An ELISA-on-a-Chip Biosensor System Coupled with Immunomagnetic Separation for the Detection of Vibrio parahaemolyticus within a Single Working Day

Ahn

et al. 2015

Sci Rep

We validated a single-stranded, DNA aptamer-based, diagnostic method capable of detecting Lipocalin-2 (LCN2), a biomarker from clinically relevant hepatocellular carcinoma (HCC) patient serum, in the sandwich assay format. Nine aptamers (LCN2_apta1 to LCN2_apta9) for LCN2 were screened with SELEX processes, and a sandwich pair (LCN2_apta2 and LCN2_apta4) was finally chosen using surface plasmon resonance (SPR) and dot blotting analysis. The result of the proposed aptamer sandwich construction shows that LCN2 was sensitively detected in the concentration range of 2.5–500 ng mL−1 with a limit of detection of 0.6 ng mL−1. Quantitative measurement tests in HCC patients were run on straight serum and were compared with the performance of the conventional antibody-based ELISA kit. The aptamer sandwich assay demonstrated an excellent dynamic range for LCN2 at clinically relevant serum levels, covering sub-nanogram per mL concentrations. The new approach offers a simple and robust method for detecting serum biomarkers that have low and moderate abundance. It consists of functionalization, hybridization and signal read-out, and no dilution is required. The results of the study demonstrate the capability of the aptamer sandwich assay platform for diagnosing HCC and its potential applicability to the point-of-care testing (POCT) system.

Section: Discussionmentioning

confidence: 99%

Aptamer-based Sandwich Assay and its Clinical Outlooks for Detecting Lipocalin-2 in Hepatocellular Carcinoma (HCC)

Ahn

et al. 2015

Sci Rep

“…This approach can be applied to detect biomarkers within the low and medium ranges of protein concentration - [picograms per milliliter to micrograms per milliliter (as shown in Tables S4 and S5)]. To date, several studies have reported the biomarkers for multipurpose applications, such as disease monitoring, early detection, and prognosis. , Compared to our findings, most of these biomarkers could detect the target in a much higher range. ,, Although highly sensitive ELISA tests are very helpful for early detection with very-low-abundance biomarkers, − they are not suitable for long-term monitoring of the disease and predicting disease progression because of their overly elevated levels in the serum. Tables S4 and S5 list the broad-spectrum biomarkers that have been shown to exist at clinical levels in patient serum.…”

Section: Discussionmentioning

confidence: 99%

Aptamer-Based Sandwich Assay for Measurement of Thymidine Kinase 1 in Serum of Cancerous Patients

et al. 2019

Thymidine kinase 1 (TK1) is traditionally a serum biomarker that is elevated in the early stages of malignancies. The diagnostic and prognostic role of TK1 for screening and monitoring human malignancies has recently been investigated. Anti-human TK1 aptamers were selected through 12 iterative rounds of systematic evolution of ligands by exponential enrichment from a DNA library. The aptamer pool of round 12 was amplified, and the polymerase chain reaction product was cloned on the TA vector. Of the 85 colonies obtained, 52 were identified as positive clones. These aptamers were screened for TK1 with surface plasmon resonance, where apta37 and apta69 showed the highest affinity for TK1. The TK1_apta37 and TK1_apta69 aptamers were used in a sandwich assay platform and successfully detected TK1 in the concentration range of 54–3500 pg mL–1. Clinical samples from 60 cancerous patients were also tested with this assay system and compared using the conventional antibody-based enzyme-linked immunosorbent assay kit. The aptamer sandwich assay demonstrated a dynamic range for TK1 at clinically relevant serum levels, covering subpicogram per milliliter concentrations. The new approach offers a simple and robust method for detecting serum biomarkers that have low and moderate abundance. The results of this study demonstrate the screening capability of the aptamer sandwich assay platform and its potential applicability to the point-of-care testing system.

“…Enzyme-linked immunosorbent assay (ELISA) is another immunological technology, which can be employed in LOC systems for rapid online detection of foodborne pathogens (Wang and Salazar, 2016;Yoon and Kim, 2012). This technology can be integrated into microchannels of LOC systems instead of the conventional micro-well plates (Seo et al, 2010). In this method, the target solution will flow continuously in the microchannels and antibodies immobilized on the surface of the microchannel can subsequently capture pathogens.…”

Section: Microfluidics Based Immunological Detection Methodsmentioning

confidence: 99%

Microfluidic devices for sample preparation and rapid detection of foodborne pathogens

Kant

Shahbazi

Dave

et al. 2018

Biotechnology Advances

152

Rapid detection of foodborne pathogens at an early stage is imperative for preventing the outbreak of foodborne diseases, known as serious threats to human health. Conventional bacterial culturing methods for foodborne pathogen detection are time consuming, laborious, and with poor pathogen diagnosis competences. This has prompted researchers to call the current status of detection approaches into question and leverage new technologies for superior pathogen sensing outcomes. Novel strategies mainly rely on incorporating all the steps from sample preparation to detection in miniaturized devices for online monitoring of pathogens with high accuracy and sensitivity in a time-saving and cost effective manner. Lab on chip is a blooming area in diagnosis, which exploits different mechanical and biological techniques to detect very low concentrations of pathogens in food samples. This is achieved through streamlining the sample handling and concentrating procedures, which will subsequently reduce human errors and enhance the accuracy of the sensing methods. Integration of sample preparation techniques into these devices can effectively minimize the impact of complex food matrix on pathogen diagnosis and improve the limit of detections. Integration of pathogen capturing bio-receptors on microfluidic devices is a crucial step, which can facilitate recognition abilities in harsh chemical and physical conditions, offering a great commercial benefit to the food-manufacturing sector. This article reviews recent advances in current state-of-the-art of sample preparation and concentration from food matrices with focus on bacterial capturing methods and sensing technologies, along with their advantages and limitations when integrated into microfluidic devices for online rapid detection of pathogens in foods and food production line.