Electrophoresis apparatus having at least one auxiliary buffer passage

• US 8,007,724 B2
• Filed: 11/17/2006
• Issued: 08/30/2011
• Est. Priority Date: 11/07/2003
• Status: Active Grant

• Current Assignee: Princeton Biochemicals, Inc.
• Original Assignee: Princeton Biochemicals, Inc.
• Inventors: Guzman, Norberto A.
• Primary Examiner(s): Jung; Unsu
• Application Number:
  US11/601,568
• Publication Number:
  US 20070111329A1
• Time in Patent Office: 1,747 Days
• Field of Search: 204/180.1, 204450-455, 204600-605
• US Class Current: 422/82.01
• CPC Class Codes: G01N 1/405 by adsorption or absorption
• CPC Class Codes: G01N 27/44708 Cooling
• CPC Class Codes: G01N 27/44743 Introducing samples
• CPC Class Codes: G01N 27/44756 Apparatus specially adapted therefor
• CPC Class Codes: G01N 33/543 with an insoluble carrier for immobilising immunochemicals
• CPC Class Codes: G01N 33/561 Immunoelectrophoresis

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.

Claims

1. 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.

2. The electrophoresis apparatus of claim 1 wherein the valve system controls the flow of sample and cleaning buffers in a sequential order from an inlet of the transport passage to an outlet of the transport passage, allowing the first and second analytes of interest in the sample to bind to the first and second analyte concentrators, respectively, followed by the flow of a cleaning buffer from the inlet of the transport passage to the outlet of the transport passage to remove excess amount of sample and unwanted material bound non-specifically to the inner wall of the transport passage and the first and second analyte concentrators.

3. The electrophoresis apparatus of claim 1 wherein the first analyte of interest released from the first analyte concentrator is separated in the first separation passage by electrophoresis, electroosmotic flow, mechanical pressure and/or a combination of electroosmotic flow and mechanical pressure, and

the second analyte of interest released from the second analyte concentrator is separated in the second separation passage by electrophoresis, electroosmotic flow, mechanical pressure, and/or a combination of electroosmotic flow and mechanical pressure.

4. The electrophoresis apparatus of claim 1 wherein the buffer supply of the first separation passage includes a first separation buffer supply and a first eluting buffer supply, allowing a plug of first eluting buffer to release the bound first analyte of interest from the first affinity ligands into the first separation passage and allowing the first separation buffer to separate the released first analyte of interest in the first separation passage, and the first separation passage being alternatively in fluid communication with the first separation buffer supply and the first eluting buffer supply;

and

wherein the buffer supply of the second separation passage includes a second separation buffer supply and a second eluting buffer supply, allowing a plug of second eluting buffer to release the bound second analyte of interest from the second affinity ligands into the second separation passage and allowing the second separation buffer to separate the released second analyte of interest in the second separation passage, and the second separation passage being alternatively in fluid communication with the second separation buffer supply and the second eluting buffer supply.

5. The electrophoresis apparatus of claim 1 wherein the first analyte concentrator includes a first matrix assembly that contains the first affinity ligands and wherein the first analyte of interest from the sample binds to the first affinity ligands;

and

wherein the second analyte concentrator includes a second matrix assembly that contains the second affinity ligands and wherein the second analyte of interest from the sample binds to the second affinity ligands.

6. The electrophoresis apparatus of claim 5 wherein the first matrix assembly includes a plurality of microstructures;

and

wherein the second matrix assembly includes a plurality of microstructures.

7. The electrophoresis apparatus of claim 1 wherein the first separation passage is capable of separating the first analyte of interest retained by the first affinity ligands after the first analyte of interest is released from the first affinity ligands and separating the released first analyte of interest by at least one mode of capillary electrophoresis;

and

wherein the second separation passage is capable of separating the second analyte of interest retained by the second affinity ligands after the second analyte of interest is released from the second affinity ligands and separating the released second analyte of interest by at least one mode of capillary electrophoresis.

8. The electrophoresis apparatus of claim 1 wherein the first separation passage is filled with an electrically conductive fluid and separation of the first analyte of interest in the first separation passage is by electrophoresis migration and/or pressure;

and

wherein the second separation passage is filled with an electrically conductive fluid and separation of the second analyte of interest in the second separation passage is by electrophoresis migration and/or pressure.

9. The electrophoresis apparatus of claim 1 wherein the first separation passage is filled with a gel matrix and an electrically conductive fluid and separation of the first analyte of interest in the first separation passage is by electrophoresis migration;

and

wherein the second separation passage is filled with a gel matrix and an electrically conductive fluid and separation of the second analyte of interest in the second separation passage is by electrophoresis migration.

10. The electrophoresis apparatus of claim 1 wherein the first analyte concentrator is a first concentrator-microreactor;

wherein the first affinity ligands are capable of performing at least one chemical or biochemical reaction;

wherein the second analyte concentrator is a second concentrator-microreactor; and

wherein the second affinity ligands are capable of performing at least one chemical or biochemical reaction.

11. The electrophoresis apparatus of claim 1 wherein the first analyte concentrator has an encapsulated cellular or subcellular structure adapted to carry out drug metabolism studies;

and

wherein the second analyte concentrator has an encapsulated cellular or subcellular structure adapted to carry out drug metabolism studies.

12. The electrophoresis apparatus of claim 1 wherein the first affinity ligands are covalently or non-covalently immobilized to a matrix assembly of the first analyte concentrator and the second affinity ligands are covalently or non-covalently immobilized to a matrix assembly of the second analyte concentrator.

13. The electrophoresis apparatus of claim 1 further comprising one or more detection systems positioned at the detection zone.

14. The electrophoresis apparatus of claim 13 wherein the detection system is an ultraviolet, fluorescence, a laser-induced fluorescence, mass spectrometry, circular dichroism, nuclear magnetic resonance, radioactive conductivity, electrochemical detection system.

15. The electrophoresis apparatus of claim 1 further comprising bulging members along corners of said first analyte concentrator wherein passage bulging members retain the first affinity ligands in the first analyte concentrator and

corners of said second analyte concentrator wherein passage bulging members retain the second affinity ligands in the second analyte concentrator.

16. The electrophoresis apparatus of claim 1 wherein an inner diameter of the transport passage is larger than inner diameters of the first and second separation passages.

17. The electrophoresis apparatus of claim 1 wherein the transport passage and the first and second separation passages are all capillaries.

18. The electrophoresis apparatus of claim 1 wherein the capillaries are fused-silica or plastic tubes.

19. The electrophoresis apparatus of claim 1 wherein the first analyte concentrator is connected to an acoustic micromixing system.

20. The electrophoresis apparatus of claim 1 wherein the second analyte concentrator is connected to an acoustic micromixing system.

21. The electrophoresis apparatus of claim 1 wherein one of an anode or cathode sides of the electrophoresis apparatus is generally at the buffer supply of at least one of the first and second separation passages and the other side is downstream of a single outlet passage.

22. The electrophoresis apparatus of claim 21 wherein the other side is downstream of the detection zone.

23. The electrophoresis apparatus of claim 1 wherein an inlet end of the first separation passage is alternatively in fluid communication with a separation buffer supply, an eluting buffer supply or a cleaning solution supply;

and

wherein an inlet end of the second separation passage is alternatively in fluid communication with a separation buffer supply, an eluting buffer supply or a cleaning solution supply.

24. The electrophoresis apparatus of claim 1 wherein the separation buffer introduced into the first separation passage through the first auxiliary passage includes preferentially an organic solvent, or a mixture of organic solvents;

and

wherein the separation buffer introduced into the second separation passage through the second auxiliary passage includes preferentially an organic solvent, or a mixture of organic solvents.

25. The electrophoresis apparatus of claim 1 wherein at least portions of the transport passage and the first separation passage are supported on a platform and the first auxiliary passage is perpendicular to a plane defined by the platform;

and

wherein a sample is introduced into an inlet of the transport passage and exits into a waste container at an outlet of the transport passage.

26. The electrophoresis apparatus of claim 25 wherein at least a portion of the transport passage and the second separation passage is supported on the platform and the second auxiliary passage is perpendicular to the plane defined by the platform.

27. The electrophoresis apparatus of claim 1 wherein an inlet end of the transport passage can be in fluid communication with a cleaning buffer supply for cleaning the first analyte concentrator with the first analyte of interest captured therein and for cleaning the second analyte concentrator with the second analyte of interest captured therein.

28. The electrophoresis apparatus of claim 1 wherein an inlet end of the first separation passage can be in fluid communication with a cleaning buffer supply for cleaning the first separation passage and the first analyte concentrator with the first analyte of interest captured therein;

and

wherein an inlet end of the second separation passage can be in fluid communication with a cleaning buffer supply for cleaning the second separation passage and the second analyte concentrator with the second analyte of interest captured therein.

29. The electrophoresis apparatus of claim 1 wherein the buffer supply of the first separation passage includes an eluting buffer supply and a different separation buffer supply, and the first separation passage can be alternatively sequentially in fluid communication with the separation buffer supply and the eluting buffer supply.

30. The electrophoresis apparatus of claim 29 wherein the buffer supply of the second separation passage includes an eluting buffer supply and a different separation buffer supply, and the second separation passage can be alternatively sequentially in fluid communication with the separation buffer supply and the eluting buffer supply.

31. The electrophoresis apparatus of claim 1 wherein one of the first and second analytes of interest is a simple molecule of interest and the other of the first and second analytes of interest is a complex molecule of interest.

32. The electrophoresis apparatus of claim 1 wherein a detector at the detection zone identifies and characterizes molecules delivered thereto via at least one of the first and second separation passages.

33. The electrophoresis apparatus of claim 1 wherein the first separation passage from an inlet of the first separation passage to an outlet of the first separation passage has electrically conductive fluid therein.

34. The electrophoresis apparatus of claim 33 wherein the electrically conductive fluid has buffer conditions that facilitate binding of the first affinity ligands with the first analyte of interest and that facilitate separation within the first separation passage of the released first analyte of interest from the first affinity ligands.

35. The electrophoresis apparatus of claim 34 wherein the second separation passage has therein from an inlet of the second separation passage to an outlet of the second separation passage electrically conductive fluid that has buffer conditions that facilitate binding of the second affinity ligands with the second analyte of interest and that facilitate separation within the second separation passage of the released second analyte of interest from the second affinity ligands.

36. The electrophoresis apparatus of claim 33 wherein the first separation passage has electrically conductive fluid and a gel matrix therein, the electrically conductive fluid in the first separation passage has buffer conditions, from an inlet of the first separation passage to a junction of the first separation passage with the first auxiliary passage, that facilitate binding of the first affinity ligands with the first analyte of interest;

and

wherein the gel matrix and the electrically conductive fluid introduced through the first auxiliary passage into the first separation passage towards the detection zone have conditions that facilitate separation in the first separation passage of the released first analyte of interest from the first affinity ligands which are immobilized to the surface of the first analyte concentrator.

37. The electrophoresis apparatus of claim 36 wherein the second separation passage has electrically conductive fluid and a gel matrix therein, the electrically conductive fluid in the second separation passage has buffer conditions, from the inlet of the second separation passage to a junction of the second separation passage with the second auxiliary passage, that facilitate binding of the second affinity ligands with the second analyte of interest;

and

wherein the gel matrix and the electrically conductive fluid introduced through the second auxiliary passage into the second separation passage towards the detection zone have conditions that facilitate separation in the second separation passage of the released second analyte of interest from the second affinity ligands which are immobilized to the surface of the second analyte concentrator.

38. The electrophoresis apparatus of claim 1 wherein an inlet end of the transport passage can be in fluid communication with a cleaning buffer supply for cleaning the transport passage and the first analyte concentrator with the first analyte of interest captured therein and the second analyte concentrator with the second analyte of interest captured therein.

39. The electrophoresis apparatus of claim 1 wherein the valve system includes a first valve at a junction of the first separation passage and the first auxiliary passage, and positioned downstream of the first concentrator-microreactor and upstream of the detection zone;

and

a second valve at a junction of the second separation passage and the second auxiliary passage, and positioned downstream of the second concentrator-microreactor and upstream of the detection zone.

40. The electrophoresis apparatus of claim 1 further comprising:

a first frit structure in an inlet of the transport passage into the first concentrator-microreactor;

a second frit structure in an outlet of the transport passage from the first concentrator-microreactor;

a third frit structure in an inlet of the first separation passage into the first concentrator-microreactor; and

a fourth frit structure in an outlet of the first separation passage from the first concentrator-microreactor;

wherein the first, second, third and fourth frit structures are positioned at connection terminals of the first concentrator-microreactor with the transport passage and the first separation passage;

wherein the control means includes a valve system which includes valves on the transport passage on both sides of the first concentrator-microreactor and valves on the first separation passage on both sides of the first concentrator-microreactor;

further comprising;

a fifth frit structure in an inlet of the transport passage into the second concentrator-microreactor;

a sixth frit structure in an outlet of the transport passage from the second concentrator-microreactor;

a seventh frit structure in an inlet of the second separation passage into the second concentrator-microreactor; and

an eighth frit structure in an outlet of the second separation passage from the second concentrator-microreactor;

wherein the fifth, sixth, seventh and eighth frit structures are positioned at connection terminals of the second concentrator-microreactor with the transport passage and the second separation passage; and

wherein the valve system includes valves on the transport passage on both sides of the second concentrator-microreactor and valves on the second separation passage on both sides of the second concentrator-microreactor.

41. The electrophoresis apparatus of claim 1 wherein the first analyte concentrator comprises a first concentrator-microreactor which includes a matrix assembly that contains the first affinity ligands;

wherein organic solvents, urea, certain detergents or other additives are preferentially incorporated in a separation buffer solution to form a complex buffer solution and to facilitate the separation of first released and concentrated analyte of interest; and

wherein the first auxiliary passage introduces the complex buffer solution into the first separation passage, towards the detection zone and away from the first concentrator-microreactor to preserve the integrity of the first affinity ligands that are immobilized to a surface of the first analyte concentrator.

42. The electrophoresis apparatus of claim 1 wherein the valve system controls fluid flow in the first and second separation passages independently and sequentially.

43. The electrophoresis apparatus of claim 1 wherein the first separation passage from an inlet of the first separation passage to a junction with the first auxiliary passage has electrically conductive fluid therein that is a facilitator with binding of molecules to facilitate binding of the of the first affinity ligands with the first analyte of interest;

and

wherein the first separation passage from the junction to an outlet of the first separation passage has electrically conductive fluid therein that is a facilitator with separation of molecules to facilitate separation of the first analyte of interest in the first separation passage.

44. The electrophoresis apparatus of claim 1 wherein the second separation passage from an inlet of the second separation passage to a junction with the second auxiliary passage has electrically conductive fluid therein that is a facilitator with binding of molecules to facilitate binding of the second affinity ligands with the second analyte of interest;

and

wherein the second separation passage from the junction to the outlet of the second separation passage has electrically conductive fluid therein that is a facilitator with separation of molecules to facilitate separation of the second analyte of interest in the second separation passage.

45. The electrophoresis apparatus of claim 1 wherein the first analyte concentrator and the second analyte concentrator is used to bind cells, organelles or bulky globules from the sample passed through the transport passage directly to the respective first affinity ligands being immobilized to a surface within the first analyte concentrator and the second affinity ligands being immobilized to a surface within the second analyte concentrator;

and

wherein the first and second separation passages merge into a single exit output capillary or channel.

46. 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.

47. The electrophoresis apparatus of claim 46 further comprising an auxiliary analyte concentrator on the first separation passage and downstream of the first analyte concentrator, the 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 auxiliary analyte concentrator into the first separation passage and towards the detection zone;

and an auxiliary analyte concentrator on the second separation passage and downstream of the second analyte concentrator, the 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 auxiliary analyte concentrator into the second separation passage and towards the detection zone.

48. The electrophoresis apparatus of claim 46 wherein the first analyte concentrator comprises a first concentrator-microreactor.

49. The electrophoresis apparatus of claim 48 wherein the second analyte concentrator comprises a second concentrator-microreactor.

50. 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.

51. The electrophoresis apparatus of claim 50 further comprising an auxiliary analyte concentrator downstream from one of the first and second analyte concentrators on one of the first and second separation passages, the auxiliary analyte concentrator having affinity ligands capable of retaining chromophores to bind to the first or second analytes of interest released from the one of the first and second analyte concentrators to improve the sensitivity and selectivity of the first or second analytes of interest.

52. 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.

53. 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.

54. 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.