Direct colorimetric diagnosis of pathogen infections by utilizing thiol-labeled PCR primers and unmodified gold nanoparticles

Abstract

We describe here a greatly simplified colorimetric detection method to identify PCR-amplified nucleic acids. Our method relies on the PCR product having thiol group at one end, which is generated by employing thiolated PCR primer. After PCR amplification reaction, unmodified gold nanoparticles (AuNPs) are added into the reaction tube followed by the addition of NaCl to induce the aggregation of AuNPs. The PCR products strongly bind to the surface of AuNPs through the interaction of the terminal thiol groups and the long chain of DNA which has abundant negative charges enhances the electrostatic and steric repulsion among AuNPs, which consequently leads to the prevention of the salt-induced aggregation. As a result, the color of AuNPs remains red in the presence of the PCR-amplified nucleic acids, while the AuNPs change its color from red to blue due to the salt-induced aggregation in the absence of the PCR products. This simple but very efficient colorimetric strategy was successfully demonstrated by diagnosing Chlamydia infection using a real human urine sample. Since the results can be clearly seen with the naked eye without any complicated step such as surface modification of AuNPs or PCR product purification, this method can be easily applied to point-of-care diagnosis.