One-Shot Single-Cell Proteome and Metabolome Analysis Strategy for the Same Single Cell.

Characterizing the profiles of proteome and metabolome at the single-cell level is of great significance in single-cell multiomic studies. Herein, we proposed a novel strategy called one-shot single-cell proteome and metabolome analysis (scPMA) to acquire the proteome and metabolome information in a single-cell individual in one injection of LC-MS/MS analysis. Based on the scPMA strategy, a total workflow was developed to achieve the single-cell capture, nanoliter-scale sample pretreatment, one-shot LC injection and separation of the enzyme-digested peptides and metabolites, and dual-zone MS/MS detection for proteome and metabolome profiling. Benefiting from the scPMA strategy, we realized dual-omic analysis of single tumor cells, including A549, HeLa, and HepG2 cells with 816, 578, and 293 protein groups and 72, 91, and 148 metabolites quantified on average. A single-cell perspective experiment for investigating the doxorubicin-induced antitumor effects in both the proteome and metabolome aspects was also performed.

Manganese(II)-doped zinc/germanium oxide nanoparticles as a viable fluorescent probe for visual and time-resolved fluorometric determination of ascorbic acid and its oxidase.

A method is described for the determination of ascorbic acid (AA) in complex biological fluids. It based on maganese(II)-doped zinc/germanium oxide nanoparticles (Mn@ZnGe NPs) with appealing time-resolved phosphorescence (TRP). TRP can provide a background-free reporter signal in analytical methods. The absorption of AA overlaps the excitation band of Mn@ZnGe NPs at 254 nm. This reduces the intensity of fluorescence via an inner filter effect (IFE) with increasing concentration of AA. Typical experimental conditions include an emission peak at 536 nm, a delay time of 50 µs and a counting time of 2 ms. This method can detect AA in a range of 5-500 µM with a 0.13 µM limit of detection. If AA is oxidized by the enzyme AA oxidase (AAOx), dehydroascorbic acid will be formed which doesn't absorb at 254 nm. Hence, the IFE cannot occur and fluorescence is not reduced. The strategy can be used to quantify AAOx in the activity range of 1-4 U·mL-1. By using a handheld UV lamp and a smart phone with a color-scanning feature, the feasibility for visual detection and real-time/onsite quantitative scanometric monitoring of AA and AAOx is demonstrated. Graphical abstract Schematic presentation of a fluorometric method for determination of ascorbic acid (AA) and ascorbic oxidase and a scanometric visual assay. It based on the use of maganese(II)-doped zinc/germanium oxide nanoparticles (Mn@ZnGe NPs) with appealing time-resolved phosphorescence (TRP) and the inner-filter effect (IFE) between AA and Mn@ZnGe NPs.

Pick-up single-cell proteomic analysis for quantifying up to 3000 proteins in a Mammalian cell.

The shotgun proteomic analysis is currently the most promising single-cell protein sequencing technology, however its identification level of ~1000 proteins per cell is still insufficient for practical applications. Here, we develop a pick-up single-cell proteomic analysis (PiSPA) workflow to achieve a deep identification capable of quantifying up to 3000 protein groups in a mammalian cell using the label-free quantitative method. The PiSPA workflow is specially established for single-cell samples mainly based on a nanoliter-scale microfluidic liquid handling robot, capable of achieving single-cell capture, pretreatment and injection under the pick-up operation strategy. Using this customized workflow with remarkable improvement in protein identification, 2449-3500, 2278-3257 and 1621-2904 protein groups are quantified in single A549 cells (n = 37), HeLa cells (n = 44) and U2OS cells (n = 27) under the DIA (MBR) mode, respectively. Benefiting from the flexible cell picking-up ability, we study HeLa cell migration at the single cell proteome level, demonstrating the potential in practical biological research from single-cell insight.

Simultaneous deep transcriptome and proteome profiling in a single mouse oocyte.

Although single-cell multi-omics technologies are undergoing rapid development, simultaneous transcriptome and proteome analysis of a single-cell individual still faces great challenges. Here, we developed a single-cell simultaneous transcriptome and proteome (scSTAP) analysis platform based on microfluidics, high-throughput sequencing, and mass spectrometry technology to achieve deep and joint quantitative analysis of transcriptome and proteome at the single-cell level, providing an important resource for understanding the relationship between transcription and translation in cells. This platform was applied to analyze single mouse oocytes at different meiotic maturation stages, reaching an average quantification depth of 19,948 genes and 2,663 protein groups in single mouse oocytes. In particular, we analyzed the correlation of individual RNA and protein pairs, as well as the meiosis regulatory network with unprecedented depth, and identified 30 transcript-protein pairs as specific oocyte maturational signatures, which could be productive for exploring transcriptional and translational regulatory features during oocyte meiosis.

Coordination polymers of Tb3+/Nucleotide as smart chemical nose/tongue toward pattern-recognition-based and time-resolved fluorescence sensing.

The abundant functional groups on guanosine monophosphate (GMP) make it possible to interact with various metal ions. The subtle difference in the structure of GMP and deoxy-guanosine monophosphate (dGMP) coupled with Tb3+ can be readily exploited to form two coordination polymers, which have been unveiled as two time-resolved fluorescence (TRF) sensing reporters (Tb-GMP and Tb-dGMP) in our study. Based on this finding, herein, we have proposed a novel TRF orthogonal sensing array (Tb-GMP/dGMP) for pattern-recognition-based sensing of various metal ions. In addition, upon integration of some thiol-affinity metal ions, Tb-GMP/dGMP can be further extended to construct two metal ion-involved pattern-recognition-based sensor arrays (Tb-GMP/dGMP-Cu, Tb-GMP/dGMP-Ag) for the TRF sensing different levels of disease-relevant biothiols in biofluids, illustrating the powerful and multifunctional capabilities of the Tb-GMP/dGMP system and would inspire simpler and more widespread designs of chemical nose/tongue-based applications.

Inorganic-Organic Hybrid Tongue-Mimic for Time-Resolved Luminescent Noninvasive Pattern and Chiral Recognition of Thiols in Biofluids toward Healthcare Monitoring.

In this work, manganese(II)-doped zinc/germanium oxide nanoparticles (Mn@ZGNPs) have been hydrothermally synthesized to equip with appealing time-resolved luminescence (TRL). Interestingly, we reveal that they can be readily quenched ("turn off") via a facile surface coating with bioinspired polydopamine (PDA) polymerized from dopamine (DA), resulting from PDA-triggered TRL resonance energy transfer (TRL-RET). By integrated with the thiol-induced inhibition of PDA formation, an ingenious inorganic-organic hybrid tongue-mimic sensor array is thus unveiled for noninvasive pattern recognition of thiols in biofluids in a TRL-RET-reversed "turn on" format toward healthcare monitoring. The sensing principle is based on the new finding that there are differential inhibitions from thiols against the polymerization of DA with various concentrations. Furthermore, density function theory (DFT) studies excellently prove our sensing principle and experimental results, reinforcing the power of the presented system. More importantly, chiral recognition of varied concentrations and mixtures of cysteine enantiomers using our platform are also been demonstrated, promising its practical usage. This is a novel concept of inorganic-organic hybrid-based pattern and chiral recognition platform for TRL background-free sensing and would sprout more novel relevant strategies toward broader applications.