In vitro drug screening models derived from different PC12 cell lines for exploring Parkinson's disease based on electrochemical signals of catecholamine neurotransmitters.

Gold nanostructures and a Nafion modified screen-printed carbon electrode (Nafion/AuNS/SPCE) were developed to assess the cell viability of Parkinson's disease (PD) cell models. The electrochemical measurement of cell viability was reflected by catecholamine neurotransmitter (represented by dopamine) secretion capacity, followed by a traditional tetrazolium-based colorimetric assay for confirmation. Due to the  capacity to synthesize, store, and release catecholamines as well as their unlimited homogeneous proliferation, and ease of manipulation, pheochromocytoma (PC12) cells were used for PD cell modeling. Commercial low-differentiated and highly-differentiated PC12 cells, and home-made nerve growth factor (NGF) induced low-differentiated PC12 cells (NGF-differentiated PC12 cells) were included in the modeling. This approach achieved sensitive and rapid determination of cellular modeling and intervention states. Notably, among the three cell lines, NGF-differentiated PC12 cells displayed the enhanced neurotransmitter secretion level accompanied with attenuated growth rate, incremental dendrites in number and length that were highly resemble with neurons. Therefore, it was selected as the PD-tailorable modeling cell line. In short, the electrochemical sensor can be used to sensitively determine the biological function of neuron-like PC12 cells with negligible destruction and to explore the protective and regenerative impact of various substances on nerve cell model.

Precise Differentiation of Wobble-Type Allele via Ratiometric Design of a Ligase Chain Reaction-Based Electrochemical Biosensor for CYP2C19*2 Genotyping of Clinical Samples.

Due to the comparable stability between the perfect-base pair and the wobble-base pair, a precise differentiation of the wobble-type allele has remained a challenge, often leading to false results. Herein, we proposed a ligase chain reaction (LCR)-based ratiometric electrochemical DNA sensor, namely, R-eLCR, for a precise typing of the wobble-type allele, in which the traditionally recognized "negative" signal of wobble-base pair-mediated amplification was fully utilized as a "positive" one and a ratiometric readout mode was employed to ameliorated the underlying potential external influence and improved its detection accuracy in the typing of the wobble-type allele. The results showed that the ratio between current of methylene blue (IMB) and current of ferrocene (IFc) was partitioned in three regions and three types of wobble-type allele were thus precisely differentiated (AA homozygote: IMB/IFc > 2; GG homozygote: IMB/IFc < 1; GA heterozygote: 1 < IMB/IFc < 2); the proposed R-eLCR successfully discriminated the three types of CYP2C19*2 allele in nine cases of human whole blood samples, which was consistent with those of the sequencing method. These results evidence that the proposed R-eLCR can serve as an accurate and robust alternative for the identification of wobble-type allele, which lays a solid foundation and holds great potential for precision medicine.

Electrochemical immunosensor based on AuNPs/ERGO@CNT nanocomposites by one-step electrochemical co-reduction for sensitive detection of P-glycoprotein in serum.

P-glycoprotein (P-gp), a transmembrane glycoprotein widely expressed on the surface of various cells, is highly associated with multidrug resistance (MDR) that heralds the malignant progress of disease after drug treatment. Notably, there have been reported that serum P-gp is a potential marker for assessing the progression of disease resistance. Currently, there are few methods for point-of-care serum P-gp detection. In this study, we proposed a gold nanoparticles/electrochemically reduced graphene oxide@carbon nanotube (AuNPs/ERGO@CNT) modified immunosensor based on a one-step electrochemical co-reduction method. The limit of detection (LOD) of our constructed electrochemical immunosensor for P-gp detection reached 0.13 ng/mL, and the detection results in serum were consistent with ELISA. The developed immunosensor is expected to provide a scientific basis for the clinical application of serum P-gp monitoring and integrated medicine.

Multimodal Ochratoxin A-Aptasensor Using 3'-FAM-Enhanced Exonuclease I Tool and Magnetic Microbead Carrier.

Thanks to its preparatory ease, close affinity, and low cost, the aptasensor can serve as a promising substitute for antibody-dependent biosensors. However, the available aptasensors are mostly subject to a single-mode readout and the interference of unbound aptamers in solution and non-target-induced transition events. Herein, we proposed a multimodal aptasensor for multimode detection of ochratoxin A (OTA) with cross-validation using the 3'-6-carboxyfluorescein (FAM)-enhanced exonuclease I (Exo I) tool and magnetic microbead carrier. Specifically, the 3'-FAM-labeled aptamer/biotinylated-cDNA hybrids were immobilized onto streptavidin-magnetic microbeads via streptavidin-biotin interaction. With the presence of OTA, an antiparallel G-quadruplex conformation was formed, protecting the 3'-FAM labels from Exo I digestion, and then anti-FAM-horseradish peroxidase (HRP) was bound via specific antigen-antibody affinity; for the aptamers without the protection of OTA, the distal ssDNA was hydrolyzed from 3' → 5', releasing 3'-FAM labels to the solution. Therefore, the OTA was detected by analyzing the "signal-off" fluorescence of the supernatant and two "signal-on" signals in electrochemistry and colorimetry through the detection of the coating magnetic microbeads in HRP's substrate. The results showed that the 3'-FAM labels increased the activity of Exo I, producing a low background due to a more thorough digestion of unbound aptamers. The proposed multimodal aptasensor successfully detected the OTA in actual samples. This work first provides a novel strategy for the development of aptasensors with Exo I and 3'-FAM labels, broadening the application of aptamer in the multimode detection of small molecules.

[Study on difference of effect of Huoxue drugs and Yiqi Huoxue drugs on tumor metastasis based on immune remodeling].

Tumor metastasis is an important cause of tumor treatment failure. Its molecular mechanism is closely related to tumor cells remodeling immune cells and immunosuppressive microenvironment, so as to create a suitable soil for tumor cell invasion and growth. "Huoxue Huayu" is one of the important therapeutic principles in cancer treatment, but the influence of Huoxue drugs on tumor metastasis has been controversial in clinical application. In this paper, we systematically summarized the comparative study of Huoxue drugs and Yiqi Huoxue drugs in tumor metastasis in recent years, and discussed the differences of molecular mechanisms of Huoxue drugs and Yiqi Huoxue drugs in anti-tumor metastasis from the perspective of immune remodeling, so as to provide scientific basis for clinical rational application of Huoxue drugs and Yiqi Huoxue drugs.