Chemistry: electrical and wave energy – Apparatus – Electrophoretic or electro-osmotic apparatus
Reexamination Certificate
1998-11-12
2001-01-23
Warden, Jill (Department: 1743)
Chemistry: electrical and wave energy
Apparatus
Electrophoretic or electro-osmotic apparatus
C204S450000, C204S451000, C204S600000, C422S105000, C435S288500
Reexamination Certificate
active
06176991
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates to electrophoretic separation devices, and in particular, to a device having a serpentine separation channel, for example, in a microfabricated device.
BACKGROUND OF THE INVENTION
Electrophoresis exploits the differential rate of migration of charged species through a separation medium, under the influence of an electric field, for purposes of separating and/or characterizing physical properties of the charged species. Typically, the sample containing the charged species to be separated is placed at one end of a separation channel (which may be a linear channel or a lane in a 2-dimensional slab) and a voltage difference is placed across opposite channel ends until a desired migration end point is reached. The separated analyte molecules may then be detected, e.g., by optical detection, radiography, or band elution.
As examples, gel electrophoresis in the presence of a charged surfactant, such as dodecyl sulfate, is widely used for protein separation and for characterizing protein molecular weight. Electrophoresis in a gel or liquid medium is commonly used to separate oligonucleotides with different numbers of bases, for example, in DNA sequencing.
One of the possible applications of microfabrication techniques that has been proposed is in the area of column separation devices, including electrophoresis devices. Jacobsen, et al. (
Anal. Chem.
66:2369 (1994);
Electrophoresis
16:481 (1995) have described a “microchip” electrophoresis device formed by etching an open electrophoresis channel, and suitable connecting reservoirs, on a glass slide. Because of the small chip dimensions, typically less than 10-15 cm on a side, it is necessary to form the separation column in the form of a serpentine pathway in order to achieve total column separation lengths suitable for most applications.
Although a serpentine column solves the problem of adequate column length on a microchip, it introduces a potentially serious limitation in terms of column resolution. When a electrophoretic band is migrating through a linear channel, the molecules making up the band, which are all migrating at roughly the same speed, tend to migrate as a tight band. However, the same molecules migrating through a turn in a serpentine pathway will migrate through the shorter inner side of the channel faster than through the longer outer side of the channel, leading to band spreading and nonuniformity across the width of the channel. At each turn in the pathway, more band resolution is lost. Heretofore, this problem has severely limited the range of practical electrophoresis applications in a microchip format.
SUMMARY OF THE INVENTION
The application includes, in one aspect, an electrophoresis channel through which one or more charged species are intended to migrate under the influence of a voltage difference placed across opposite ends of the channel. The channel includes (i) a pair of channel segments disposed at an angle a with respect to one another, and (ii) an angled channel region connecting the two channel segments.
The angled channel region has a first curved channel portion subtending an angle &agr;
f
>&agr;, where &agr; is the angle between the two channel segments, and a second curved channel portion subtending an angle &agr;
s
=&agr;
f
−&agr;. The first curved portion defines inner and outer tracks or channel sides, such that an analyte migrating through the first channel portion under the influence of such voltage difference will traverse the inner track in a time interval &dgr;t
f
faster than that of the same analyte traversing the outer track. The second curved portion defines second inner and outer tracks such that an analyte migrating through the second channel portion under the influence of the same voltage difference will traverse the outer track in a time interval &dgr;t
s
faster than that of the same analyte traversing the inner track. The angles and cross-sections of the two channel portions are such that &dgr;t
f
is approximately equal to &dgr;t
s
.
The channel is typically part of a serpentine pathway containing a plurality of such segments, each pair of adjacent channel segments being connected by an associated angled channel region.
Where the two channel segments are disposed at right angles with respect to one another, &agr;
f
is preferably between about 110° and 160°, and &agr;
s
, between about 20° and 70°, respectively. Where the two channel segments are disposed substantially parallel to one another, &agr;
f
is preferably between about 200° and 250°, and &agr;
s
, between about 20° and 70°, respectively.
In a microfabricated chip format, the channel has preferred width dimensions between about 25-250 microns, and preferred depth dimensions between about 5-100 microns.
In one general embodiment, the first and second curved portions have substantially constant channel widths W
f
and W
s
, respectively, where W
f
<W
s
. In this embodiment, the angled channel region further includes tapered-width segments joining the second curved channel portion to the first channel portion and to one of the two channel segments. An approximate relationship between W
f
and W
s
is given by the relationship W
s
=(&agr;
f
W
f
2
R
f
/&agr;
s
R
s
)
½
, where R
f
and R
s
are the radii of curvature of the first and second curved portions, respectively.
In another general embodiment, the first curved channel portion has a preferably fixed channel width, and the second channel portion, a variable width that expands on progressing inwardly from each end.
In yet another embodiment, the first curved channel portion has a channel depth which increases on progressing toward the second channel portion, and the second curved channel portion has a channel depth which decreases on progressing away from the first curved channel portion. The channel width may be substantially constant in the channel segments and the channel connecting region therebetween.
More generally, the invention includes an analyte separation channel through which one or more analytes is intended to migrate under the influence of a motive force applied to opposite ends of the channel. The device includes (i) a pair of channel segments disposed at an angle &agr; with respect to one another, and (ii) an angled channel region of the type just described connecting the two channel segments. The motive force may be a voltage difference applied across the opposite ends of the channel, or a force producing fluid movement through the channel or a combination of the two.
In a related aspect, the invention includes a microfabricated device for electrophoretic separation of analytes in a mixture. The device includes a substantially planar-surface substrate having formed thereon, first and second reservoirs and a serpentine electrophoretic channel extending therebetween. The channel has a plurality of linear segments, and connecting the adjacent ends of each pair of adjacent segments, an angled channel region of the type described above. The channel, including the linear segments and angled channel regions, has preferred channel width dimensions between about 25-250 microns, and depth dimensions between about 5-100 microns.
These and other objects and features of the invention will become more fully apparent when the following detailed description of the invention is read in conjunction with the accompanying drawings.
REFERENCES:
patent: 5599432 (1997-02-01), Manz et al.
patent: 5958694 (1999-12-01), Nikiforov
patent: 5993750 (1999-11-01), Ghosh et al.
patent: 6033546 (2000-03-01), Ramsey
patent: 6033628 (2000-03-01), Kaltenbach et al.
patent: 91/16966 (1991-11-01), None
patent: 96/04547 (1996-02-01), None
Jacobson, S.C., et al., “Effects of Injection Schemes and Column Geometry on the Performance of Microchip Electrophoresis Devices”Anal. Chem. (1994) 66:1107-1113.
Jacobson, S.C., and Ramsey J.M., “Microchip electrophoresis with sample stacking,” Electrophoresis 16:481-486 (1995).
Jacobson, S.C., et al., “Open Channel Electrochromatography on a Microchip,” Anal. Che
Dehlinger Peter J.
Starsiak Jr. John S.
The Perkin-Elmer Corporation
Warden Jill
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