Dual Enhancement with a Nanoparticle-Based Lateral Flow Biosensor for the Determination of DNA

ABSTRACT

An attractive methodology for the one-step signal amplification in a gold nanoparticle-based lateral flow biosensor was successfully applied for nucleic acid determination. This relatively new methodology, namely the “dual gold signal enhancement method” utilizes two gold nanoparticle-antibody conjugates, responsible for both the oligonucleotide detection, as well as a significant increase on the signal intensity of the biosensors’ test zone. Herein, the first conjugate consisted of anti-biotin antibody on gold nanoparticles, blocked with bovine serum albumin and the second conjugate contained anti-bovine serum albumin antibody on gold nanoparticles of different sizes, compared to the first conjugate. Any biotinylated oligonucleotide (polymerase chain reaction or oligonucleotide ligation reaction products) may be efficiently detected. The gold nanoparticle size was found to be the critical factor for the signal formation for the biosensor test and control zones. Gold nanoparticles with relative sizes of 30 nm for the first and 10 nm for the second conjugate were used. The application of the second conjugate, that is, 10-nm gold nanoparticle antibovine serum albumin, resulted in increased sensitivity. The detection limit was improved by approximately an order of magnitude compared to conventional methodology for a reference oligonucleotide. The reference oligonucleotide was detected as low as 0.05 fmol in twenty minutes by using the dual gold signal enhancement. To our knowledge, this is the first time that the methodology has been reported on a nucleic acid lateral flow biosensor.

KEYWORDS:

• Dual gold nanoparticle conjugates
• gold nanoparticles
• lateral flow strip biosensor
• nucleotide detection

Conflicts of interest

The authors declare no conflict of interest. The founding sponsors had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; and in the decision to publish the results.

Acknowledgments

A. Sklavounos and Prof. A. C. Calokerinos (National and Kapodistrian University of Athens, Department of Chemistry) are acknowledged for performing the ultraviolet-visible spectroscopy measurements.