Search

We describe the synthesis and characterization of metal-chelating polymers with a degree of polymerization of 67 and 79, high diethylenetriaminepentaacetic acid (DTPA) functionality, [M.sub.w]/[M.sub.n], [less than or equal to] 1.17, and a maleimide as an orthogonal functional group for conjugation to antibodies. The polymeric disulfide form of the [DP.sub.n] = 79 DTPA polymer was analyzed by thermogravimetric analysis to determine moisture and sodium-ion content and by isothermal titration calorimetry (ITC) to determine the [Gd.sup.3+] binding capacity. These results showed each chain binds 68 [+ or -] 7 [Gd.sup.3-] per chain. Secondary goat antimouse IgG was covalently labeled with the maleimide form of the DTPA polymer ([DP.sub.n] = 79) carrying [sup.159]Tb. Conventional ICPMS analysis of this conjugate showed each antibody carded an average of 161 [+ or -] 4 [sup.159]Tb atoms. This result was combined with the ITC result to show there are an average of 2.4 [+ or -] 0.3 polymer chains attached to each antibody. Eleven monoclonal primary antibodies were labeled with different lanthanide isotopes using the same labeling methodology. Single cell analysis of whole umbilical cord blood stained with a mixture of 11 metal-tagged antibodies was performed by mass cytometry. 10.1021/ac101901x

A novel instrument for real time analysis of individual biological cells or other microparticles is described. The instrument is based on inductively coupled plasma timeof-flight mass spectrometry and comprises a threeaperture plasma--vacuum interface, a dc quadrupole turning optics for decoupling ions from neutral components, an rf quadrupole ion guide discriminating against low-mass dominant plasma ions, a point-to-parallel focusing dc quadrupole doublet, an orthogonal acceleration rellectron analyzer, a discrete dynode fast ion detector, and an 8-bit 1 GHz digitizer. A high spectrum generation frequency of 76.8 kHz provides capability for collecting multiple spectra from each particle-induced transient ion cloud, typically of 200-300 [mu]s duration. It is shown that the transients can be resolved and characterized individually at a peak frequency of 1100 particles per second. Design considerations and optimization data are presented. The figures of merit of the instrument are measured under standard inductively coupled plasma (ICP) operating conditions (< 3% cerium oxide ratio). At mass resolution (full width at half-maximum) M/[DELTA]M > 900 for m/z = 159, the sensitivity with a standard sample introduction system of > 1.4 x [10.sup.8] ion counts per second per mg [L.sup.-1] of Tb and an abundance sensitivity of (6 x [10.sup.-4])-(1.4 x [10.sup.-3]) (trailing and leading masses, respectively) are shown. The mass range (m/z = 125-215) and abundance sensitivity are sufficient for elemental immunoassay with up to 60 distinct available elemental tags. When 500) can be used, which provides >2.4 x [10.sup.8] cps per nag [L.sup.-1] of Tb, at (1.5 x [10.sup.-3]) - (5.0 x [10.sup.-3]) abundance sensitivity. The real-time simultaneous detection of multiple isotopes from individual 1.8 [micro]m polystyrene beads labeled with lanthanides is shown. A real time single cell 20 antigen expression assay of model cell fines and leukemia patient samples immuno-labeled with lanthanide-tagged antibodies is presented.