Patent Application
20070110628
The present invention relates to an apparatus and method for dispensing low volumes of an analyte solution for subsequent testing or analysis.
Apparatus used in analytical chemistry include mass spectrometers. One type of mass spectrometer is a matrix assisted laser desorption ionization (MALDI) mass spectrometer of which there are also several types. MALDI is typically operated as an off-line ionization technique, where a solid or liquid sample, mixed with a suitable matrix, is deposited on a MALDI target or medium to form dry mixed crystals and, subsequently, placed in a source chamber of the mass spectrometer for analysis. An example of a MALDI target is a rectangular plate having a plurality of microfabricated wells on an upper surface thereof for receiving pL-nL sample volumes of analyte solution. The analyte solution may be generated by a variety of separation or processing techniques or apparatus including liquid chromatography apparatus.
Liquid chromatography (LC) involves the separation of chemical substances and particles by differential movements through a two phase system. A mixture of materials is typically applied to a column containing a suitable chosen absorbent (e.g. an ion-exchange material) and caused to flow therethrough. Materials in the mixture are absorbed at differential rates, with the least absorbed materials emerging first from the column and the more strongly absorbed materials emerging later.
When analyzing liquids eluted from an LC apparatus using a mass spectrometer, it is important to dispense very low volume samples (e.g. pL-nL samples) with no or minimal carry-over. That is, it is desirable to keep the components of a sample as well as the components of subsequent samples from mixing together when being deposited on a sample target or medium. While apparatus and methods exist for dispensing low volumes of liquid, there is a problem with carryover of liquid samples being dispensed. The present invention is intended to address this problem.
In accordance with a first aspect of the invention, there is provided an apparatus for dispensing fractions of an analyte solution, the apparatus comprising:
The ejection cavity may have a volume of between 0.1 nl and 1000 nl, or less than 500 nl, 200 nl, 100 nl, or 50 nl.
The dispensing mechanism may comprise an actuator chosen from solenoid, piezoelectric, electro kinetic, mechanical, valve, thermal, magnetic, and pressurized fluid actuators. Furthermore, the analyte generating apparatus may comprise a component chosen from liquid chromatography, capillary electrophoresis, and capillary electro chromatography apparatus.
The dispensing mechanism may further include an aspirator operable to selectively reverse the flow of buffer from and out of the ejection cavity and upstream along the second fluid conduit.
The outlet of the apparatus may be circular and have a diameter of from 5 to 200 micrometers, or less than 100, 50, or 20 micrometers.
In one embodiment, the apparatus is used to dispense fractions of an analyte solution onto a collection medium for subsequent testing by matrix assisted laser desorption ionization, and includes a matrix flow generator, the matrix flow generator having a matrix supply and a third conduit having a first end in communication with the matrix supply and a second end in communication with the ejection cavity, the matrix flow generator being operable to deliver matrix through the third conduit to the ejection cavity, whereby a mixture of the analyte solution and matrix is dispensed when the dispensing mechanism is actuated. The second end of the third conduit may be in communication with the first flow path upstream of the ejection cavity whereby mixing of the matrix and analyte may occur upstream of the ejection cavity.
In accordance with another aspect of the invention, there is provided a method for dispensing fractions of an analyte solution comprising the steps of:
The analyte solution may be eluted material exiting a liquid chromatography apparatus, and may also be supplied to the ejection cavity at a rate of from 1 nl per minute to 2 ml per minute, from 10 nl per minute to 5000 nl per minute, or from 50 nl per minute to 2000 nl per minute.
Embodiments of the invention will now be described with reference to the drawings in which:
As shown in
In this embodiment, the LC fractions are later analyzed using a matrix assisted laser desorption (MALDI) apparatus. Thus, the apparatus 10 includes a matrix flow generator in the form of a syringe pump 40 containing a matrix solution. Alternatives to the syringe pump include any other suitable pump which is effective to supply a flow of matrix to the ejection cavity. The person skilled in the art would understand which materials would be suitable for use as matrices in MALDI applications. Common matrix components include alpha-cyano hydroxy cynnamic acid, 2,5-dehydroxy benzoic acid, sinappinic acid, succinic acid, glycerol, and picolinic acid. The matrix flow generator includes a conduit 42 having a first end 44 in communication with the matrix supply and a second end 46 in communication with the ejection cavity 14. The matrix flow generator 40 is operable to deliver matrix through the matrix conduit 42 to the ejection cavity 14 via the first fluid flow path 22, whereby a mixture of the analyte solution and matrix is formed upstream of the ejection cavity 14 and enters the ejection cavity 14. The mixture is then dispensed when the dispensing mechanism 30 is actuated.
In this embodiment, the dispensing mechanism 30 comprises an aspirator 48 which is selectively operable to reverse the flow of buffer solution 32 from and out of the ejection cavity 14 and upstream along the second fluid conduit 20. The aspiration is effected by a second solenoid actuator 50, as is also known in the art. When the second actuator 50 is actuated, the buffer solution 32 flows into a waste conduit 52 from the second fluid conduit 20 and into a waste reservoir in the form of a waste container 5 containing waste buffer solution. In this embodiment, the waste container 54 has an internal pressure of from 0 to 15 psi.
In use, the liquid chromatography apparatus 26 delivers LC fractions eluted from the LC column through the LC conduit 28 to the ejection cavity 14 via the first fluid flow path 22. The solution is delivered at a rate of 100 nl per minute. Matrix solution is dispensed continuously at a similar rate and a mixture of matrix solution and LC fractions arrives at the ejection cavity at a rate of 200 nl per minute. In this embodiment, the ejection cavity has a volume of 20 nl. Since the rate of flow of analyte solution into the ejection cavity 14 is known, and since the volume of the ejection cavity 14 is known, the time within which the ejection cavity 14 is filled with matrix-containing analyte solution can be easily determined. The apparatus 10 includes a timer which is used to actuate the dispensing mechanism 30 to cause buffer solution to flow into the ejection cavity at select intervals to dispense the analyte solution after the ejection cavity 14 is filled. The buffer solution serves to clean the ejection cavity 14 during dispensing so as to prevent cross-contamination of LC fractions being dispensed. Because the outlet 16 does not come in contact with the steel plate 38, the risk of cross-contamination of samples being dispensed is greatly reduced. To make room for the next sample to be dispensed, the second solenoid actuator 50 is actuated after each dispensing operation to aspirate the buffer and cause it to leave the ejection cavity 14. Eluted material flowing from the LC column may then flow into the ejection cavity 14 after exit of the buffer.
Referring now to
While specific embodiments have been described, the person skilled in the art will appreciate that many modifications may be made to the present invention. For example, the ejection cavity 14 may have a volume of anywhere between 0.1 and 1000 nl, less than 500 nl, less than 200 nl, or less than 100 nl.
Furthermore, the dispensing mechanism may be any one of a number of direct or indirect actuators, some of which are shown in
Instead of a liquid chromatography apparatus, any apparatus which operates to generate analyte solution to be dispensed may be used, including capillary electrophoresis and capillary electro chromatography apparatus. It will be appreciated that the present apparatus may be used to dispense low volume samples of any test solution.
In the above examples, buffer is injected into the ejection cavity 14 once the ejection cavity is filled with analyte solution. However, buffer may be injected prior to the ejection cavity 14 being filled completely to dispense volumes less than the internal volume of the ejection cavity. It will also be appreciated that an analyte solution may be supplied to the ejection cavity 14 at varying rates. Typically, solution will be supplied at a rate of from 1 nl per minute to 2 ml per minute. The typical rate of flow of eluted material out of the liquid chromatography apparatus is between 50 nl/min and 1 ml/min and often between 50 nl/min and 5000 nl/min for nano/micro LC applications. Furthermore, any non-reactive gas may be used as the gaseous buffer, including nitrogen, argon and helium.
The aspiration portion of the dispensing mechanism 30 is optional even in the case in which the buffer is a liquid. For example, the pressure generator may be selected and configured to inject only enough buffer so as to displace an amount of analyte solution equivalent to the volume of the meniscus of analyte solution forming at the outlet. In this case, the ejection cavity 14 would still contain mostly analyte solution after each dispensing operation and there would be no need to aspirate the buffer to make room for analyte solution entering the ejection cavity 14.
Furthermore, the dispensing and aspiration functions of the dispensing mechanism 30 may be combined in a single device. For example, an electrokinetic pump may be used to cause liquid to flow in opposite directions depending on the polarity of the voltage applied. Thus, when dispensing, the voltage of the pump will be of one polarity, and when aspirating, the voltage will be of the opposite polarity. There are other devices which may be used in place of an electrokinetic pump to achieve this combined function. The person skilled in the art will readily understand which alternative devices would be suitable in the present application.
The foregoing description is by way of example only and shall not be construed so as to the limit the scope of the following claims.
| Filing Document | Filing Date | Country | Kind | 371c Date |
|---|---|---|---|---|
| PCT/CA04/00792 | 5/28/2004 | WO | 1/23/2007 |
| Number | Date | Country | |
|---|---|---|---|
| 60473932 | May 2003 | US |
