Chemistry: electrical and wave energy – Apparatus – Electrophoretic or electro-osmotic apparatus
Reexamination Certificate
2000-08-30
2001-07-10
Warden, Jill (Department: 1743)
Chemistry: electrical and wave energy
Apparatus
Electrophoretic or electro-osmotic apparatus
C204S451000, C204S452000, C204S601000
Reexamination Certificate
active
06258238
ABSTRACT:
FIELD OF THE INVENTION
The present invention deals quite generally with a method of producing a capillary, and in particular with a capillary of the type which is adapted to be used for an electrophoresis device.
BACKGROUND OF THE INVENTION
There are various techniques for rapidly and accurately identifying small quantities of inorganic and organic substances. When such substances, whose molecular structure is comparatively small, are to be detected, gas chromatographs and liquid chromatographs have been used for this detection for years. Although chromatography proved to be a successful attempt to determine metals and inorganic mixtures as well as small organic ions, this technique used for determining a constituent part in a sample is not suitable for determining large and complex molecules, such as amino acids, proteins and DNA.
Hence, another analysis technique, which is called electrophoresis, has been used with great success for some years. This technique comprises the step of applying an electric field across the length of a capillary that contains a mixture of an unknown sample and a non-reactive liquid, which is referred to as sample solution. The electric field causes the constituents of the unknown sample to migrate through the capillary due to the electrical attraction created by the electric field. Different components within the sample are attracted at different rates due to their varying molecular properties (molecular drag) and varying electrical charges. Hence, the substances become increasingly separated into distinct zones or groups as they progress along the capillary. Each band of constituent material that makes up the original, unseparated mixture of the unknown sample material passes through the capillary. At some point along the capillary, the band is examined and identified by a detector. One typical detector for electrophoresis separation measures the electrical conductivity of the bands in the capillary. An alternative detection scheme uses fluorescence, for example laser induced fluorescence. Although this technique is comparatively sensitive, it is costly and limited to specific mixtures that can be stimulated to fluoresce.
In other electrophoresis systems, an optical detection technique is used, which comprises the step of measuring the light absorption caused by the bands, which are separated from one another over the electrophoresis path. In conventional electrophoresis devices, the sensitivity of this type of optical detection is only very low due-to the short path that the light travels through the interior of the capillary. An increase in the inner diameter of the capillary is, however, disadvantageous, since otherwise turbulences or eddy currents might occur, whereby the separation of the constituent parts caused by electrophoresis would become null and void.
However, such known electrophoresis systems with capillaries having a uniform outer and inner diameter throughout their length entail also additional problems. The voltage applied to the capillary ends drops uniformly across the whole capillary, when the capillary is filled completely. The part of this voltage which will result in a separation of the substances of the sample is only the part that drops across the separation path between the sample input or rather the inlet of the capillary and the detector. The residual voltage drop between the detector and the outlet of the capillary does not contribute in any way to the separation and will only cause losses. It follows that, in connection with such systems, comparatively complicated and costly measures are normally taken so as to obtain the shortest possible length of the capillary between the detector and the outlet end of the capillary.
An additional problem of this technique resides in the fact that the advantage of a capillary inner width which is as small as possible, viz. low loss power in combination with high mass selectivity, can only be obtained at the price of a loss of detection sensitivity.
U.S. Pat. No. 5,061,361 disclosed for the first time an electrophoretic system in which high mass sensitivity and low loss power can be obtained without having to put up with a low detection sensitivity. In the electrophoresis system according to U.S. Pat. No. 5,061,361, the capillary has an enlarged inner width at least in the area of the detector device so that a longer, effective detection path of the light between the light source and the sensor of the detector device through the fluid containing the constituent part to be detected will be obtained without any necessity of enlarging the capillary inner width of the residual part of the capillary. U.S. Pat. No. 5,061,361 describes a method of producing a capillary of this type comprising the steps of closing a capillary, which initially has a constant outer width and a constant inner width, on one side thereof and applying a low pressure to the interior thereof, whereupon said capillary is locally heated and simultaneously rotated so that a local gas bubble, which has to be arranged in the area of the detector device, will be formed. This production method, which is based on the principle of glass-blowing, is comparatively complicated. Due to the enlargement of the outer width of the capillary in the area of the bubble, it is not possible to use such a modified capillary as a substitute for a standard capillary in commercially available capillary electrophoresis devices.
SUMMARY OF THE INVENTION
One object of the present invention is to provide a simplified method of producing a capillary whose inner width can be varied along its length.
An additional object of the present invention is to provide a capillary which is adapted to be used for electrophoresis devices and which, in spite of having a low loss power, permits a high detection sensitivity in combination with a good separation effect.
Another object of the present invention is to provide a capillary which is adapted to be inserted into existing electrophoretic devices as a substitute for standard capillaries having an invariable inner width so as to enhance the detection sensitivity of such existing electrophoresis devices.
Finally, it is an object of the present invention to provide an electrophoresis device having a high detection sensitivity in combination with a low loss power and a high mass selectivity.
In accordance with a first aspect of the invention, this object is achieved by a method of producing a capillary comprising a first capillary area having a first inner width and a second capillary area having a second inner width which is enlarged in comparison with said first inner diameter, said method including the following steps:
providing a capillary which has an essentially uniform inner width over its length,
introducing into the interior of the capillary an etchant by means of which the material of the capillary can be etched,
producing a temperature gradient over the capillary in such a way that the temperature of the second capillary area is higher than the temperature of the first capillary area, and
carrying out the etching process until the second capillary region has the desired inner width.
In a preferred embodiment, the method step of carrying out the etching process comprises the measure of causing the etchant to flow through the capillary.
In a further preferred embodiment, the method step of producing the temperature gradient comprises the following steps:
introducing one of the two capillary areas of the capillary into a hose,
introducing the other capillary area into a bath having a predetermined temperature,
passing through said hose a medium whose temperature differs from that of the bath.
In a further preferred embodiment, the bath is an ice water bath, and the medium passing through the hose is steam.
In a further preferred embodiment, the etchant flowing through the capillary is supplied to the capillary with overpressure.
In a further preferred embodiment, the capillary consists of quartz glass and the etchant is hydrofluoric acid HF.
In accordance with a second aspect of the invention, this
Büttner Christian
D T. Beck Wolfgang
Agilent Technologie,s Inc.
Starsiakm Jr. John S.
Warden Jill
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