System and method for the separation of analytes
View Patent ↗A separation module operates to fractionate or separate an analyte into fractions according to pI, i.e., pI bands, utilizing capillary isoelectric focusing (“CIEF”) within a first microchannel. The fractions are stacked to form plugs, the number of which is determined by a number of parallel second microchannels integrally connected to the first microchannel, into which the fractions are directed according to the buffer characteristics found in each of the individual microchannels. Within the microchannels the plugs are separated into proteins according to a different chemical property, i.e., “m/z,” utilizing capillary electrophoresis (“CE”).
1. A process for separating a sample comprising:
introducing a sample into a first microchannel formed in a module;
separating the sample into a plurality of sub-samples based on varying isoelectric points of the plurality of sub-samples;
causing with a first buffer a first of the plurality of sub-samples to enter a first of a plurality of second parallel microchannels integrally connected to the first microchannel, said first of a plurality of second parallel microchannels containing a second buffer having an ionic strength different from said first buffer for base stacking the first plurality of sub-samples between said first and second buffers; and
separating the first of the plurality of sub-samples into a plurality of protein components according to electrophoresis.
2. The process according to claim 1 , further comprising applying opposite charges to either end of the first microchannel to facilitate separating the sample into a plurality of sub-samples; and
applying a first charge to a non-intersecting end of a third microchannel integrally connected to the first microchannel and applying a second, opposite charge to the non-intersecting end of the first of the plurality second microchannels to facilitate the first of the plurality of sub-samples entering the first of the plurality of second microchannels and further separating according to electrophoresis.
3. The process according to claim 1 , further comprising:
aligning a first detector with the first microchannel in order to detect the separation of the sample into the plurality of sub-samples based on varying isoelectric points of the plurality of sub-samples within the first microchannel.
4. The process according to claim 3 , further comprising:
aligning a second detector with the non-intersecting end of the first of the plurality of second microchannels; and
detecting the plurality of protein components for the first of the plurality of sub-samples.
5. The process according to claim 4 , further comprising:
creating first spectral data indicative of the plurality of protein components of the first of the plurality of sub-samples.
6. The process according to claim 5 , further comprising:
aligning the first detector with the first of the plurality of second microchannels;
detecting the separation of first of the plurality of sub-samples into the plurality of protein components; and
creating second spectral data indicative of the protein components of the first of the plurality of sub-samples.
7. The process according to claim 6 , farther comprising:
reducing the first spectral data and the second spectral data to facilitate identification of each of the plurality of protein components.
8. The process according to claim 3 , further comprising:
subsequently applying a first charge to a non-intersecting end of a third microchannel and applying a second, opposite charge to a non-intersecting end of a second of the plurality of second microchannels to facilitate a second of the plurality of sub-samples entering the second of the plurality of second microchannels and further separating according to electrophoresis;
aligning a second detector with a non-intersecting end of the second of the plurality of second microchannels; and
detecting the plurality of protein components for the second of the plurality of sub-samples.