Design and Development of a Novel Lateral Flow Strip Test Based on Gold-Silver Nanoparticles for the Rapid Detection of Human Papillomavirus

Human papillomavirus (HPV) is considered one of the main causative agents of cervical cancer, and its rapid and accurate detection plays a critical role in the timely prevention and treatment of the disease. The present study involved the synthesis and subsequent characterization of gold-silver nanoparticles, conjugating anti-HPV antibodies with these nanoparticles, and designing a rapid detection kit based on a lateral flow assay (LFA) to identify HPV antigens. Initially, gold-silver nanoparticles were synthesized using the citrate reduction method and characterized via UV-Vis spectroscopy, dynamic light scattering (DLS), scanning electron microscopy (SEM), and Fourier-transform infrared spectroscopy (FTIR). Subsequently, anti-HPV antibodies were conjugated to the surface of the nanoparticles using the carbodiimide method. Different concentrations of HPV antigen, ranging from 1 nanomolar to 10⁻⁹ nanomolar, along with positive and negative clinical samples, were examined to determine the minimum detectable antigen concentration and the shortest detection time. This study demonstrated that the gold-silver nanoparticles were synthesized by reducing chloroauric acid and silver nitrate, with a resultant color change of the solution to reddish-brown. The UV-Vis spectrum of the synthesized nanoparticles exhibited a maximum absorbance peak at 528 nm. SEM images confirmed the presence of approximately spherical nanoparticles ranging in size from 50 to 250 nm, with some degree of aggregation and adhesion. DLS analysis revealed that particle sizes ranged from 90 to 250 nm, with an average diameter of 170 nm and a zeta potential of –18 mV . FTIR spectra of both the antibody-conjugated nanoparticles and the free antibodies showed characteristic peaks in the wavenumber regions corresponding to amide bands (Amide I, II, III), indicating successful antibody attachment to the synthesized nanoparticle surfaces. Furthermore, results obtained from clinical samples indicated a minimum detection time of 30 seconds, with the sensitivity, specificity, accuracy, and precision of the designed kit’s all reaching 100%.