporated within the bead. 
     
     
         35 . The method of  claim 31  wherein the spectrometer comprises a mass spectrometer. 
     
     
         36 . The method of  claim 35  wherein the mass spectrometer comprises a time of flight mass spectrometer. 
     
     
         37 . The method of  claim 31  wherein the spectrometer comprises an optical emission spectrometer. 
     
     
         38 . A kit for the sequential analysis of particles by spectrometry, comprising: at least one reagent for labeling a particle; and instructions for the sequential analysis of the particles by spectrometry. 
     
     
         39 . A particle for the analysis of an analyte in a biological sample containing at least one of an element or an isotope, wherein on vaporization, atomization, excitation or ionization of the particle, the elements or isotopes are capable of measurement by either an optical spectrometer or mass spectrometer. 
     
     
         40 . The use of the particle of  claim 39  in either an optical spectrometer or mass spectrometer to produce signals or ratios of the elements or isotopes to distinguish the particle from other particles. 
     
     
         41 . The particle of  claim 39 , wherein the particle is a bead 
     
     
         42 . The particle of  claim 41 , wherein the bead comprises of polystyrene, agarose or silica. 
     
     
         43 . The particle of  claim 39 , wherein particle is less than 10 micrometer in diameter. 
     
     
         44 . The particle of  claim 43 , wherein the particle is less than 1 micrometer in diameter. 
     
     
         45 . The particle of  claim 44 , wherein the particle is about 150 nanometer in diameter. 
     
     
         46 . The particle of  claim 39 , wherein the element is selected from the group consisting of Ru, Rh, Pd, Ag, In, La, Ce, Pr, Nd, Sm, Eu, Th, Dy, Ho, Er, Tm, Yb, Lu, Hf, Re, Ir, Pt, Au, Tb and isotopes of the foregoing 
     
     
         47 . The particle of  claim 39 , further comprising an affinity substance capable of binding an analyte. 
     
     
         48 . The particle of  claim 47 , wherein the element or isotope is bound to the affinity substance. 
     
     
         49 . The particle of  claim 47 , wherein the affinity substance is selected from the group consisting of an oligonucleotide, aptamer, antibody and antigen. 
     
     
         50 . The particle of  claim 46 , wherein the element is Au. 
     
     
         51 . The particle of  claim 46 , wherein the element is Eu. 
     
     
         52 . The particle of  claim 46 , wherein the element is Tb. 
     
     
         53 . A method of detecting the presence of a biological analyte in a sample comprising:
 providing particles for the analysis of an analyte in a biological sample containing at least one of an element or an isotope, wherein the particle comprises an affinity substance capable of binding the analyte;   exposing the particles to the sample;   introducing the particles sequentially into a device that vaporizes, atomizes, excites or ionizes the particles; and   individually analyzing elemental composition of one or more of the vaporized, atomized, excited or ionized particles;   wherein on vaporization, atomization, excitation or ionization of the particle, the elements or isotopes are capable of measurement by either an optical spectrometer or mass spectrometer.   
     
     
         54 . The method of  claim 53 , wherein following the step of exposing the particles to the sample, there is an additional step of exposing the particles to a second affinity substance capable of binding the analyte and associated with a different at least one of an element or an isotope; 
     
     
         55 . A set of particles, each particle being a particle of  claim 39 , wherein the at least one of an element or an isotope is identical for all particles in the set and is present in sufficiently identical concentration in each particle. 
     
     
         56 . A group of sets of particles, each set according to  claim 55  and wherein the at least one of an element or an isotope is different for each set. 
     
     
         57 . A kit comprising the set of particles of  claim 55  and instructions for use. 
     
     
         58 . The kit of  claim 57 , wherein the instructions correspond to the steps of the method of  claim 53 . 
     
     
         59 . A kit comprising the group of  claim 56  and instructions for use. 
     
     
         60 . The kit of  claim 59 , wherein the instructions correspond to the steps of the method of  claim 53 .