Morphological determination of localization and function of Golgi proteins.

In animal cells, the Golgi apparatus serves as the central hub of the endomembrane secretory pathway. It is responsible for the processing, modification, and sorting of proteins and lipids. The unique stacking and ribbon-like architecture of the Golgi apparatus forms the foundation for its precise functionality. Under cellular stress or pathological conditions, the structure of the Golgi and its important glycosylation modification function may change. It is crucial to employ suitable methodologies to study the structure and function of the Golgi apparatus, particularly when assessing the involvement of a target protein in Golgi regulation. This article provides a comprehensive overview of the diverse microscopy techniques used to determine the specific location of the target protein within the Golgi apparatus. Additionally, it outlines methods for assessing changes in the Golgi structure and its glycosylation modification function following the knockout of the target gene.

Electrochemiluminescence cytosensing platform based on Ru(bpy)32+@silica-Au nanocomposite as luminophore and AuPd nanoparticles as coreaction accelerator for in situ evaluation of intracellular H2O2.

An electrochemiluminescence (ECL) cytosensor was fabricated onto a microfluidic paper-based analytical device (µ-PAD) in order to detect hydrogen peroxide (H2O2) which was released from tumor cells. The ECL probe Ru(bpy)32+@silica-Au nanocomposite (Ru@SiO2-Au) was fabricated and served as ECL reagent. The ECL of Ru@SiO2-Au nanocomposite was quenched by the ferrocene (Fc). AuPd nanoparticles (AuPd NPs), which were modified on the paper working electrode (PWE), were served as the catalyst of H2O2 to produce hydroxyl radicals (•OH) for cleaving Fc-labelled DNA to achieve "signal-on", and AuPd NPs also severed as coreaction accelerator. H2O2 was released from cells under the stimulation of phorbol myristate acetate. Fc-labelled DNA strand was cleaved via •OH. Fc molecule departed from the PWE surface, The ECL could be recovered. An ECL cytosensor on a 3D origami device was constructed. The ECL response of the Ru@SiO2-Au-Fc system was related to the number of cells. The ECL intensity was quantitatively related with the logarithm of MCF-7 cells number and H2O2 concentration, the detection limit was 30 cells mL-1. As a consequence, this work provided a really low-cost and disposable µ-PAD for sensitive detection of intracellular H2O2. What's more, this work had potential application value for studying cellular biology and pathophysiology.

Ultrasensitive Microfluidic Paper-Based Electrochemical Biosensor Based on Molecularly Imprinted Film and Boronate Affinity Sandwich Assay for Glycoprotein Detection.

In this work, we proposed a strategy that combined molecularly imprinted polymers (MIPs) and hybridization chain reaction into microfluidic paper-based analytical devices for ultrasensitive detection of target glycoprotein ovalbumin (OVA). During the fabrication, Au nanorods with a large surface area and superior conductibility were grown on paper cellulosic fiber as a matrix to introduce a boronate affinity sandwich assay. The composite of MIPs including 4-mercaptophenylboronic acid (MPBA) was able to capture target glycoprotein OVA. SiO2@Au nanocomposites labeled MPBA and cerium dioxide (CeO2)-modified nicked DNA double-strand polymers (SiO2@Au/dsDNA/CeO2) as a signal tag were captured into the surface of the electrode in the presence of OVA. An electrochemical signal was generated by using nanoceria as redox-active catalytic amplifiers in the presence of 1-naphthol in electrochemical assays. As a result, the electrochemical assay was fabricated and could be applied in the detection of OVA in the wide linear range of 1 pg/mL to 1000 ng/mL with a relatively low detection limit of 0.87 pg/mL (S/N = 3). The results indicated that the proposed platform possessed potential applications in clinical diagnosis and other related fields.

Polyhedral-AuPd nanoparticles-based dual-mode cytosensor with turn on enable signal for highly sensitive cell evalution on lab-on-paper device.

A novel dual-mode cytosensor based on polyhedral AuPd alloy nanoparticles (PH-AuPd NPs) and three-dimentional reduced graphene oxide (3D-rGO) was constructed for highly sensitive detection of MCF-7 cells. The 3D-rGO was in situ synthesized on the paper working electrode (PWE) by a pollution-free hydrothermal method, increasing the specific surface area and further facilitating the modification of Au nanoparticles (AuNPs). After modified with AuNPs, the Au@ 3D-rGO/PWE was then functionalized by aptamer H1 to trap MCF-7 cells. To construct the cytosensor, PH-AuPd NPs was prepared as a novel catalytic material, and further modified with aptamer H2 for recognizing MCF-7 cells. With the occurrence of efficient recognition of MCF-7 cells, PH-AuPd NPs were bound onto the surface of the cells, and could catalyze H2O2 to generate •OH, leading to an amplified electrochemical signal. Meanwhile, as the electrolyte solution flowed, the •OH are transferred outward to the colorimetric detection zone, and catalyzed a chromogenic substrate TMB forms a colored product. The electrical signal measurement and colorimetric detection were carried out on a compatibly designed lab-on-paper device (LPD), realizing a dual-mode signal readout. This paper-based dual-mode cytosensor provided a relatively low detection limit of 20 cells mL-1 and a sensitive detection from 50 cells mL-1 to 107 cells mL-1 for MCF-7 cells, providing a reliable pathway of sensitively detecting cancer cells in clinical applications.

Electrochemiluminescence based detection of microRNA by applying an amplification strategy and Hg(II)-triggered disassembly of a metal organic frameworks functionalized with ruthenium(II)tris(bipyridine).

An electrochemiluminescence (ECL) biosensor is described for the detection of microRNA (miRNA-155) based on tris(bipyridine)ruthenium(II) functionalized metal organic framework (RuMOF) materials. The material was prepared by a solvothermal method and was found to be stable even in acidic solution. However, it is selectively and sensitively disassembled by Hg(II) ions, resulting in the release of large quantities of Ru(II)(bpy)3 ions, which produces a strong ECL signal. In view of the ion-selective disassembly and release and strand displacement process, an ultrasensitive ECL sensing method was established for detection of microRNAs. In the presence of the target, the hairpin structure of H1 can open and hybridize with the hairpin probe H2 to form a more stable H1-H2 duplex structure than the H1-target hybrid. The target of hybridization to H1 was immediately freed from the structure and the released target re-entered the new hairpin assembly target recovery process. The remaining H2 single fragment can bind to the I-RuMOFs-conjugates. The more hairpin probes H1, the more I-RuMOFs-conjugates load the DNA fragments, leading to the signal amplification. The method works in the 0.8 f. to 1.0 nM miRNA-155 concentration range and has a detection limit of 0.3 fM. The assay is sensitive, fairly specific and remarkably stable. In our perception, it offers an attractive tool for the sensitive detection of microRNAs in clinical samples. Graphical abstract An electrochemiluminescence (ECL) based biosensor is described for the detection of microRNA (miRNA-155) based on the use of a metal organic framework functionalized with ruthenium(II)tris(bipyridine) that was deposited on a glassy carbon electrode (GCE) modified with gold nanoparticles.

Ultrasensitive microfluidic paper-based electrochemical/visual biosensor based on spherical-like cerium dioxide catalyst for miR-21 detection.

In this work, an electrochemical biosensor based on Au nanorods (NRs) modified microfluidic paper-based analytical devices (µPADs) were constructed for sensitive detection of microRNA (miRNA) by using cerium dioxide - Au@glucose oxidase (CeO2-Au@GOx) as an electrochemical probe for signal amplification. Au NRs were synthesized by in-situ growth method in µPADs surface to enhance the conductivity and modified hairpin probe through Au-S bonds. The construction of "the signal transducer layer" was carried out by GOx catalyzing glucose to produce H2O2, which was further electrocatalyzed by CeO2. After the biosensor was constructed, an obvious electrochemical signal was observed from the reduction of H2O2. In order to make the detection more convincing, the visual detection was performed based on the oxidation of 3,3',5,5'-tetramethylbenzidine by H2O2 with the help of Exonuclease I. The electrochemical biosensor provided a wide linear range of 1.0fM to 1000fM with a relatively low detection limit of 0.434fM by the electrochemical measurement. Linear range of 10fM to 1000fM with a relatively low detection limit of 7.382fM was obtained by visual detection. The results indicated the proposed platform has potential utility for detection of miRNA.