Radiant energy – Ionic separation or analysis – With sample supply means
Reexamination Certificate
2002-02-25
2004-03-09
Lee, John R. (Department: 2881)
Radiant energy
Ionic separation or analysis
With sample supply means
C250S283000, C250S281000
Reexamination Certificate
active
06703611
ABSTRACT:
FIELD OF THE INVENTION
The present invention generally relates to devices for performing standard electrospray ionization and nanoelectrospray ionization.
BACKGROUND OF THE INVENTION
Electrospray ionization is used to transform a liquid sample into gaseous ions. A sample solution is forced or pulled through a small sprayer needle so that a fine mist of nebulized sample droplets is created. The sample is sprayed toward a counter-electrode with a high voltage applied between the solution and the counter-electrode. The high voltage causes charged molecules to be formed from the solution.
One application of electrospray ionization has been the formation of ions from an analyte sample for analysis by mass spectrometry, which can produce an analysis based on very few molecules. A sample is typically sprayed at a source orifice of a mass spectrometer with high voltage applied between the solution and the orifice to generate the ions for analysis. Because of the importance of analyzing small amounts of biological samples (particularly complex biological samples), a great deal of interest has arisen in the use of low flow rate electrospray ionization devices.
Nanoelectrospray ionization is a subset of the electrospray ionization technique that uses very low flow rates to allow the analysis of very small amounts of sample by mass spectrometry. Common volumetric flow rates for nanoelectrospray ionization are in the nL/min range. In order to achieve a stable electrospray at such low flow rates, very small sprayers must be used. Typical sprayer needles used with nanoelectrospray ionization have openings with diameters in the 1-75 &mgr;m range, whereas standard electrospray ionization sprayer needles usually have openings of 75-300 &mgr;m in diameter. Such nanoelectrospray needles are fabricated using special techniques, usually by melting and pulling a larger capillary down to a smaller opening. In order to prevent sample carryover between experiments and because the tips of the nanoelectrospray needles are very fragile, the needles are usually only used for a single sample and are then discarded.
Capturing the entire plume of ions created with standard electrospray ionization is difficult because the plume can be several centimeters in diameter and the inlet orifice (e.g., a transfer capillary) on the vacuum system of the mass spectrometer is typically less than a millimeter in diameter. Any portion of the electrospray ionization plume not captured by the transfer capillary is wasted sample. One solution to this problem is U.S. Pat. No. 6,107,628 to Smith et al., which describes an apparatus for directing ions generated at atmospheric pressures into a region under vacuum. The apparatus of the '628 patent comprises a plurality of elements contained within a region maintained at a pressure between 10
−1
Millbrae and 1 bar, each of the elements having progressively larger apertures to form an “ion funnel” having an entry at the largest aperture and an exit at the smallest aperture. An RF voltage is applied to each of the elements so that the RF voltage applied to each of the elements is out of phase with the RF voltage applied to the adjacent element or elements. Although the apparatus of the '628 may achieve the goal of focusing a dispersion of charged particles, it does so by complicating the design of the electrospray ionization source.
In nanoelectrospray ionization, the small aperture size of the nanoelectrospray needles reduces the applied voltage necessary to sustain a spray, and the sprayer needle is thus positioned much closer to the sampling orifice than in electrospray. As a result of the shorter distance between the sprayer needle and orifice, and because of the smaller diameter sprayer needle, the plume from nanoelectrospray ionization is much smaller in size than the plume from standard electrospray ionization and most, if not all, of the ions created by nanoelectrospray ionization may be captured by the transfer capillary and sent to the mass spectrometer for analysis. This increase in efficiency is one of the main reasons nanoelectrospray ionization produces higher sensitivity than standard electrospray ionization. However, in order to have most or all of the ions that are created transferred into the mass spectrometer, the nanoelectrospray ionization needle must be precisely aligned with the small orifice into the mass spectrometer vacuum system. This alignment is difficult and is often only achieved using complicated and expensive cameras and microscope lenses. Additionally, because the nanoelectrospray needles are not commonly re-used (as is the case with standard electrospray ionization needles), the alignment has to be performed for every sample to be analyzed.
Researchers at Bruker Daltonics Inc. recently proposed a zero adjustment device for nanospray mass spectrometry as a solution to this problem. (See Wang et al., “Zero Adjustment Device for Nanospray Mass Spectrometry”,
Proceedings of the
48
th
ASMS Conference on Mass Spectrometry and Allied Topics,
Long Beach, Calif., 2000; pp. 379-380.) The zero adjustment device is a sub-unit that can be detached from an electrospray ionization source for the sample loading, nanospray needle exchanging, and source cleaning. A pre-opened nanospray needle is self-aligned by a needle mounting union and is inserted into an ionization channel when mounted. The needle position is fixed and no fine adjustment is needed. The ionization channel is attached to a pre-capillary used as an interface between the ionization channel and the main electrospray ionization capillary. The zero adjustment nanospray device can be operated with the needle tip in a wide range of positions, which allows more tolerances on spray needle mounting. However, neither the construction of the metal ionization channel used in the zero adjustment nanospray device nor the task of interfacing the zero adjustment nanospray device with different source designs are simple tasks.
Internal calibration of a mass spectrum produces the most accurate peak assignments of an analyte solution because the calibration ions experience essentially the same conditions as the analyte ions. Typically, a calibration solution is added to the analyte solution before it is electrosprayed. However, when electrospraying two solutions containing ions of interest, ionization suppression can occur. Ionization suppression occurs when one of the species present (i.e., either the analyte or the calibrant) is more easily ionized, thereby effectively suppressing the signal of the other species contained in the sample. In addition, mixing two solutions with different solvent systems can cause problems with adduct formation, solubility, and/or reactivity.
In order to try to avoid ion suppression and other problems occurring when electrospraying mixed solutions for internal calibration of a mass spectrum, multiple sprayer standard electrospray ionization has been proposed. The analyte solution is loaded into one of the electrospray ionization needles while a calibration solution is loaded in another. The needles are either aimed at a single sampling orifice or separate orifices are used and the streams are mixed once inside the vacuum system of the mass spectrometer. The use of two or more spray needles with standard electrospray ionization sources has been demonstrated by several research groups. (See, e.g., Andrien et al., “Multiple Inlet Probes for Electrospray and APCI Sources”,
Proceedings of the
46
th ASMS Conference on Mass Spectrometry and Allied Topics,
Orlando, Fla., 1998; p. 889.; Dresch et al., “Accurate Mass Measurements with a High Resolution Dual-Electrospray Time-of-Flight Mass Spectrometer”,
Proceedings of the
47
th ASMS Conference on Mass Spectrometry and Allied Topics,
Dallas, Tex., 1999; p. 1865-1866.; Jiang et al., “Development of Multi-ESI-Sprayer, Multi-Atmospheric-Pressure-Inlet Mass Spectrometry and Its Application to Accurate Mass Measurement Using Time-of-Flight Mass Spectrometry”,
Anal. Chem.
2000, 72, 20-24; Hannis et al., “A Dual Electrospray
Danell Ryan M.
Glish Gary L.
Burns Doane Swecker & Mathis L.L.P.
Hughes James P.
Lee John R.
The University of North Carolina at Chapel Hill
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