Analyte injection system

Abstract

This invention provides methods and systems for injection of analytes into a separation channel for resolution and detection. Samples can be preconditioned and concentrated by isotachophoresis (ITP) before the injection is triggered by a detected voltage event. Separation of analytes from other sample constituents can be enhanced using skewing channel ITP.

1 . A method of applying a stacked analyte to a channel segment comprising: 
 stacking one or more analytes in a channel;    detecting a voltage potential in the channel; and,    applying an electric field or a pressure differential along a channel segment when a selected voltage event is detected;    thereby applying the stacked analytes to the channel segment.    
     
     
         2 . The method of  claim 1 , wherein stacking comprises transient stacking or steady state stacking.  
     
     
         3 . The method of  claim 1 , wherein the channel comprises a microscale channel.  
     
     
         4 . The method of  claim 1 , wherein the channel comprises a stacking channel segment or a separation channel segment.  
     
     
         5 . The method of  claim 4 , wherein the stacking channel segment comprises a trailing electrolyte or a leading electrolyte.  
     
     
         6 . The method of  claim 4 , wherein the stacking channel segment comprises a trailing electrolyte and a leading electrolyte, which electrolytes comprise different mobilities.  
     
     
         7 . The method of  claim 6 , wherein the trailing electrolyte and the leading electrolyte differ in one or more of: a pH, a viscosity, a conductivity, size exclusion, an ionic strength, an ion composition, or a temperature.  
     
     
         8 . The method of  claim 6 , further comprising: adjusting the trailing electrolyte to comprise a mobility less than the one or more analytes, or adjusting the leading electrolyte to comprise a mobility greater than the one or more analytes.  
     
     
         9 . The method of  claim 6 , further comprising: adjusting the trailing electrolyte to comprise a mobility greater than one or more sample constituents not of interest, or adjusting the leading electrolyte to comprise a mobility less than one or more sample constituents not of interest.  
     
     
         10 . The method of  claim 1 , wherein the channel comprises a skewing channel segment.  
     
     
         11 . The method of  claim 10 , wherein the skewing channel segment comprises: a serpentine curve, a helix, a coil, an angle, or a spiral.  
     
     
         12 . The method of  claim 10 , wherein the skewing channel segment comprises conditions providing a dispersion Peclet number more than  0 . 1  times a ratio of a skewing channel length over a skewing channel width.  
     
     
         13 . The method of  claim 10 , wherein the channel comprises a skewing channel internal width greater than a skewing channel depth.  
     
     
         14 . The method of  claim 10 , wherein the skewing channel segment comprises a travel surface distance on a first side of the skewing channel greater than a travel surface distance on a second side of the skewing channel.  
     
     
         15 . The method of  claim 10 , wherein the stacking comprises selective isotachophoresis.  
     
     
         16 . The method of  claim 4 , wherein said applying an electric field comprises switching from a substantial lack of current in the separation channel segment and a current in the stacking channel segment to a current in the separation channel segment and a substantial lack of current in the stacking channel segment.  
     
     
         17 . The method of  claim 16 , wherein the substantial lack of current comprises a float voltage or a lack of a complete circuit.  
     
     
         18 . The method of  claim 4 , wherein the separation channel segment comprises one or more of: a pH gradient, size selective media, ion exchange media, a hydrophobic media, or a viscosity enhancing media.  
     
     
         19 . The method of  claim 4 , further comprising detecting analytes in the separation channel segment or detecting analytes eluting from the separation channel segment.  
     
     
         20 . The method of  claim 19 , wherein said detecting analytes comprises monitoring: a conductivity, a fluorescence, a light absorbance, or a refractive index.  
     
     
         21 . The method of  claim 1 , wherein said stacking comprises consecutively stacking two or more samples of the analytes in the channel.  
     
     
         22 . The method of  claim 21 , wherein said stacking the two or more samples comprises: 
 loading a first sample into a loading channel segment;    applying an electric field across the sample, thereby stacking sample analytes;    loading a second sample into the loading channel segment; and,    applying an electric field across the stacked sample analytes and the second sample.    
     
     
         23 . The method of  claim 22 , further comprising flowing the stacked first sample analytes towards the loading channel segment.  
     
     
         24 . The method of  claim 1 , wherein said stacking comprises loading samples of the analytes in a loading channel segment comprising a cross-section greater than a stacking channel segment cross-section.  
     
     
         25 . The method of  claim 1 , wherein said stacking comprises loading one or more spacer electrolytes between two or more sample analytes, which spacer electrolytes comprise a mobility greater than a trailing electrolyte and a mobility less than a leading electrolyte.  
     
     
         26 . The method of  claim 25 , wherein one or more of the two or more sample analytes comprise a stacked sample analyte.  
     
     
         27 . The method of  claim 25 , further comprising adjusting the spacer electrolytes to provide a mobility between mobilities of two or more of the analytes.  
     
     
         28 . The method of  claim 25 , further comprising adjusting the mobility of the spacer electrolytes by selecting one or more of: a spacer electrolyte pH, a spacer electrolyte viscosity, or a spacer electrolyte conductivity.  
     
     
         29 . The method of  claim 1 , further comprising determining a pK of the analytes.  
     
     
         30 . The method of  claim 29 , further comprising adjusting the pH of a trailing electrolyte or a leading electrolyte to be higher or lower than the determined pK.  
     
     
         31 . The method of  claim 1 , wherein said detecting comprises monitoring a float voltage.  
     
     
         32 . The method of  claim 1 , wherein the voltage event comprises: a voltage peak, a voltage trough, a predesignated voltage, a relative voltage, or a rate of voltage change.  
     
     
         33 . The method of  claim 1 , wherein said applying an electric field or pressure differential along the channel segment is automatic when the voltage event is detected.  
     
     
         34 . The method of  claim 1 , wherein the analytes comprise one or more of: a protein, a nucleic acid, a carbohydrate, a glycoprotein, a derivitized molecule, or an ion.  
     
     
         35 . A system for application of a stacked analyte to a channel segment comprising: 
 a channel;    an analyte stacking in the channel; and,    a voltage detector in electrical contact with the channel, and in communication with a controller;    wherein the controller initiates a flow of electric current in a channel segment or a pressure differential along the channel segment when a selected voltage event is detected by the voltage detector.    
     
     
         36 . The system of  claim 35 , wherein the analyte comprises one or more of: a protein, a nucleic acid, a carbohydrate, a glycoprotein, a derivatized molecule, or an ion.  
     
     
         37 . The system of  claim 35 , wherein the channel comprises a microscale channel.  
     
     
         38 . The system of  claim 35 , wherein the channel comprises a stacking channel segment or a separation channel segment.  
     
     
         39 . The system of  claim 38 , wherein the stacking channel segment comprises a trailing electrolyte or a leading electrolyte, which electrolytes comprise different mobilities.  
     
     
         40 . The system of  claim 39 , wherein the trailing electrolyte and the leading electrolyte differ in one or more of: a pH, a viscosity, a conductivity, a size exclusion, an ionic strength, an ion composition, or a temperature.  
     
     
         41 . The system of  claim 39 , wherein the trailing electrolyte comprises a mobility less than a mobility of the analyte of interest or a mobility greater than a mobility of a sample constituent not of interest.  
     
     
         42 . The system of  claim 39 , wherein the leading electrolyte comprises a mobility greater than a mobility of the analyte of interest or a mobility less than a mobility of a sample constituent not of interest.  
     
     
         43 . The system of  claim 38 , wherein the separation channel segment comprises one or more of: a pH gradient, size selective media, ion exchange media, a hydrophobic media, or a viscosity enhancing media.  
     
     
         44 . The system of  claim 35 , wherein the controller comprises a logic device or a system operator.  
     
     
         45 . The system of  claim 38 , further comprising substantial elimination of current in the stacking channel segment when the voltage event is detected.  
     
     
         46 . The system of  claim 38 , wherein the channel further comprises a loading channel segment in fluid contact with the stacking channel segment.  
     
     
         47 . The system of  claim 46 , wherein the loading channel segment comprises a cross-section greater than a stacking channel segment cross-section.  
     
     
         48 . The system of  claim 46 , further comprising a pressure differential across the stacking channel segment, whereby a stacked sample can be flowed toward the loading channel segment.  
     
     
         49 . The system of  claim 46 , further comprising a collector tube through which an analyte sample can flow into the loading channel segment.  
     
     
         50 . The system of  claim 38 , further comprising a spacer electrolyte between two or more analyte sample segments in the stacking channel segment.  
     
     
         51 . The system of  claim 50 , wherein the spacer electrolyte comprises a mobility between mobilities of two or more of the analytes in the sample segments.  
     
     
         52 . The system of  claim 35 , further comprising a float voltage regulator or a switch in electrical contact with the channel.  
     
     
         53 . The system of  claim 35 , wherein the voltage event comprises one or more of: a voltage peak, a selected voltage, a voltage trough, a relative voltage, or a rate of voltage change.  
     
     
         54 . The system of  claim 35 , wherein the flowing of electric current in the channel segment or application of the pressure differential along the channel segment is automatic on detection of the voltage event.  
     
     
         55 . The system of  claim 38 , further comprising an analyte detector directed to monitor: analytes in the separation channel segment, or analytes eluted from the separation channel segment.  
     
     
         56 . The system of  claim 55 , wherein the analyte detector comprises: a fluorometer, a spectrophotometer, a refractometer, or a conductivity meter.  
     
     
         57 . The system of  claim 35 , further comprising a microfluidic chip.  
     
     
         58 . The system of  claim 38 , wherein the channel comprises a skewing channel segment.  
     
     
         59 . The system of  claim 58 , wherein the skewing channel segment comprises: a serpentine curve, a helix, an angle, or a spiral.  
     
     
         60 . The system of  claim 58 , further comprising skewing channel segment conditions that provide a dispersion Peclet number more than about 0.1 times a ratio of a skewing channel length over a skewing channel width.  
     
     
         61 . The system of  claim 58 , wherein the skewing channel segment comprises a skewing channel internal width greater than a skewing channel depth.  
     
     
         62 . The system of  claim 58 , wherein the skewing channel segment comprises a travel surface distance on a first side of the skewing channel greater than a travel surface distance on a second side of the skewing channel.  
     
     
         63 . The system of  claim 58 , wherein the stacking comprises selective isotachophoresis.  
     
     
         64 . A method of separating an analyte of interest from a sample constituent not of interest, the method comprising: 
 stacking the analyte by isotachophoresis in a channel comprising a skewing channel segment; and,    flowing the analyte and sample constituent not of interest through the skewing channel segment during or before the isotachophoresis,    wherein the skewing channel segment comprises conditions providing a dispersion Peclet number more than 0.1 times a ratio of a skewing channel length over a skewing channel width.    
     
     
         65 . The method of  claim 64 , wherein the analytes comprise one or more of: a protein, a nucleic acid, a carbohydrate, a glycoprotein, a derivitized molecule, or an ion.  
     
     
         66 . The method of  claim 64 , wherein the isotachophoresis comprises selective isotachophoresis.  
     
     
         67 . The method of  claim 64 , wherein the skewing channel comprises: a serpentine curve, a helix, an angle, a coil, or a spiral.  
     
     
         68 . The method of  claim 64 , wherein the skewing channel segment comprises conditions providing a dispersion Peclet number more than the ratio of the skewing channel length over the skewing channel width.  
     
     
         69 . The method of  claim 64 , wherein the channel has a greater internal width at the skewing channel segment.  
     
     
         70 . The method of  claim 64 , wherein the skewing channel segment comprises a skewing channel internal width greater than a skewing channel depth.  
     
     
         71 . The method of  claim 64 , wherein the skewing channel segment comprises a travel surface distance on a first side of the skewing channel greater than a travel surface distance on a second side of the skewing channel.  
     
     
         72 . The method of  claim 71 , wherein the difference between the travel surface distances of the first side and the second side is at least about 25%.  
     
     
         73 . The method of  claim 64 , further comprising: 
 detecting a voltage potential in the channel; and,    applying an electric field or a pressure differential along a channel segment when a selected voltage event is detected;    thereby applying the stacked analytes to the channel segment.    
     
     
         74 . The method of  claim 73 , wherein the channel segment comprises a separation channel.  
     
     
         75 . A isotachophoresis system comprising: 
 a channel comprising a skewing channel segment; and,    an analyte in the channel stacking by isotachophoresis;    wherein the skewing channel segment comprises conditions that provide a dispersion Peclet number more than  0 . 1  times a ratio of a skewing channel length over a skewing channel width.    
     
     
         76 . The isotachophoresis system of  claim 75 , wherein the channel comprises a micro scale channel.  
     
     
         77 . The isotachophoresis system of  claim 75 , wherein the skewing channel comprises segment: a serpentine curve, a helix, an angle, or a spiral.  
     
     
         78 . The isotachophoresis system of  claim 75 , wherein the channel has a greater internal width in the skewing channel segment.  
     
     
         79 . The isotachophoresis system of  claim 75 , wherein the skewing channel comprises segment a skewing channel internal width greater than a skewing channel depth.  
     
     
         80 . The isotachophoresis system of  claim 75 , wherein the skewing channel segment comprises a travel surface distance on a first side of the skewing channel greater than a travel surface distance on a second side of the skewing channel.  
     
     
         81 . The isotachophoresis system of  claim 75 , wherein the dispersion Peclet number is more than about the ratio of the skewing channel length over the skewing channel width.  
     
     
         82 . The isotachophoresis system of  claim 75 , wherein the analyte comprises one or more of: a protein, a nucleic acid, a carbohydrate, a glycoprotein, a derivitized molecule, or an ion.  
     
     
         83 . The isotachophoresis system of  claim 75 , wherein the isotachophoresis comprises selective isotachophoresis.  
     
     
         84 . The isotachophoresis system of  claim 75 , wherein the isotachophoresis comprises a leading electrolyte comprising a mobility greater than a mobility of the analyte or a trailing electrolyte comprising a mobility less than the mobility of analyte.  
     
     
         85 . The isotachophoresis system of  claim 75 , wherein the selective isotachophoresis comprises a leading electrolyte comprising a mobility less than a mobility of sample constituent not of interest or a trailing electrolyte comprising a mobility greater than the mobility of the sample constituent not of interest.  
     
     
         86 . The isotachophoresis system of  claim 75 , further comprising: 
 a voltage detector in electrical contact with the channel, and in communication with a controller;    wherein the controller initiates a flow of electric current in a channel segment or a pressure differential along the channel segment when a selected voltage event is detected by the voltage detector.