Enhanced Isotachophoresis Assays Using Additives with Spatial Gradients
Techniques for enhanced isotachophoresis assays using additives with spatial gradients include forming a concentration gradient of an additive along a channel from an input port to an output port. The channel is used for isotachophoresis with ions of a leading electrolyte having a first mobility greater than a mobility of an analyte, and ions of a trailing electrolyte having a second mobility less than the mobility of the analyte. The additive is different from both the leading electrolyte and the trailing electrolyte; and the additive has a third mobility that assures the analyte will encounter the additive. The method further comprises introducing a mixture of the trailing electrolyte and a sample including the analyte. The method further comprises applying an electric field to the channel; and, measuring the analyte.
1 . A method comprising:
forming a concentration gradient of each of one or more additives along a channel from an input port to an output port, wherein:
the channel is used for isotachophoresis with ions of a leading electrolyte having a first effective mobility magnitude greater than an effective mobility of an analyte, and ions of a trailing electrolyte having a second effective mobility magnitude less than the effective mobility of the analyte;
each additive is different from both the leading electrolyte and the trailing electrolyte; and
each additive has a third mobility that assures the analyte will encounter the additive;
contacting a sample including the analyte to the leading electrolyte;
contacting the trailing electrolyte to the sample;
applying an electric field to the channel; and
measuring the analyte.
2 . A method as recited in claim 1 , wherein each additive operates most effectively on a component of the sample along a portion of the channel where the concentration of the additive is in a particular range of concentrations.
3 . A method as recited in claim 1 , wherein the channel is a microchannel.
4 . A method as recited in claim 1 , wherein the one or more additives comprise a polymer that provides a sieving matrix.
5 . A method as recited in claim 1 , wherein the analyte is a nucleic acid comprising a plurality of nucleotides and an additive comprises a denaturing agent.
6 . A method as recited in claim 1 , wherein the one or more additives comprise a fluorescent label.
7 . A method as recited in claim 1 , wherein the analyte is a nucleic acid comprising a plurality of nucleotides and the one or more additives comprise a nucleic acid probe with a fluorescent label.
8 . A method as recited in claim 1 , wherein the analyte is a nucleic acid comprising a plurality of nucleotides and the one or more additives comprise a molecular beacon that comprises a nucleic acid probe with a fluorescent label at one end and a fluorescence quencher at a different end.
9 . A method as recited in claim 4 , wherein forming the concentration gradient of each of one or more additives along the channel from the input port to the output port further comprises:
disposing a photo-initiator and monomer within the channel;
covering the channel with a photo-mask that varies along the length of the channel; and
exposing the photo-mask to polymerizing light that causes the monomer to form the polymer that provides the sieving matrix.
10 . A method as recited in claim 1 , wherein contacting the trailing electrolyte to the sample further comprises contacting the trailing electrolyte to the sample and a solution of spacer ions, wherein each spacer ion has a third effective mobility magnitude between the effective mobility magnitude of a product of the analyte and the effective mobility magnitude the analyte.
11 . A method as recited in claim 10 , wherein the product of the analyte is a reporter molecule.
12 . An apparatus comprising:
a channel connecting an input port to an output port;
a concentration gradient of each of one or more additives along the channel:
means for contacting a sample including an analyte to a leading electrolyte, wherein the leading electrolyte has a first effective mobility magnitude greater than an effective mobility of the analyte and the leading electrolyte is different from the additive;
means for contacting a trailing electrolyte to the sample, wherein the trailing electrolyte has a second effective mobility magnitude less than the effective mobility of the analyte and the trailing electrolyte is different from the additive;
means for applying an electric field to the channel; and
means for measuring the analyte.