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Journal of chromatography. A

Incorporation of polyamidoamine sweeping and electrokinetic supercharging for in-line DNA fragment preconcentration.


PMID 23374369

Abstract

We report an approach for in-line preconcentration of DNA fragments using dendritic polyamidoamine generation 2.0 (PAMAM G 2.0) as sweeping agent. During the experiment, a plug of PAMAM G 2.0 was hydrodynamically injected first, followed by field-amplified sample injection (FASI) of DNA fragments, which were concurrently swept by PAMAMs via DNA-PAMAM complexation. Then, a segment of releasing agent, sodium dodecyl sulfate (SDS), was hydrodynamically introduced and subsequently electrophoretically driven to interact with the DNA-PAMAM complexes, forming more stable supramolecular SDS-PAMAM complexes and releasing the initially bound DNA fragments. The excess dodecyl sulfate anion also acted as terminating electrolyte in the separation, thereby the DNA fragments were enriched by the joint effects of FASI, sweeping and transient isotachophoresis. We term the approach PAMAM sweeping-electrokinetic supercharging (EKS). Because the mobility of the DNA-PAMAM complex was very low, the proposed method allowed long-time sample injection without notable loss in separation efficiency. Under the optimum conditions, the PAMAM sweeping-EKS strategy improved the detection sensitivity of DNA fragments by more than 30 folds relative to the conventional FASI. Moreover, due to the sweeping process incorporated, the approach can be applied to enrichment of DNA fragments in highly saline matrix.