摘要：

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 for 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, i.e., 10 nm gold nanoparticle anti-bovine 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. As low as 0.05 fmol of the reference oligonucleotide was detected, 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.

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