Fluorescence Aptasensor of Tuberculosis Interferon-γ in Clinical Samples Regulated by Steric Hindrance and Selective Identification

Abstract

Although there are many interferon gamma (IFN-γ)-based tools for tuberculosis (TB) diagnosis, they are less sensitive and laborious. Here, we developed an IFN-γ aptasensor using pyrophosphate–cerium coordination polymeric nanoparticles (PPi-Ce CPNs) as signal reporters and a double-stranded DNA as a probe. The sensor was realized by sterically regulating the polymerization elongation of terminal deoxynucleotidyl transferase (TdT) and the selective recognition reaction of PPi-Ce CPNs. This method employs PPi-Ce CPNs to selectively identify Cu2+ and polyT-templated copper nanoparticles (Cu NPs), as well as a TdT-assisted amplification technique. Our data showed that under optimized experimental conditions, a limit of detection of as low as 0.25 fg/mL was achieved, with a linear range of 1–100 fg/mL, and a good target protein specificity. The detection sensitivity was an order of magnitude higher than that observed with Cu NPs when used as signal reporters. This IFN-γ quantification technique was further validated in clinical samples using 57 clinical TB patients (22 negative and 35 positive). Our findings agreed with those from enzyme-linked immunosorbent assay, GeneXpert MTB/rifampin assay, and polymerase chain reaction detection of TB-DNA and those from clinical imaging techniques. Therefore, our analytical system may provide an additional and more sensitive tool for the early diagnosis of TB.

Current progress of functional nanobiosensors for potential tuberculosis diagnosis: The novel way for TB control?

“…In a fluorescence nanobiosensor, when biometric elements or targeted biomolecules are labeled by fluorescent labels, the fluorescence intensity is used to reveal how strong about the interactions between the biometric molecules and the target biomolecules. Chen et al (2022) developed a novel aptamer sensor for IFN-γ analysis using PPI-CE CPNs (cerium pyrophosphate coordination polymer nanoparticles) as a signal reporter molecule and using double-stranded DNAs as probes. The sensor is implemented by spatially regulating polymeric extension of terminal deoxynucleotide transferase (TdT) as well as the selective recognition of PPiCe CPNs.…”

Steric Hindrance-Mediated Enzymatic Reaction Enable Homogeneous Dual Fluorescence Indicators Aptasensing of Hepatocellular Carcinoma CTCs

“…Nevertheless, due to the low concentration of CTCs, amplification is required after capture to achieve ultrasensitive detection. Most nucleic acid amplification techniques are enzyme based, including helicase, ligase, and polymerase. − Among them, terminal deoxynucleotidyl transferase (TdT) provides rapid amplification of nucleic acid without the need for a template or a specific primer. , Any single- or double-stranded DNA molecule with a protruding, concave, or smooth 3′-OH end can serve as a primer for TdT. The main product of TdT-mediated nucleic acid amplification is a long oligonucleotide chain (from tens to hundreds of bp) whose base sequence depends on the proportion of the substrate deoxyribonucleoside triphosphate (dNTP) composition .…”