Chemistry: electrical and wave energy – Processes and products – Electrophoresis or electro-osmosis processes and electrolyte...
Reexamination Certificate
1999-08-23
2001-11-20
Warden, Jill (Department: 1743)
Chemistry: electrical and wave energy
Processes and products
Electrophoresis or electro-osmosis processes and electrolyte...
C204S451000
Reexamination Certificate
active
06319379
ABSTRACT:
BACKGROUND OF THE INVENTION
The present invention relates to chromatography and electrophoresis analysis, particularly to a method of sample injection into a number of horizontal separation/analysis channels, and more particularly to an improved method that enables the efficient introduction of analyte samples wherein the amount of sample is significantly reduced and a means of sample stacking is provided.
Chromatography and electrophoresis analysis systems use columns filled with various media to separate analytes based on differing analyte mobilities due to an applied electric field. In all cases the introduction of sample is paramount to achieving quality results. Specifically, samples containing multiple analytes must be introduced in a tight zone to achieve high separation resolution. Discrete capillary systems have addressed this issue by using a time gated electrokinetic injection process whereby the lead end of the capillary is suspended vertically into a sample vial that contains the sample resuspended in a loading buffer. An electrode is placed into the loading buffer and a potential is applied on the order of seconds to drive the sample into the capillary. Typically it takes on the order of 5 microliters of sample solution to permit the required electrical contact to drive the sample. Once the sample has been introduced, the capillary is removed from the sample loading buffer and placed in a running buffer for the extent of the analysis. Due to the high costs of sample preparation, particularly in massively parallel applications such as DNA sequencing, there has been a need to reduce the amount of sample solution required to inject a small amount of sample into the separation/analysis channels.
The present invention provides a method for reducing the typical use of 5 microliters of sample solution to an amount of under 0.2 microliters, thus providing a great cost savings in sample preparation. Basically, the method of the present invention comprises a modified electrokinetic sample injection procedure, which involves preparation of the input of the separation channel, the physical sample introduction, and subsequent removal of excess material. This method is very useful for high throughput analysis systems such as DNA sequencing, and it will enable uniquely controllable sample stacking for increased loading efficiency electrophoresis.
SUMMARY OF THE INVENTION
It is an object of the present invention to provide a modified electrokinetic sample injection method for chromatography and electrophoresis analysis systems.
A further object of the invention is to provide an improved method for loading horizontal separation/analysis channels.
A further object of the invention is to reduce the quantity of sample solution used in horizontal microchannel separation/analysis systems.
Another object of the invention is to provide a method for loading when taken in conjunction with horizontal microchannel which allows much reduced sample volumes.
Another object of the invention is to provide a method for loading samples which provides a means of sample stacking to greatly reduce the concentration of the sample.
Another object of the invention is to provide a modified electrokinetic sample injection method which greatly reduces the amount of sample required for separation/analysis and the associated cost savings in sample preparation.
Other objects and advantages of the present invention will become apparent from the following description and accompanying drawings. The essence of the present invention is in the preparation of the input of the separation channel, the physical sample introduction, and subsequent removal of excess material. The invention is a modified electrokinetic sample injection method for chromatography and electrophoresis analysis, wherein the amount of sample volume is significantly reduced (i.e. 5 &mgr;liters down to ~0.2 &mgr;liters). By the reduction of the amount of sample used for analysis systems incorporating multiple horizontal microchannels, the cost of sample preparation is greatly reduced, which is significant in massively applications such as DNA sequencing. Basically the method of this invention involves: (1) filling the microchannels with separation media, (2) removal of the media from the input sample load well, (3) loading the sample resuspended in a high density buffer, (4) applying an electric field to load sample into the separation media, (5) removing the electric field and decanting excess sample, and (6) applying separation electric field for analysis.
Balch Joseph W.
Davidson J. Courtney
Carnahan L. E.
Noguerola Alex
The Regents of the University of California
Thompson Alan H.
Warden Jill
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