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    Comparison of Six Aptamer-Aptamer Pairs on Rapid Detection of SARS-CoV-2 by Lateral Flow Assay
    (Oxford Univ Press Inc, 2024) Derin, Dilek Cam; Gultekin, Enes; Gunduz, Elif; Otlu, Baris
    Background SARS-CoV-2 is a threat to humanity. Both the spike (S) protein and its receptor binding domain (sRBD) are extensively used for rapid detection. Although real-time reverse transcription polymerase chain reaction (rRT-PCR) is mostly used method for the molecular detection of SARS-CoV-2, rapid assays for antigenic detection are always needed. Lateral flow assays (LFAs) are the most commonly used tests for this purpose, and aptamers having stability and long shelf life are used as capture reagents.Objective This study aimed to develop the LFAs based on the aptamer pairs for the antigenic detection of SARS-CoV-2 with the naked eye.Method Gold nanoparticles (AuNPs) were used as labels, and six sandwich models by three different aptamers were prepared using 4 mu M and 8 mu M probes and two kinds of membranes for developing the LFAs.Results The 8 mu M probe concentration and M2 membrane showed the best recognition of both the synthetic sRBD and SARS-CoV-2 coming from the naso/oropharingeal swabs by designed LFAs as 100% sensitivity and 93.3% specificity compared to the antibody-detecting LFAs.Conclusions Our developed strip assays based on aptamer pairs recognized the target directly in 5-6 min with the naked eye. It was also concluded that aptamer pairs, membrane types, assay buffers, and probe concentrations have a significant role in the detection of SARS-CoV-2 by LFAs.Highlights The detection of SARS-CoV-2 in clinical samples was demonstrated with the best aptamer pairs, sensitively and selectively among the designed six aptamer pairs for LFAs. Developed LFAs can be an alternative method to the conventional antibody-based LFAs for SARS-CoV-2 detection.
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    Multiple detection method for SARS-CoV-2 with aptamer cocktails depending on their localization in paper-based assays
    (Bmc, 2025) Cam Derin, Dilek; Gunduz, Elif
    BackgroundThe nucleocapsid (N) and spike (S) proteins are key markers for SARS-CoV-2 diagnosis and treatment because of their antigenic regions. Aptamers targeting these proteins have been developed for various diagnostic platforms, including Lateral Flow Assays (LFAs). However, existing studies have focused mainly on the use of single aptamer or antibody/aptamer sandwiches in capture zones, lacking the advantages of using aptamer cocktails. The present study aimed to develop multiple aptamer cocktail-based lateral flow assays (mACLFAs) for rapid and accurate detection of SARS-CoV-2 in clinical samples.MethodsSpherical gold nanoparticles were synthesized and used as labels for aptamer conjugation for naked-eye analysis. Aptamers specific to the S and N proteins of SARS-CoV-2 were chosen and used as either capture or detection reagents for the sandwich assay model. A nitrocellulose membrane was used to develop the mACLFAs.ResultsModel 2 was the most efficient multiple-assay model for viral diagnosis, although models 1 and 3 correctly detected the virus in clinical samples. Model 4 and Model 5 had nonspecific interactions and were not preferred for multiple assay development. The specificity of models 1, 2 and 3 was 100%, and the sensitivity of models 2 and 3 was 100%, while it was 96.6% for Model 1.ConclusionOur findings demonstrated that the accuracy of viral detection was improved by the use of aptamer cocktails, as both the N and S proteins were captured in clinical samples. The results also highlighted the importance of the position of the detector aptamers in the capture zones in terms of the accuracy and interaction of the aptamers in the sandwich assay. Thus, these findings will be valuable for developing diagnostic tools to monitor and prevent future outbreaks, such as monkey pox virus.