Limits...
Magnetically assisted DNA assays: high selectivity using conjugated polymers for amplified fluorescent transduction.

Xu H, Wu H, Huang F, Song S, Li W, Cao Y, Fan C - Nucleic Acids Res. (2005)

Bottom Line: We demonstrate that the use of magnetic microparticles significantly improves the selectivity of this class of DNA sensors.Compared with previously reported DNA sensors with CP amplification, this novel sensing strategy displays excellent discrimination against non-cognate DNA in the presence of a protein mixture or even human serum.We also demonstrate that the magnetically assisted DNA sensor can conveniently identify even a single-nucleotide mismatch in the target sequence.

View Article: PubMed Central - PubMed

Affiliation: Division of Nanobiology and Nanomedicine, Shanghai Institute of Applied Physics, Chinese Academy of Sciences Shanghai 201800, China.

ABSTRACT
We report a strategy for conjugated polymer (CP)-based optical DNA detection with improved selectivity. The high sensitivity of CP-based biosensors arises from light harvesting by the CP and the related amplified fluorescent signal transduction. We demonstrate that the use of magnetic microparticles significantly improves the selectivity of this class of DNA sensors. Compared with previously reported DNA sensors with CP amplification, this novel sensing strategy displays excellent discrimination against non-cognate DNA in the presence of a protein mixture or even human serum. We also demonstrate that the magnetically assisted DNA sensor can conveniently identify even a single-nucleotide mismatch in the target sequence.

Show MeSH
Histograms for fluorescence intensity at different conditions. From left to right: 1 nM target DNA in the pure buffer; 1 nM target DNA in the artificial matrix containing proteins; 1 nM target DNA in the human serum sample; 5 μM non-cognate DNA in the human serum sample and a human serum sample ([PF] = 5.4 × 10−7 M). Error bars were estimated from at least three independent measurements.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC1131937&req=5

fig4: Histograms for fluorescence intensity at different conditions. From left to right: 1 nM target DNA in the pure buffer; 1 nM target DNA in the artificial matrix containing proteins; 1 nM target DNA in the human serum sample; 5 μM non-cognate DNA in the human serum sample and a human serum sample ([PF] = 5.4 × 10−7 M). Error bars were estimated from at least three independent measurements.

Mentions: In an attempt to test the applicability of this DNA sensor to detection in real samples, we prepared an artificial matrix containing 1 nM target DNA 3 in a solution containing a 1000-fold higher concentration of proteins (Hb, Lys and BSA) and random DNA sequences. As a further step, we employed diluted human serum samples (1:10) containing either target DNA or non-complementary DNA. With the assistance of magnetic separation, we were able to observe FRET signals corresponding to the presence of target DNA. More importantly, we have shown that the FRET signal intensity is comparable to that obtained in pure DNA solutions (Figure 4), implying that the DNA detection is insensitive to non-specific species, and that it is possible to perform DNA detection even in blood samples.


Magnetically assisted DNA assays: high selectivity using conjugated polymers for amplified fluorescent transduction.

Xu H, Wu H, Huang F, Song S, Li W, Cao Y, Fan C - Nucleic Acids Res. (2005)

Histograms for fluorescence intensity at different conditions. From left to right: 1 nM target DNA in the pure buffer; 1 nM target DNA in the artificial matrix containing proteins; 1 nM target DNA in the human serum sample; 5 μM non-cognate DNA in the human serum sample and a human serum sample ([PF] = 5.4 × 10−7 M). Error bars were estimated from at least three independent measurements.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC1131937&req=5

fig4: Histograms for fluorescence intensity at different conditions. From left to right: 1 nM target DNA in the pure buffer; 1 nM target DNA in the artificial matrix containing proteins; 1 nM target DNA in the human serum sample; 5 μM non-cognate DNA in the human serum sample and a human serum sample ([PF] = 5.4 × 10−7 M). Error bars were estimated from at least three independent measurements.
Mentions: In an attempt to test the applicability of this DNA sensor to detection in real samples, we prepared an artificial matrix containing 1 nM target DNA 3 in a solution containing a 1000-fold higher concentration of proteins (Hb, Lys and BSA) and random DNA sequences. As a further step, we employed diluted human serum samples (1:10) containing either target DNA or non-complementary DNA. With the assistance of magnetic separation, we were able to observe FRET signals corresponding to the presence of target DNA. More importantly, we have shown that the FRET signal intensity is comparable to that obtained in pure DNA solutions (Figure 4), implying that the DNA detection is insensitive to non-specific species, and that it is possible to perform DNA detection even in blood samples.

Bottom Line: We demonstrate that the use of magnetic microparticles significantly improves the selectivity of this class of DNA sensors.Compared with previously reported DNA sensors with CP amplification, this novel sensing strategy displays excellent discrimination against non-cognate DNA in the presence of a protein mixture or even human serum.We also demonstrate that the magnetically assisted DNA sensor can conveniently identify even a single-nucleotide mismatch in the target sequence.

View Article: PubMed Central - PubMed

Affiliation: Division of Nanobiology and Nanomedicine, Shanghai Institute of Applied Physics, Chinese Academy of Sciences Shanghai 201800, China.

ABSTRACT
We report a strategy for conjugated polymer (CP)-based optical DNA detection with improved selectivity. The high sensitivity of CP-based biosensors arises from light harvesting by the CP and the related amplified fluorescent signal transduction. We demonstrate that the use of magnetic microparticles significantly improves the selectivity of this class of DNA sensors. Compared with previously reported DNA sensors with CP amplification, this novel sensing strategy displays excellent discrimination against non-cognate DNA in the presence of a protein mixture or even human serum. We also demonstrate that the magnetically assisted DNA sensor can conveniently identify even a single-nucleotide mismatch in the target sequence.

Show MeSH