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Systems for microchip capillary electrophoresis absent electrokinetic injection

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Systems for microchip capillary electrophoresis absent electrokinetic injection


Microchip capillary electrophoresis (CE) utilizing a sample injector based on a mechanical valve rather than electrokinetic injection can provide improved sample injections, enhanced capabilities, and can eliminate the need for changing the electric field in the separation channel to induce sample injection. In one instance CE electrodes continuously apply an electric field for CE separation along a separation channel. A sample channel is connected to the separation channel at an intersection and has a sample pressure that is greater than that which is present in the separation channel near the intersection. The sample channel does not have electrodes that apply voltages for electrokinetic injection. A sample injector in the sample channel or at the intersection comprises a mechanical valve to control sample injection from the sample channel to the separation channel.

Browse recent Battelle Memorial Institute patents - Richland, WA, US
Inventors: Xuefei Sun, Ryan T. Kelly, Keqi Tang, Richard D. Smith
USPTO Applicaton #: #20120318672 - Class: 204604 (USPTO) - 12/20/12 - Class 204 
Chemistry: Electrical And Wave Energy > Apparatus >Electrophoretic Or Electro-osmotic Apparatus >Capillary Electrophoresis Type >With Injector

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The Patent Description & Claims data below is from USPTO Patent Application 20120318672, Systems for microchip capillary electrophoresis absent electrokinetic injection.

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CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims priority to, and is a division of U.S. patent application Ser. No. 12/889,108 filed on Sep. 23, 2010, which is incorporated herein by reference.

STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT

This invention was made with Government support under Grant Number RR018522 from the U.S. National Institute of Health and Contract Number DE-ACO576RL01830 awarded by the U.S. Department of Energy. The Government has certain rights in the invention.

BACKGROUND

When performing microchip capillary electrophoresis (CE), sample introduction and injection can significantly affect CE performance. Currently, electrokinetic injection is used almost exclusively for microchip CE sample injection. In a particular type of electrokinetic injection known as “pinched” injection, various electrodes in a first mode apply voltages in four intersecting channels to drive sample through the intersection to a waste reservoir. In a second mode, the voltages induce injection of only a plug of sample occupying the small volume in the intersection towards a separation channel. The voltage applied to the separation channel is different in the first mode, wherein sample is diverted to waste, compared to the second mode, wherein the plug of sample is injected into the separation channel. Except for the sample plug, the vast majority of sample is wasted.

While electrokinetic injection can yield small sample plugs for improved separation efficiency and can minimize electrophoretic injection bias under certain conditions, it also has several significant limitations. For example, a considerable amount of time is required to achieve steady state in the first mode. Steady state is a necessary condition to avoid sample bias and/or injection bias caused by high mobility species arriving more quickly than low mobility species. During prolonged operation, the high mobility species can be depleted preferentially and prematurely from the sample supply. Sample utilization is extremely inefficient because the total volume required is very large compared to the actual injected plug volume, which is very small. Furthermore, the injection volume is fixed because it is determined by the geometry of the intersection. In order to change the injection volume, the geometry of the intersection must typically be altered. Further still, the rate at which sequential injections can be analyzed, and the total number of sample plugs that can be injected into the separation channel, is limited by the steady-state flow requirements and by the changing voltages in the separation channel associated with electrokinetic injection.

In view of at least the limitations described above, a need for an improved sample injector and microchip CE system exists.

SUMMARY

The present invention is a microchip CE system that utilizes a sample injector based on a mechanical valve rather than electrokinetic injection. The mechanical valve can be operated to provide rapid sequential sample injections and to eliminate the need for changing the electric field in the separation channel to induce sample injection. Instead, sample injection is accomplished by pressure gradients and by opening the mechanical valve. A constant electric field can, therefore, be continuously applied along the separation channel. Varying the sample pressure and/or the duration of time that the valve remains in an open position can vary sample injection volume. Preferably mass spectrometry is performed to analyze the sample after CE separation.

In one embodiment, the microchip CE system comprises a separation channel and CE electrodes continuously applying an electric field for CE separation along the separation channel. As used herein, the continuous application of an electric field along the separation channel for CE separation is significant because any sample injection provided to the separation channel will be subject to a continuously applied CE separation field. There is no required change in voltage between an injection mode and a CE separation mode.

A sample channel is connected to the separation channel at an intersection and has a sample pressure that is greater than that which is present in the separation channel near the intersection. The sample channel does not have electrodes that apply voltages for electrokinetic injection. A sample injector in the sample channel, or at the intersection, comprises a mechanical valve to control sample injection from the sample channel to the separation channel. When the valve is opened for a short time, a small volume of sample solution is pushed into the separation channel under a low pressure. When the valve is closed, the sample solution is completely isolated from the CE run buffer in the separation channel such that there is no risk of sample leakage during the operation, and a discrete, well defined sample plug is injected with each valve opening event.

A significant characteristic of the system is that the sample injection is independent of the CE separation. During operation, a high voltage is applied only along the separation channel and no voltage switching is needed. The sample is directly provided into the separation channel for subsequent CE separation. There is no need to wait for production of a steady-state, stable sample plug as would be required in the traditional electrokinetic injection. Discrete sample plugs can be injected repeatedly over relatively long periods of time. The injection and separation frequency is only determined by the actuation of the mechanical valve. A valve having a high duty cycle makes it possible to perform continuous flow monitoring, high throughput analysis, and/or multiplexed separations.

Embodiments of the present invention can further comprise a plurality of discrete injections of a sample from the sample channel to the separation channel in a rapid sequence. The sequence can preferably be pseudo-random. A detector at the end of the separation channel detects the discrete injections after CE-induced overlap, which comprises mixing of at least one component from at least one of the discrete injections to another discrete injection. A processing device executes programming to deconvolute the CE-induced overlap in data collected by the detector so that a spectrum can be reconstructed.

Some embodiments of a microchip CE system can further comprise a plurality of CE channels within the separation channel as well as a manifold within the separation channel distributing one or more discrete injections among the plurality of CE channels. Preferably, an electrospray ionization (ESI) emitter is connected at the end of each CE channel.

In some embodiments, liquid chromatography (LC) separation is performed in conjunction with CE separation. Accordingly, the microchip CE system can further comprise a LC column connected to the sample channel and providing LC separation prior to injection into the separation channel.

Embodiments of the present invention also include methods for analyzing a sample having a plurality of components using microchip CE. In a particular embodiment, the method includes the steps of applying a sample pressure in the sample channel greater than the sample pressure in a separation channel. The sample channel is connected to the separation channel at an intersection and lacks electrodes associated with electrokinetic-based injectors. A continuous electric field for CE separation is applied along the separation channel. Injection of the sample occurs by mechanically opening for a duration a mechanical valve, not an electrokinetic-based injector. The mechanical valve is located in the sample channel or at the intersection. The electric field in the separation channel can then separate the components in the injection.

The method can further comprise repeating the mechanical opening in a rapid, pseudo-random sequence to provide a plurality of discrete injections of the sample from the sample channel to the separation channel. CE-induced overlap can be the result of mixing at least one component from at least one of the discrete injections with another discrete injection. The discrete injections are then detected at the end of the separation channel after CE-induced overlap. Finally, the CE-induced overlap in data collected by the detector is deconvoluted so that a spectrum can be reconstructed.

Alternatively, the method can further include distributing one or more injections among a plurality of CE channels within the separation channel. In preferred embodiments, an electrospray can be generated at the end of each CE channel.

In another embodiment, LC separations can be performed in conjunction with the CE separations. Accordingly, the method can further comprise separating the sample in a liquid chromatography column prior to providing an injection to the separation channel.

The purpose of the foregoing abstract is to enable the United States Patent and Trademark Office and the public generally, especially the scientists, engineers, and practitioners in the art who are not familiar with patent or legal terms or phraseology, to determine quickly from a cursory inspection the nature and essence of the technical disclosure of the application. The abstract is neither intended to define the invention of the application, which is measured by the claims, nor is it intended to be limiting as to the scope of the invention in any way.

Various advantages and novel features of the present invention are described herein and will become further readily apparent to those skilled in this art from the following detailed description. In the preceding and following descriptions, the various embodiments, including the preferred embodiments, have been shown and described. Included herein is a description of the best mode contemplated for carrying out the invention. As will be realized, the invention is capable of modification in various respects without departing from the invention. Accordingly, the drawings and description of the preferred embodiments set forth hereafter are to be regarded as illustrative in nature, and not as restrictive.



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stats Patent Info
Application #
US 20120318672 A1
Publish Date
12/20/2012
Document #
13596360
File Date
08/28/2012
USPTO Class
204604
Other USPTO Classes
International Class
01N27/447
Drawings
11



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