Sensitive detection of alkaline phosphatase based on terminal deoxynucleotidyl transferase and endonuclease IV-assisted exponential signal amplification

Single-Molecule Biosensing of Alkaline Phosphatase in Cells and Serum Based on Dephosphorylation-Triggered Catalytic Assembly and Disassembly of the Fluorescent DNA Chain

“…The integration of these two technologies will allow the simple detection of ALP with high accuracy and sensitivity. In comparison with the reported fluorescent ALP sensors, the proposed biosensor is much simpler by eliminating laborious procedures for the preparation of functional fluorescent nanomaterials − and small-molecule fluorescent probes, − and avoiding the enzyme-assisted nucleic acid amplification for ALP signal amplification, ,− Although it cannot be applied for the imaging of ALP activity in living cells like some reported fluorescent ALP assays, , the proposed ALP biosensor can sensitively detect ALP in cell extracts and serum samples.…”

“…We have constructed a new single-molecule fluorescent biosensor for accurate and sensitive monitoring of ALP level in human cells and clinical serum samples. All probes involved in this biosensor are commercially available, which avoids laborious procedures for the preparation of functional fluorescent nanomaterials − and small-molecule fluorescent probes. − Superior to enzyme-assisted nucleic acid amplification-based ALP assays, ,− the signal amplification and transduction involved in this biosensor can be achieved with enzyme-free assembly and disassembly processes, greatly simplifying the assay procedures and eliminating nonspecific signals. This biosensor can sensitively detect ALP activity with a detection limit of 2.61 × 10 –6 U/mL, and it can be applied for ALP inhibition assays and kinetic analysis.…”

“…Moreover, non-nucleic acid molecules, including phenolphthalein monophosphate, hydroquinone diphosphate, peptide, , p -nitrophenylphosphate (PNPP), pyrophosphate, , guanine monophosphate, and adenosine triphosphate, are frequently exploited as the ALP sensing substrates, but they are not easily amplified like DNA/RNA and are not suitable for accurate detection of low-abundant ALP target in rare samples. To overcome this issue, a few nucleic acid signal amplification methods, such as DNA ligase-mediated ligase amplification reaction, DNA polymerase- and nicking enzyme-mediated circular exponential amplification, DNA RNA polymerase- and Cas13a nuclease-mediated RNA transcription-coupled cyclic cleavage reaction, RNA polymerase- and duplex specific nuclease-mediated RNA transcription-induced dual signal amplification, and DNA polymerase- and endonuclease-mediated exponential signal amplification, have been adapted for amplified detection of ALP, but they require multiple specific enzymes to trigger the amplification system, which inevitably increases the assay cost and complicates the reaction process. Besides, the instability of enzyme activities and frequent nonspecific amplification may impair the assay accuracy and reproducibility.…”

Ginsenoside Rk3 is a novel PI3K/AKT-targeting therapeutics agent that regulates autophagy and apoptosis in hepatocellular carcinoma

“…Pathological staining was performed according to previous studies with slight modifications [ [34] , [35] , [36] , [37] ]. Liver, tumor, and other tissues were fixed with 10% formalin and then embedded in paraffin.…”