JP2006317467A - Modified siphons for improving metering precision - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a centrifugal rotor for delivering a premeasured volume of liquid to a chamber in the rotor. <P>SOLUTION: The rotor, in particular, comprises a siphon 134 for delivering the premeasured volume of liquid between a first chamber and a second chamber 136 in the rotor. The siphon 134 is designed, such that the inlet 138 of the siphon on the first chamber is directed radially outward of the siphon outlet 139 on the second chamber 136. The first chamber is emptied to a level equivalent to the radial position of the siphon outlet 139. <P>COPYRIGHT: (C)2007,JPO&INPIT

Description

  This application is a continuation-in-part of pending US patent application Ser. No. 08 / 115,162, incorporated herein by reference.

  The present invention relates generally to an apparatus and method for analyzing biological fluids. In particular, it relates to the design and use of an improved centrifugal rotor having a siphon that allows a precise amount of liquid to be pumped into a chamber within the rotor.

  Biological testing of plasma and other biological fluids often requires that these fluids be rapidly divided into predetermined amounts for analysis with a wide variety of optical tests or assays. It is often also desirable to separate the potentially interfering cytoplasmic components of this material from other liquids before testing. Such measurement and separation steps are typically typically performed, for example, using centrifugation to separate plasma from cytoplasmic components, and then manually or automatically placing a predetermined amount of plasma in separate test reservoirs. It was done by pipetting. Such a procedure is labor intensive and time consuming. As a result, various automated systems and methods have been proposed to obtain multiple aliquots that are suitable for more efficient testing.

  A major advance in the analysis of biological fluids has been the use of centrifugal rotors. These rotors are designed to measure the amount of biological fluid, remove cytoplasmic components, and mix this fluid with a suitable diluent for analysis, eg, by optical testing. Typically, these rotors place multiple separate amounts of sample in separate cuvettes in which the sample is optically analyzed.

  In order to ensure accurate and consistent results, such rotors need to deliver precisely measured amounts of liquid to the various chambers of the rotor. This often must be done in an environment where the rotor is rapidly accelerated or decelerated during operation or otherwise confused. This confusion often can lead to inaccurate measurements. The present invention addresses these and other needs.

  U.S. Pat. Nos. 4,894,204 and 5,160,702 disclose siphons for transferring liquid between chambers in a rotor. U.S. Pat. No. 4,244,916 discloses a rotor that includes a plurality of cuvettes disposed radially outward of a central receptacle. Each cuvette is connected to this central receptacle by a duct and includes another air escape orifice. U.S. Pat. No. 4,314,968 relates to a rotor having cells arranged on the outer periphery of the rotor. Each cell includes a peripheral orifice for removing liquid introduced into the cell. U.S. Pat. No. 4,902,479 discloses a multi-cuvette rotor that includes a long, radially extending cuvette. Each long cuvette includes a first chamber for receiving a first component and a second chamber for receiving a second component. The partition structure between the first chamber and the second chamber prevents mixing of these components before a predetermined time. Mixing occurs when the rotor rotates at a sufficient speed. U.S. Pat. No. 4,963,498 discloses an apparatus that relies on capillaries, chambers, and orifices to pump and mix liquids for optical analysis. U.S. Pat. No. 5,077,013 discloses a rotor including a peripheral cuvette connected to a holding chamber disposed radially inward.

  The present invention provides a centrifugal rotor that includes a siphon for delivering a pre-measured amount of a liquid, typically a biological sample such as plasma, between a first chamber and a second chamber in the rotor. . The siphon of the present invention has an elbow that is radially inward from the innermost point in the radial direction of the liquid in the first chamber. As the rotor rotates, no liquid flows through the elbow. After the rotor has stopped, the capillary force "primes" the siphon by pulling the liquid immediately around the elbow, i. When the rotor is restarted, centrifugal force pulls the remaining liquid from the metering chamber into the receiving chamber until the liquid level in the metering chamber is the same radial distance as the siphon outlet. The siphon of the present invention is designed so that the inlet of this siphon above the first chamber is radially outward from the siphon outlet above the second chamber.

  This arrangement of the inlet and outlet of the siphon of the present invention produces many advantages. For example, the siphon inlet is always radially outward from the final position of the liquid meniscus in the first chamber after the liquid has been transferred to the second chamber. Thus, this meniscus is minimized so that measurement inaccuracies associated with different liquids and different meniscus shapes are minimized. Moreover, those skilled in the art will know that all the siphons are semi-stable, although the fluid connection in the siphon is stable, but if the rotor is confused it is easily lost. When this liquid connection is broken by centrifugal force, the liquid in the siphon will flow radially to the outermost point. In the prior art siphon, this is the siphon outlet. Thus, an unmeasured amount of liquid may be delivered to the receiving chamber. In the siphon of this invention, the outermost point in the radial direction of the siphon is the siphon inlet. In this design, when the liquid connection is broken, the liquid flows back into the first chamber, thereby avoiding the problem of delivering this unmeasured amount of liquid.

  The chambers connected by the siphon of the present invention can be used for a number of functions such as liquid metering, separation of solid components from the sample, mixing of sample and diluent. In the preferred embodiment, the siphon connects the plasma metering chamber to the mixing chamber to mix a pre-measured amount of plasma with the diluent.

  Moreover, the rotor of the present invention includes an unmodified inlet channel that connects the distribution ring to a cuvette that contains reagents for optical analysis of biological samples. These inlet channels are sized such that as the rotor rotates, liquid escapes through the inlet channel as liquid enters the cuvette through the inlet channel. An “uncorrected inlet channel” as used herein is typically not modified (eg, by changing cross-sectional shape, surface texture, etc.) to create a passage for gas that is not otherwise bleed to escape from the cuvette. In particular, it refers to a simple inlet channel with a rectangular cross section.

  The present invention provides a method and apparatus for delivering liquid to the chamber of an analytical rotor. The rotor of the present invention includes a siphon that ensures that a metered amount of liquid is accurately delivered to a desired chamber within the rotor.

  The rotor of this invention is suitable for the analysis of any liquid, typically a biological sample such as whole blood or plasma. It is also useful for many other biological fluids such as urine, sputum, semen, saliva, ocular lens fluid, brain fluid, cerebrospinal fluid, amniotic fluid. Other liquids that can be tested include tissue culture media, food and industrial chemicals, environmental samples, and the like.

  The rotor typically separates cellular components from a biological sample (eg, whole blood), measures the exact amount of a liquid sample (eg, plasma), and mixes this sample with an appropriate diluent. And a chamber in which the diluted sample can be delivered to a cuvette for optical analysis. The liquid delivered to these cuvettes undergoes reactions in these cuvettes, for example, with reagents that form part of an analytical procedure for detecting one or more analytes. This sample may be further optically analyzed while in the rotor, regardless of previous reactions.

  The apparatus of the present invention includes an analytical rotor having a rotor body that can be attached to a conventional commercial laboratory centrifuge, the centrifuge being a spinco division of Beckman Instruments, Inc., Fullerton, California. Available from vendors such as Fisher Scientific, Pittsburgh, Pennsylvania; VWR Scientific, San Francisco, California; Generally, the centrifugal rotor has a receptacle or other coupling device suitable for mounting on a vertical drive shaft included in the centrifuge. The specific design of the receptacle or coupling device depends on the nature of the centrifuge, so that the centrifugal rotors of the invention can be made available now or in the future for all or most types of centrifuges You will find that you can adapt it for use.

  The rotor body has a structure that maintains a desired geometric style or relationship between the plurality of chambers, interconnecting passages, and vents, as described in more detail below. Various specialized chambers and channels suitable for use in the rotor of the present invention are described in U.S. Pat. Nos. 5,061,381 and 5,122,284 and U.S. patent application no. 07 / 678,762 and 07 / 783,041.

  Typically, the body is a substantially strip or disk that has channels and passages formed as spaces or cavities in the strip. Such a strip can be conveniently formed by laminating a plurality of separate layers together as a composite structure, and creating a chamber and a horizontal passageway generally between adjacent layers. Vertical passages may be made through these layers. The individual layers may be made by injection molding, machining, or combinations thereof and are typically joined together using a suitable adhesive or by ultrasonic welding. Final encapsulated volumes are created when these layers are brought together.

  Of course, this centrifugal rotor can also be made by arranging a plurality of individual parts, such as tubes, containers, chambers, etc., in a suitable framework. However, such individual component assemblies are generally more difficult to manufacture and therefore less desirable than those made in substantially stripped boards.

  The rotor body may be made from a wide variety of materials and may optionally include more than one material. Typically these materials are transparent so that the presence and distribution of biological fluids, cellular components, and reagents in the various internal chambers and passages can be observed. Optionally, the contents of these cuvettes can be analyzed spectroscopically, fluorimetrically, or other optically, as long as an analytical room, for example a cuvette, or other test reservoir is made in this rotor. It is desirable for the rotor to have an appropriate optical path so that the evaluation instrument can be observed. A suitable cuvette configuration in which a special optical path therethrough is made is disclosed in US Pat. No. 5,173,193, incorporated herein by reference. In this preferred embodiment, the rotor is made of an acrylic resin having suitable optical properties, at least in the region forming the optical path.

  The devices and methods of this invention are suitable for performing a wide variety of analytical procedures and assays that are beneficially or necessarily performed on plasma and other samples. These analytical procedures involve the sample with one or more reagents in order to cause certain detectable changes that may be related to the presence and / or amount of a particular component (analyte) or the characteristics of the sample. May require combining. For example, the sample may undergo reactions or other changes that result in changes in color, fluorescence, luminescence, etc. that can be measured with conventional spectrophotometers, fluorometers, photodetectors, and the like. In some cases, immunoassays and other specific binding assays may be performed in a cell-free liquid collection chamber or in a cuvette coupled to this collection chamber. In general, such an assay procedure should be homogeneous and should not require a separation step. However, in other cases, the heterogeneous assay can be performed by providing a means for separating a sample (eg, plasma) from this collection chamber or another test reservoir or cuvette after the immunological reaction step has taken place. It may be possible to accommodate the system. One skilled in the art will recognize that the means for analyzing the sample is not an important aspect of the invention. Any of a number of analytical methods can be adapted for use with the rotor of the present invention, depending on the particular sample to be analyzed and the components to be detected.

  For blood analysis, a conventional blood assay is typically performed. Examples of assays that can be performed include glucose, lactate dehydrogenase, serum glutamate-oxaloacetate transaminase (SGOT), serum glutamate-pyruvate transaminase (SGPT), blood urea nitrogen (BUN), total protein, alkalinity, phosphatase Some are designed to detect bilirubin, calcium, chloride, sodium, potassium, magnesium, and the like. This table is not exhaustive and is merely provided as an example of an assay that can be performed using the apparatus and method of the present invention. Typically, these tests require the combination of blood and plasma with one or more reagents, which produce changes in the plasma that are optically detectable, usually detectable with a photometer. These necessary reagents are well known and are described extensively in the patent and scientific literature.

These reagents are preferably prepared in lyophilized form to increase stability. Ideally, they are provided in the form of lyophilized reagent spheres as described in US patent application Ser. No. 07 / 747,179, incorporated herein by reference.
Referring now to FIGS. 1A-1F, an analytical rotor including the channel of the present invention can be seen. FIG. 1A shows the position of this sample within the blood application chamber 104 after the blood sample 102 has been placed in the rotor body 100. The diluent container in the chamber 106 is connected to the centrifuge as described in the co-pending, co-assigned US patent application Ser. No. 07 / 873,327, incorporated herein by reference. Opens when attached to the spindle.

  FIG. 1B shows the position of diluent 108 and blood sample 102 after rotating the rotor at 4,000 rpm. Blood sample 102 begins to exit blood application chamber 104 and enters plasma metering chamber 110. At the same time, the diluent 108 is poured from the diluent container into the holding chamber 112. This diluent immediately begins to enter diluent diluent chamber 114 through channel 116.

  FIG. 1C shows the position of these liquids as the rotor 100 continues to rotate. Here, the blood sample 102 empties the blood application chamber 104, overflows from the plasma measurement chamber 110 to the overflow chamber 118, and flows from there to the hemoglobin cuvette 120 and the excess blood disposal site 122. On the other hand, diluent 108 fills diluent metering chamber 114, with excess flowing through channel 124 to diluent-only cuvette 126 and excess diluent dump 127.

  FIG. 1D shows the position of these liquids at the end of this first rotation. Blood sample 102 is divided into cells 128 and plasma 130. The diluent only cuvette 126 is filled and a predetermined amount of diluent remains in the diluent metering chamber 114. Next, the rotor 100 stops and the siphon 132 from the diluent metering chamber 114 and the siphon 134 from the plasma metering chamber 110 are primed as described above. Siphon 134 is the siphon of the present invention. It is connected to the plasma metering chamber 110 at the inlet 138. The inlet 138 is located radially outward from the siphon outlet 139, and the siphon 134 communicates with the mixing chamber 136 through the outlet.

  FIG. 1E shows the position of these liquids during the second rotation of the rotor. Diluent metering chamber 114 leads to mixing chamber 136 through siphon 132. A predetermined amount of plasma 130 is metered and supplied to the mixing chamber 136, and the two liquids are mixed, thereby producing diluted plasma 131. The amount of plasma 130 delivered to the mixing chamber 136 is determined by the position of the outlet 139 on the siphon 134. As can be seen in this figure, the final height 133 of the plasma in the plasma metering chamber 110 is at the same radial position as the outlet 139. Thus, the amount of plasma delivered to the mixing chamber 136 is determined by the volume between the outlet 129 to the overflow chamber of the plasma metering chamber 110 and the final height 133 of the plasma. After the plasma and diluent are mixed in the mixing chamber 136, the rotor stops again and the outlet siphon 140 is primed.

  FIG. 1F shows the position of diluted plasma 131 as it rotates during the third rotation of the rotor. This figure shows that diluted plasma 131 travels through distribution ring 142 and inlet channel 144 to cuvette 146 and surplus plasma dump 147. The resistance to flow in the outlet siphon 140 is chosen to be higher than the resistance to flow in the distribution ring 142 and inlet channel 144 so that the air in the cuvette 146 can escape as these cuvettes are filled. In particular, the siphon 140 is dimensioned such that the ratio of the cross-sectional area of the inlet channel 144 to the cross-sectional area of the liquid therein is 2: 1 or higher, preferably 4: 1 or higher. The cross-sectional area of the inlet channel 144 is typically the same or slightly smaller than that of the distribution channel 142 so that gas in the cuvette that is not bleed escapes through the inlet channel 144 and the distribution channel 142. If the sample is plasma or diluted plasma and the channel cross section is rectangular, their dimensions are typically as follows: siphon: depth 0.150 mm, width 0.200 mm; distribution channel depth 0.300 mm, width 0.500 mm; inlet channel: depth 0.150, width 0.500.

  After filling the cuvettes, the reagents in these cuvettes are mixed with the solution and the required photometric analysis is performed on the sample. Such an analysis is performed according to methods known to those skilled in the art as described above.

  Although the foregoing invention has been described in detail for purposes of clarity of understanding, it will be apparent that modifications can be made that are within the scope of the appended claims.

FIG. 3 is a plan view of the rotor of the present invention showing the flow of liquid through the rotor chamber and channel as the rotor rotates. FIG. 3 is a plan view of the rotor of the present invention showing the flow of liquid through the rotor chamber and channel as the rotor rotates. FIG. 3 is a plan view of the rotor of the present invention showing the flow of liquid through the rotor chamber and channel as the rotor rotates. FIG. 3 is a plan view of the rotor of the present invention showing the flow of liquid through the rotor chamber and channel as the rotor rotates. FIG. 3 is a plan view of the rotor of the present invention showing the flow of liquid through the rotor chamber and channel as the rotor rotates. FIG. 3 is a plan view of the rotor of the present invention showing the flow of liquid through the rotor chamber and channel as the rotor rotates.

Claims (19)

A centrifugal rotor having a rotor body, the rotor body having a liquid dispensing chamber, a liquid receiving chamber, and a siphon;
The siphon is connected to the liquid dispensing chamber via a siphon inlet, and is connected to the liquid receiving chamber via a siphon outlet, the siphon inlet being radially outward of the siphon outlet; and the siphon A centrifugal rotor that forms an elbow structure by first extending radially inward at a point radially inward of the siphon inlet and then radially outward at the siphon outlet. A centrifugal rotor having a rotor body, the rotor body having a liquid dispensing chamber, a liquid receiving chamber, and a siphon;
The siphon is connected to the liquid dispensing chamber via a siphon inlet and is connected to the liquid receiving chamber via a siphon outlet, the siphon inlet being radially inward of the siphon outlet; and the siphon Form an elbow structure by first extending radially outward of the liquid dispensing chamber, then radially inward to a point radially inward of the siphon inlet, and then radially outward to the siphon outlet. Centrifugal rotor. A centrifugal rotor having a rotor body, the rotor body having a liquid dispensing chamber, a liquid receiving chamber, and a siphon;
The siphon is connected to the liquid dispensing chamber via a siphon inlet, and is connected to the liquid receiving chamber via a siphon outlet, the siphon inlet being radially outward of the siphon outlet; and the siphon Form an elbow structure by first extending radially inward at a point radially inward of the siphon inlet and then radially outward at the siphon outlet;
The rotor is
A distribution ring disposed radially outward of the liquid receiving chamber;
A centrifugal rotor further comprising a delivery channel connecting the distribution ring to the liquid receiving chamber, wherein the distribution ring is connected to a cuvette via an inlet channel. A centrifugal rotor having a rotor body, the rotor body having a liquid dispensing chamber, a liquid receiving chamber, and a siphon;
The siphon is connected to the liquid dispensing chamber via a siphon inlet and is connected to the liquid receiving chamber via a siphon outlet, the siphon inlet being radially inward of the siphon outlet; and the siphon Form an elbow structure by first extending radially outward of the liquid dispensing chamber, then radially inward to a point radially inward of the siphon inlet, and then radially outward to the siphon outlet. ,
The rotor is
A distribution ring disposed radially outward of the liquid receiving chamber;
A centrifugal rotor further comprising a delivery channel connecting the distribution ring to the liquid receiving chamber, wherein the distribution ring is connected to a cuvette via an inlet channel. A liquid dispensing chamber;
An unbleeded liquid receiving chamber disposed radially outward from the liquid dispensing chamber;
A delivery channel connected to the liquid dispensing chamber for removing liquid from the liquid dispensing chamber under centrifugal force;
A distribution ring directly connected to the delivery channel for removing liquid from the delivery channel under centrifugal force;
A centrifugal rotor having an inlet channel connected to the unbleeded liquid receiving chamber for removing liquid from the liquid dispensing chamber via the delivery channel and the distribution ring,
The resistance to liquid flow in the delivery channel is greater than the resistance to liquid flow in the inlet channel and the distribution ring, so that the cross-sectional area of the liquid flowing through the inlet channel and the distribution ring Being smaller than the cross-sectional area of the ring and rotating the rotor acts to cause the liquid to flow from the liquid dispensing chamber through the delivery channel, the distribution ring and the inlet channel to the receiving chamber. A centrifugal rotor, characterized in that when the liquid enters the receiving chamber, air escapes from the receiving chamber through the inlet channel.   The rotor according to any one of claims 3 to 5, wherein the delivery channel is a siphon.   The rotor according to claim 3 or 4, wherein the cuvette contains a reagent necessary for analysis of a biological sample, and the cuvette is radially outside the liquid receiving chamber.   The rotor according to any one of claims 3 to 5, wherein the inlet channel has at least a cross-sectional area of about 1.5 times the cross-sectional area of the delivery channel.   The rotor according to any one of claims 3 to 5, wherein the inlet channel has at least a cross-sectional area of about twice the cross-sectional area of the delivery channel. The rotor according to any one of claims 3 to 5, wherein a cross-sectional area of the delivery channel is about 0.03 mm ² .   The rotor according to claim 1, wherein the liquid distribution measurement chamber is a plasma measurement chamber and the liquid receiving chamber is a mixing chamber.   The rotor according to claim 1, wherein the liquid distribution measurement chamber is a diluent measurement chamber, and the liquid receiving chamber is a mixing chamber.   The rotor according to claim 5, wherein the liquid distribution measuring chamber is a mixing chamber.   The rotor according to claim 5, wherein the liquid receiving chamber is a cuvette containing a reagent necessary for analysis of a biological sample.   The rotor according to claim 5, further comprising a diluent dividing chamber that is disposed radially inward of the liquid dispensing and metering chamber and connected to the liquid dispensing and metering chamber via a siphon.   The rotor according to claim 5, further comprising a plasma metering chamber disposed radially inward of the liquid dispensing metering chamber and connected to the liquid dispensing metering chamber via a siphon. In the method of delivering a pre-measured amount of liquid from the first chamber to the second chamber in the rotor,
Rotating a rotor, thereby introducing an unmeasured amount of liquid into the first chamber, the rotor comprising a first chamber having a first volume, a second chamber, and a siphon inlet through the siphon inlet; A siphon connected to the first chamber and connected to the second chamber via a siphon outlet, wherein the siphon inlet is radially outward of the siphon outlet, and the siphon is first connected to the siphon inlet Forming an elbow structure by extending radially inward to a radially inner point and then radially outward to the siphon outlet; and
Stopping the rotation of the rotor, thereby priming the siphon connecting the first chamber to the second chamber;
Rotating the rotor, thereby initiating operation of the siphon and delivering a pre-measured amount of liquid from the first chamber to the second chamber. In the method of delivering a pre-measured amount of liquid from the first chamber to the second chamber in the rotor,
Rotating a rotor, thereby introducing an unmeasured amount of liquid into the first chamber, the rotor comprising a first chamber having a first volume, a second chamber, and a siphon inlet through the siphon inlet; A siphon connected to the first chamber and connected to the second chamber via a siphon outlet, wherein the siphon inlet is radially inward of the siphon outlet, the siphon initially being the liquid dispensing Forming an elbow structure by extending radially outward to a point radially inward of the chamber, then radially inward to a point radially inward of the siphon inlet, and then radially outward to the siphon outlet; and ,
Stopping the rotation of the rotor, thereby priming the siphon connecting the first chamber to the second chamber;
Rotating the rotor, thereby initiating operation of the siphon and delivering a pre-measured amount of liquid from the first chamber to the second chamber.   The method of claim 17, wherein the pre-measured amount is determined by a radial position of the siphon outlet and the first volume of the first chamber.

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