Deep‐dLAMP: Deep Learning‐Enabled Polydisperse Emulsion‐Based Digital Loop‐Mediated Isothermal Amplification

Abstract

Digital nucleic acid amplification tests enable absolute quantification of nucleic acids, but the generation of uniform compartments and reading of the fluorescence requires specialized instruments that are costly, limiting their widespread applications. Here, the authors report deep learning‐enabled polydisperse emulsion‐based digital loop‐mediated isothermal amplification (deep‐dLAMP) for label‐free, low‐cost nucleic acid quantification. deep‐dLAMP performs LAMP reaction in polydisperse emulsions and uses a deep learning algorithm to segment and determine the occupancy status of each emulsion in images based on precipitated byproducts. The volume and occupancy data of the emulsions are then used to infer the nucleic acid concentration based on the Poisson distribution. deep‐dLAMP can accurately predict the sizes and occupancy status of each emulsion and provide accurate measurements of nucleic acid concentrations with a limit of detection of 5.6 copies µl‐1 and a dynamic range of 37.2 to 11000 copies µl‐1. In addition, deep‐dLAMP shows robust performance under various parameters, such as the vortexing time and image qualities. Leveraging the state‐of‐the‐art deep learning models, deep‐dLAMP represents a significant advancement in digital nucleic acid tests by significantly reducing the instrument cost. We envision deep‐dLAMP would be readily adopted by biomedical laboratories and be developed into a point‐of‐care digital nucleic acid test system.

CoID-LAMP: Color-Encoded, Intelligent Digital LAMP for Multiplex Nucleic Acid Quantification

“…The paired primer and sample droplets were then merged by destabilizing the interface using corona treatment, completing the mixing between the sample and different primers. The device was then incubated at 63 °C to perform LAMP, generating precipitates of magnesium pyrophosphate in droplets where the encapsulated target matched with the primer. − The droplets were then imaged on-chip to determine the target occupancy by detecting the presence of the precipitate and decode the encapsulated primer by detecting the droplet color, and the data were then used to quantify the concentrations of each target (see the Methods Section). The assay turnaround time was about 2 h, including 10 min for sample droplet generation, 15 min for sample droplet loading, 60 min for amplification, 20 min for chip imaging, and 10 min for data analysis.…”

Deep Learning-Assisted Droplet Digital PCR for Quantitative Detection of Human Coronavirus

“…These methods allow fully automated analysis of the dPCR images and enable high-throughput, accurate, and rapid detection. With the recent advances of deep learning technology, the image processing methods have further been improved [ 24 ]. After training dataset of the images, the location of the compartments is predicted by a convolutional neural network (CNN) and segmentation is conducted with remarkable accuracy.…”

Hydrogel Digital LAMP with Suppressed Nonspecific Amplification for Rapid Diagnostics of Fungal Disease in Fresh Fruits

“…alternata isolation culture and morphology scanning, consuming 7–8 days with low specificity . Recently, molecular techniques have been extensively applied for fungal species identification, such as polymerase chain reaction (PCR) and real-time PCR (qPCR) using specific primers to amplify target genomic segments. , Digital PCR is a single-molecule detection technique that dispenses reaction solution into thousands of microwells, followed by single nucleic acid amplification and end-point digital counting, which is faster and more sensitive than conventional semiquantitative methods. , However, PCR relies on the costly specialized equipment and thermal cycle with long amplification time . Alternatively, loop-mediated isothermal amplification (LAMP) has become an effective assay for point-of-care test. , LAMP can be operated at a constant temperature with high accuracy and high sensitivity. − Nevertheless, nonspecific amplification is an inevitable process in the rapid nucleic acid test, adversely affecting the diagnostic accuracy and sensitivity of fungal diseases with false positives.…”

“…16,17 However, PCR relies on the costly specialized equipment and thermal cycle with long amplification time. 18 Alternatively, loopmediated isothermal amplification (LAMP) has become an effective assay for point-of-care test. 19,20 LAMP can be operated at a constant temperature with high accuracy and high sensitivity.…”