Abstract:Coronavirus disease 2019 (COVID-19) manifests with high clinical variability and warrants sensitive and specific assays to analyze immune responses in infected and vaccinated individuals.U sing Single Molecule Arrays( Simoa), we developed an assay to assess antibody neutralization with high sensitivity and multiplexing capabilities based on antibodymediated blockage of the ACE2-spike interaction. The assay does not require live viruses or cells and can be performed in ab iosafety level 2l aboratory within two … Show more
“…12 C). This device was also adapted for COVID-19 detection with LOD of 0.015 ng mL −1 for IgG and 0.099 pg mL −1 for N protein [ [371] , [372] , [373] , [374] ]. Fig.…”
Section: Luminescence Immunoassays For Covid-19 Poctsmentioning
“…12 C). This device was also adapted for COVID-19 detection with LOD of 0.015 ng mL −1 for IgG and 0.099 pg mL −1 for N protein [ [371] , [372] , [373] , [374] ]. Fig.…”
Section: Luminescence Immunoassays For Covid-19 Poctsmentioning
“…By distinguishing the number of fluorescence-positive microwells, subfemtomolar protein targets could be digitally detected. With this system, various types of biomarkers, including miRNAs and virus, were successfully analyzed [ 77 – 82 ]. Although the Simoa-based strategies can achieve efficient analysis of biomarkers, they subject to the low sample loading and the requirement of special instruments, which hamper their widespread application.…”
Section: Single-entity Counting-based Digital Bioassaysmentioning
Ultrasensitive detection of biomarkers is of paramount importance in various fields. Superior to the conventional ensemble measurement-based assays, single-entity assays, especially single-entity detection-based digital assays, not only can reach ultrahigh sensitivity, but also possess the potential to examine the heterogeneities among the individual target molecules within a population. In this review, we summarized the current biomolecular analysis methods that based on optical counting and imaging of the micro/nano-sized single entities that act as the individual reactors (e.g., micro-/nanoparticles, microemulsions, and microwells). We categorize the corresponding techniques as analog and digital single-entity assays and provide detailed information such as the design principles, the analytical performance, and their implementation in biomarker analysis in this work. We have also set critical comments on each technique from these aspects. At last, we reflect on the advantages and limitations of the optical single-entity counting and imaging methods for biomolecular assay and highlight future opportunities in this field.
“…The dSimoa platform achieved attomolar limits of detection for human interleukin under a dropcasting workflow. However, current digital bioaffinity assays, whether dSimoa or Simoa, rely on antibodies, necessitating a competitive format for small-molecule detection. ,,, These approaches suffer from a narrow dynamic concentration due to the restricted antibody microenvironment for antigen competitive reaction . Actually, much fewer single-molecule detection technologies are available for small molecules , compared with those for biomacromolecules such as proteins and nucleic acids.…”
Determination of trace amounts of targets or even a single molecule target has always been a challenge in the detection field. Digital measurement methods established for single molecule counting of proteins, such as single molecule arrays (Simoa) or dropcast single molecule assays (dSimoa), are not suitable for detecting small molecule, because of the limited category of small molecule antibodies and the weak signal that can be captured. To address this issue, we have developed a strategy for single molecule detection of small molecules, called small molecule detection with single molecule assays (smSimoa). In this strategy, an aptamer is used as a recognition element, and an addressable DNA Nanoflower (DNF) attached on the magnetic beads surface, which exhibit fluorescence imaging, is employed as the output signal. Accompanied by digital imaging and automated counting analysis, E2 at the attomolar level can be measured. The smSimoa breaks the barrier of small molecule detection concentration and provides a basis for high throughput detection of multiple substances with fluorescence encoded magnetic beads.
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