May 6, 2004
Gold nanoparticles and bio-bar codes bring sensitive DNA detection
Researchers at Northwestern University, US, have shown that their “bio-bar-code-based” DNA detection technique has a sensitivity similar to that of the commonly used polymerase chain reaction (PCR) method. The technique makes use of both gold nanoparticles attached to “bar code” DNA and magnetic microparticles.
“We are the first to demonstrate technology that can compete with - and beat - PCR in many of the relevant categories,” said Chad Mirkin, director of Northwestern’s Institute for Nanotechnology. “Nanoscience has made this possible. Our alternative method promises to bring diagnostics to places PCR is unlikely to go - the battlefield, the post office, a village in a developing country, the hospital and, perhaps ultimately, the home.”
To demonstrate their technique the researchers detected a DNA sequence associated with anthrax lethal factor (one of the three proteins making up anthrax toxin). They attached 30 nm diameter gold nanoparticles to two types of oligonucleotide sequence, one that was complementary to the target-DNA sequence and another that was complementary to a bar-code DNA sequence chosen as a unique identification tag for the target. Each gold nanoparticle was attached to around 360 oligonucleotide strands, and the ratio of bar-code-binding DNA to target-binding DNA was around 70 to 1. In fact the bio-bar-code amplification technique measures the bar-code DNA rather than the target DNA. Before starting the test the scientists loaded the functionalized gold nanoparticles with bar-code DNA using a hybridization process, so that the bar-code DNA attached to the nanoparticle became double-stranded.
The technique also employed iron-oxide magnetic microparticles linked to an oligonucleotide complementary to part of the target-DNA sequence. When both the gold nanoparticles and magnetic microparticles were introduced to the test sample, they attached themselves to any strands of the target-DNA sequence that were present. As a result, each target-DNA sequence became sandwiched between a gold nanoparticle and a magnetic microparticle. The researchers were then able to use a magnetic field to separate the target-DNA sequences and the unreacted magnetic microparticles from the other components.
Next, the scientists released the bar-code DNA from the gold nanoparticles by heating them, and used a chip-based method to detect the bar-codes. The chips contained spots of capture DNA. These attracted the bar-codes and a second set of gold nanoparticle probes. Enhancing the spots with silver enabled the researchers to measure the scattered light from the spots using a commercial reader. What’s more, the spot intensity gave a measure of the concentration of the target DNA.
The bar-code amplification technique was able to detect 10 strands of target DNA in a 30 microlitre sample, giving it a sensitivity comparable to those of PCR-based techniques. The technique was also able to differentiate single-base mismatches.
“For each molecule of captured target DNA, thousands of bar-code DNA strands are released, which is a powerful way of amplifying the signal for a DNA target of interest, such as anthrax,” said Mirkin. “There is power in its simplicity.”
The team has also used the bio-bar-code amplification technique to detect low levels of prostate-specific antigen, a protein that can be an indicator of prostate cancer.
The researchers reported their work in the Journal of the American Chemical Society.
About the author
Liz Kalaugher is editor of nanotechweb.org.