Apparatus and method for electrical detection of oligonucleotides through pore blockades

摘要

Systems and methods for specific nucleic acid (NA) sequence detection that do not rely on polymerase chain reaction (PCR) for target sequence amplification and do not require any special reagents other than a complementary sequence capture probe conjugated to spherical beads.

主权项

1. A method for detecting the presence of nucleic acids with a target sequence in a sample, the method comprising:
(a) providing a detection apparatus, said detection apparatus comprising:
(i) a first chamber;(ii) a second chamber;(iii) a membrane separating said first and second chambers, said membrane having one or more pores extending through the membrane between the first and second chambers;(iv) a first electrode disposed in the first chamber; and(v) a second electrode disposed in the second chamber; (b) introducing a liquid into said first and second chambers; (c) applying a direct electric current to said first and second electrodes with a negative polarity being connected to the first electrode; (d) measuring electric current between the electrodes as a reference current; (e) preparing at least one bead which is conjugated with one or more strands of capture probe nucleic acid that are complementary to a desired target sequence of a sample to be detected; (f) capping charged groups on the prepared beads to render the beads with a neutral charge; (g) introducing one or more of the capped beads and a sample into the first chamber to hybridize the capture probe to the desired target sequence; and (h) detecting the target sequence by measuring electric current between the electrodes after introduction of the beads and sample and comparing the measured current with the reference current; (i) wherein a decrease in measured electric current indicates the presence of one or more strands of target nucleic acid that are complementary to the probe sequence conjugated to the bead; and (j) wherein said decrease in measured electric current results from electrophoretic movement of the beads bound to charged target nucleic acid strands to a pore of diameter less than the bead diameter or tapered to a lesser diameter and the beads causing sustained blockage of one or more pores.