US 8,007,724 B2 · Electrophoresis apparatus having at least one auxiliary buffer passage · patentgrep

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Abstract

An electrophoresis apparatus is generally disclosed for sequentially analyzing a single sample or multiple samples having one or more analytes in high or low concentrations. The apparatus comprises a relatively large-bore transport capillary which intersects with a plurality of small-bore separation capillaries and includes a valve system. Analyte concentrators, having antibody-specific (or related affinity) chemistries, are stationed at the respective intersections of the transport capillary and separation capillaries to bind one or more analytes of interest. The apparatus allows the performance of two or more dimensions for the optimal separation of analytes. The apparatus may also include a plurality of valves surrounding each of the analyte concentrators to localize each of the concentrators to improve the binding of one or more analytes of interest.

Drawings

Claims54

Dependent claims

An electrophoresis apparatus, comprising:

a transport passage;
a first analyte concentrator including first affinity ligands which are attracted to a first analyte of interest;
the first affinity ligands being immobilized to a surface within the first analyte concentrator;
a first separation passage to convey the first analyte of interest from a sample introduced into the transport passage and concentrated by the first analyte concentrator at a first location of the transport passage to an analyte identification and characterization detection zone where the first analyte of interest can be identified and characterized;
the first separation passage being communicable upstream of the first location with a buffer supply;
a first auxiliary passage through which a separation buffer can be provided to the first separation passage, and positioned downstream of the first analyte concentrator and positionable upstream of the detection zone;
a second analyte concentrator including second affinity ligands which are attracted to a second analyte of interest;
the second affinity ligands being immobilized to a surface within the second analyte concentrator;
a second separation passage to convey the second analyte of interest from the sample introduced into the transport passage and concentrated by the second analyte concentrator at a second location of the transport passage to the detection zone where the second analyte of interest can be identified and characterized;
the second separation passage being communicable upstream of the second location with a separation or elution buffer supply;
a second auxiliary passage through which a separation buffer can be provided to the second separation passage, and positioned downstream of the second analyte concentrator and positionable upstream of the detection zone;
a valve system for respectively controlling the flow in the transport passage and in the first and second separation-passages and controlling flow of separation buffer from the first auxiliary passage to the first separation passage and from the second auxiliary passage to the second separation passage independently and sequentially;
and
a first auxiliary analyte concentrator on the first separation passage and downstream of the first analyte concentrator, the first auxiliary analyte concentrator having affinity ligands capable of retaining chromophores to bind the first analyte of interest released from the first analyte concentrator to improve the sensitivity and selectivity of the first analyte of interest once the first analyte of interest is released from the first auxiliary analyte concentrator into the first separation passage and towards the detection zone wherein the first auxiliary analyte concentrator is upstream of the first auxiliary passage;
and
a second auxiliary analyte concentrator on the second separation passage and downstream of the second analyte concentrator, the second auxiliary analyte concentrator having affinity ligands capable of retaining chromophores to bind the second analyte of interest released from the second analyte concentrator to improve the sensitivity and selectivity of the second analyte of interest once the second analyte of interest is released from the second auxiliary analyte concentrator into the second separation passage and towards the detection zone wherein the second auxiliary analyte concentrator is upstream of the second auxiliary passage.

13 more elements

An electrophoresis apparatus, comprising:

a transport passage;
a first analyte concentrator including first affinity ligands which are attracted to a first analyte of interest, the first analyte concentrator comprises a first concentrator-microreactor;
the first affinity ligands being immobilized to a surface within the first analyte concentrator;
a first separation passage to convey the first analyte of interest from a sample introduced into the transport passage and concentrated by the first analyte concentrator at a first location of the transport passage to an analyte identification and characterization detection zone where the first analyte of interest can be identified and characterized;
the first separation passage being communicable upstream of the first location with a buffer supply;
a first auxiliary passage through which a separation buffer can be provided to the first separation passage, and positioned downstream of the first analyte concentrator and positionable upstream of the detection zone;
a second analyte concentrator including second affinity ligands which are attracted to a second analyte of interest, the second analyte concentrator comprises a second concentrator-microreactor;
the second affinity ligands being immobilized to a surface within the second analyte concentrator;
a second separation passage to convey the second analyte of interest from the sample introduced into the transport passage and concentrated by the second analyte concentrator at a second location of the transport passage to the detection zone where the second analyte of interest can be identified and characterized;
the second separation passage being communicable upstream of the second location with a separation or elution buffer supply;
a second auxiliary passage through which a separation buffer can be provided to the second separation passage, and positioned downstream of the second analyte concentrator and positionable upstream of the detection zone;
and
a valve system for controlling the flow in the transport passage and in the first and second separation passages and for controlling fluid flow of separation buffer from the first auxiliary passage to the first separation passage and from the second auxiliary passage to the second separation passage independently and sequentially wherein the first concentrator-microreactor is positioned at an intersection of the transport passage and the first separation passage and has a staggered elongated portion connected to the first separation passage and two shorter portions connected to the transport passage;
and
wherein the second concentrator-microreactor is positioned at an intersection of the transport passage and the second separation passage and has a staggered elongated portion connected to the second separation passage and two shorter portions connected to the transport passage.

12 more elements

An electrophoresis apparatus, comprising:

a transport passage;
a first analyte concentrator including first affinity ligands which are attracted to a first analyte of interest, the first analyte concentrator comprises a first concentrator-microreactor;
the first affinity ligands being immobilized to a surface within the first analyte concentrator;
a first separation passage to convey the first analyte of interest from a sample introduced into the transport passage and concentrated by the first analyte concentrator at a first location of the transport passage to an analyte identification and characterization detection zone where the first analyte of interest can be identified and characterized;
the first separation passage being communicable upstream of the first location with a separation or elution buffer supply;
a first auxiliary passage through which a separation buffer can be provided to the first separation passage, and positioned downstream of the first analyte concentrator and positionable upstream of the detection zone;
a second analyte concentrator including second affinity ligands which are attracted to a second analyte of interest, the second analyte concentrator comprises a second concentrator-microreactor;
the second affinity ligands being immobilized to a surface within the second analyte concentrator;
a second separation passage to convey the second analyte of interest from the sample introduced into the transport passage and concentrated by the second analyte concentrator at a second location of the transport passage to the detection zone where the second analyte of interest can be identified and characterized;
the second separation passage being communicable upstream of the second location with a buffer supply;
a second auxiliary passage through which a separation buffer can be provided to the second separation passage, and positioned downstream of the second analyte concentrator and positionable upstream of the detection zone;
and
a valve system for controlling the flow in the transport passage and in the first and second separation passages and for controlling fluid flow of separation buffer from the first auxiliary passage to the first separation passage and from the second auxiliary passage to the second separation passage independently and sequentially wherein the first concentrator-microreactor is positioned at an intersection of the transport passage and the first separation passage and has a cruciform configuration with four symmetrical portions, two of which are connected to the first separation passage and the other two of which are connected to the transport passage;
and
wherein the second concentrator-microreactor is positioned at an intersection of the transport passage and the second separation passage and has a cruciform configuration with four symmetrical portions, two of which are connected to the second separation passage and the other two of which are connected to the transport passage.

12 more elements

An electrophoresis apparatus, comprising:

a transport passage;
a first analyte concentrator including one or more first affinity ligands which are attracted to one or more first analytes of interest;
the one or more first affinity ligands being immobilized to a surface within the first analyte concentrator;
a first separation passage to convey the one or more first analytes of interest from a sample introduced into the transport passage and concentrated by the first analyte concentrator at a first location of the transport passage to an analyte identification and characterization detection zone where the one or more first analytes of interest can be identified and characterized;
the first separation passage being communicable upstream of the first location with one or more buffer supplies;
a first auxiliary passage through which a separation buffer can be provided to the first separation passage, and positioned downstream of the first analyte concentrator and positionable upstream of the detection zone;
a second analyte concentrator including one or more second affinity ligands which are attracted to one or more second analytes of interest;
the one or more second affinity ligands being immobilized to a surface within the second analyte concentrator;
a second separation passage to convey the one or more second analytes of interest from the sample introduced into the transport passage and concentrated by the second analyte concentrator at a second location of the transport passage to the detection zone where the one or more second analytes of interest can be identified and characterized;
the second separation passage being communicable upstream of the second location with one or more buffer supplies;
a second auxiliary passage through which a separation buffer can be provided to the second separation passage, and positioned downstream of the second analyte concentrator and positionable upstream of the detection zone;
a valve system for controlling the flow in the transport passage and in the first and second separation-passages and for respectively controlling fluid flow of the separation buffer from the first auxiliary passage to the first separation passage and from the second auxiliary passage to the second separation passage independently and sequentially;
and
an auxiliary analyte concentrator on the first separation passage and downstream of the first analyte concentrator, the auxiliary analyte concentrator having one or more affinity ligands capable of retaining chromophores to bind the one or more first analytes of interest released from the first analyte concentrator to improve the sensitivity and selectivity of the one or more first analytes of interest once the one or more first analytes of interest are released from the auxiliary analyte concentrator into the first separation passage and towards the detection zone wherein the auxiliary analyte concentrator is upstream of the first auxiliary passage.

11 more elements

An electrophoresis apparatus, comprising:

a transport passage;
a first analyte concentrator including one or more first affinity ligands which are attracted to one or more first analytes of interest, the first analyte concentrator comprises a first concentrator-microreactor;
the one or more first affinity ligands being immobilized to a surface within the first analyte concentrator;
a first separation passage to convey the one or more first analytes of interest from a sample introduced into the transport passage and concentrated by the first analyte concentrator at a first location of the transport passage to an analyte identification and characterization detection zone where the one or more first analytes of interest can be identified and characterized;
the first separation passage being communicable upstream of the first location with one or more buffer supplies;
a first auxiliary passage through which a separation buffer can be provided to the first separation passage, and positioned downstream of the first analyte concentrator and positionable upstream of the detection zone;
a second analyte concentrator including one or more second affinity ligands which are attracted to one or more second analytes of interest, the second analyte concentrator comprises a second concentrator-microreactor;
the one or more second affinity ligands being immobilized to a surface within the second analyte concentrator;
a second separation passage to convey the one or more second analytes of interest from the sample introduced into the transport passage and concentrated by the second analyte concentrator at a second location of the transport passage to the detection zone where the one or more second analytes of interest can be identified and characterized;
the second separation passage being communicable upstream of the second location with one or more buffer supplies;
a second auxiliary passage through which a separation buffer can be provided to the second separation passage, and positioned downstream of the second analyte concentrator and positionable upstream of the detection zone;
and
a valve system for controlling the flow in the transport passage and in the first and second separation passages and for controlling fluid flow of the separation buffer from the first auxiliary passage to the first separation passage and from the second auxiliary passage to the second separation passage wherein the first concentrator-microreactor is positioned at an intersection of the transport passage and the first separation passage and has a staggered elongated portion connected to the first separation passage and two shorter portions connected to the transport passage;
and
wherein the second concentrator-microreactor is positioned at an intersection of the transport passage and the second separation passage and has a staggered elongated portion connected to the second separation passage and two shorter portions connected to the transport passage.

12 more elements

An electrophoresis apparatus, comprising:

a transport passage;
a first analyte concentrator including one or more first affinity ligands which are attracted to one or more first analytes of interest, the first analyte concentrator comprises a first concentrator-microreactor;
the one or more first affinity ligands being immobilized to a surface within the first analyte concentrator;
a first separation passage to convey the one or more first analytes of interest from a sample introduced into the transport passage and concentrated by the first analyte concentrator at a first location of the transport passage to an analyte identification and characterization detection zone where the one or more first analytes of interest can be identified and characterized;
the first separation passage being communicable upstream of the first location with one or more buffer supplies;
a first auxiliary passage through which a separation buffer can be provided to the first separation passage, and positioned downstream of the first analyte concentrator and positionable upstream of the detection zone;
a second analyte concentrator including one or more second affinity ligands which are attracted to one or more second analytes of interest, the second analyte concentrator comprises a second concentrator-microreactor;
the one or more second affinity ligands being immobilized to a surface within the second analyte concentrator;
a second separation passage to convey the one or more second analytes of interest from the sample introduced into the transport passage and concentrated by the second analyte concentrator at a second location of the transport passage to the detection zone where the one or more second analytes of interest can be identified and characterized;
the second separation passage being communicable upstream of the second location with one or more buffer supplies;
a second auxiliary passage through which a separation buffer can be provided to the second separation passage, and positioned downstream of the second analyte concentrator and positionable upstream of the detection zone;
and
a valve system for controlling the flow in the transport passage and in the first and second separation passages and for controlling fluid flow of the separation buffer from the first auxiliary passage to the first separation passage and from the second auxiliary passage to the second separation passage wherein the first concentrator-microreactor is positioned at an intersection of the transport passage and the first separation passage and has a cruciform configuration with four symmetrical portions, two of which are connected to the first separation passage and the other two of which are connected to the transport passage;
and
wherein the second concentrator-microreactor is positioned at an intersection of the transport passage and the second separation passage and has a cruciform configuration with four symmetrical portions, two of which are connected to the second separation passage and the other two of which are connected to the transport passage.

12 more elements

Dependencies are inferred from the claim text.

Description23,322 words

Summary1,649 words

#1. Field of the Invention

#The present invention relates generally to the analysis of chemical and biological materials and, more particularly, to an improved electrophoresis apparatus which simultaneously performs multiple analyses on a plurality of analytes.

#2. General Background and State of the Art

Brief description of the drawings41 figures

#FIG. 1 is a perspective view of the electrophoresis apparatus of the present invention;

#FIG. 2 is an enlarged, elevated view of a plurality of analyte concentrators stationed at the respective intersections of a large bore transport capillary and an equal plurality of small bore separation capillaries;

#FIG. 3 is an elevated view of a second embodiment of the present invention, showing a plurality of analytes concentrators stationed at the respective intersections of an alternative transport channel and an equal plurality of separation channels;

Detailed description20,759 words

#FIG. 1 illustrates electrophoresis apparatus 10 of the present invention. In its elementary mode (e.g., FIG. 8), apparatus 10 performs single sample studies on chemical or biological matrices having constituents or analytes of interest. But, according to the operating principles shown and described, apparatus 10 can perform multiple analyses by detecting and measuring the presence of a plurality of analytes (for example, three). Suitable and representative analytes may include narcotics, glucose, cholesterol or pharmaceutical drugs that may be present in urine or whole blood, as well as small and large molecular weigh substances having simple and complex structures.

#As shown in FIG. 1, apparatus 10 includes platform 12 having side wall 14. Sample cup 15 is mounted laterally on side wall 14. A large-bore (150-300 mm in length×500-2000 μm I.D.), nonselective introduction capillary 16 and large-volume (1-3 ml) analyte concentrator 17 connect sample cup 15 to a first input of valve 18 which is coupled, by capillary 20, to waste container 22 positioned on side wall 14 adjacent to sample cup 15. In a typical configuration, analyte concentrator 17 comprises a matrix-like assembly of the type shown in U.S. Pat. No. 5,202,010. The collective mass of the matrix is provided by large quantities of microstructures such as beads, platelets, chips, fibers, filament of the like. Individual substrates can be made from glass, plastic or other polymeric material, ceramic, or metallic compositions, and mixtures thereof. Coated or otherwise deposited onto the microstructures are immobilized analyte-specific antibodies or other affinity chemistries which are suitable for characterizing and separating particular analytes of interest. Representative antibodies include those which act against peptide hormones such as insulin, human growth hormone and erythropoietin. These antibodies are readily available from commercial vendors such as Sigma-Aldrich Co., St. Louis, Miss., and Peninsula Laboratories, Belmont, Calif.

#The present invention contemplates a user-friendly, sample preparation step which is designed to eliminate unwanted analytes that occupy binding sites and contaminate the inner walls of capillaries or channels. This procedure will now be described with specific reference to apparatus 10 of FIG. 2.