A near-infrared xanthene-based fluorescent probe for selective detection of hydrazine and its application in living cells.

Developing fluorescent probes for selective determination of the toxic and carcinogenic hydrazine are pretty significant. Herein, a rhodamine dye coupled to naphthalene was selected as a near-infrared fluorophore and acetyl group as a trigger unit for hydrazine sensing with a Stokes shifts of 62 nm. The probe showed about 77-fold NIR fluorescence enhancement in the presence of hydrazine. In addition, the detection limit was as low as 3.4 ppb, and the fluorescence intensity at 654 nm showed a satisfactory linearity with the concentration range of hydrazine from 0 to 120 µM. More importantly, the practical utility of probe has been successfully proved through the fluorescence bioimaging of hydrazine in living cells with low cytotoxicity and quantitative N2H4 detection in environmental water samples.

A colorimetric and turn-on NIR fluorescent probe based on xanthene system for sensitive detection of thiophenol and its application in bioimaging.

Developing fluorescent probes for specific detection of extremely toxic thiophenols is pretty significant in the field of environment, chemistry and biology. We report herein a turn-on red fluorescent xanthene-based probe (RD-Probe) for detecting thiophenol with high selectivity over other analytes including aliphatic thiols. The probe could play the part of a "naked-eye"colorimetric indicator toward thiophenol. Moreover, the relative fluorescence intensity at 653 nm displayed good linearity with the concentration of thiophenol ranging from 0 to 6 µM, and the limit of detection for thiophenol could be as low as 15 nM. Furthermore, the practicability of RD-Probe has been successfully proved through the quantitative thiophenol detection in real water samples and fluorescence bioimaging of thiophenol in HeLa cells.

Reaction-based fluorescent probe for the selective and sensitive detection of thiophenols with a large Stokes shift and its application in water samples.

Although widely used in organic synthesis, pharmaceuticals and agrochemicals, thiophenol has brought about a series of ecological problems due to its high toxicity. Therefore, the development of efficient methods to discriminate thiophenols from aliphatic thiols is of great importance. In this work, a new reaction-based turn-on red fluorescence probe for the detection of thiophenols has been developed for the first time by employing dicyanomethylene-4H-pyran (DCM) as a fluorescence reporter and 2,4-dinitrobenzene-sulfonate (DNBS) as a recognition unit. The probe displayed a highly selective and sensitive (63 fold-fluorescence enhancement) response to thiophenols over aliphatic thiols. Additionally, the probe also exhibited a large Stokes shift (159 nm) and the detection limit reached as low as 8.3 nM. Moreover, this probe was also proved suitable for the quantification of thiophenol in real environmental water samples.

Dissection of DEN-induced platelet proteome changes reveals the progressively dys-regulated pathways indicative of hepatocarcinogenesis.

Due to the lack of precise markers indicative of its occurrence, progression, and malignant stages, hepatocellular carcinoma (HCC) is currently associated with high mortality. Given the fact that thrombocytopenia is associated with chronic liver diseases, and the multifunctional nature of platelets we reason that phenotype-specific platelets could be the systemic barometer for hepato-carcinogenesis. The mass spectrometry (MS)-based proteomic efforts to discover novel biomarkers in plasma or serum are largely compromised by a few of the overwhelmingly abundant proteins that comprise over 95% of the total protein mass of plasma or sera. Platelets however are free of these MS signal-suppressing proteins. On the basis of a HCC animal model where diethyl nitrosamine (DEN) administration on male rats specifically induces HCC, by using a multiplex quantitative proteomic approach, we profiled the phase-to-phase proteome changes in a series of viable phenotype-specific platelets along with the DEN-induced progressive liver transformation. The platelet proteome was found highly responsive to each physiological stage of liver inflammation or pathogenesis. Using data-dependent bioinformatics network analysis, we found that certain pathway modules involved in immune response, tissue wound repair, apoptosis, cell proliferation, and catabolism and metabolism were differentially regulated, which were uncovered by the DEN-induced differential expression of the corresponding pathway components. The phase-specific presentations of these pathways suggested that the DEN-induced progression of immune suppression and apoptosis resistance is dynamically coordinated in the platelets. These novel platelet signatures are interconnected in the dynamic networks along with HCC progression and could be identified noninvasively for HCC prognosis and early diagnosis.

Global analysis of the rat and human platelet proteome - the molecular blueprint for illustrating multi-functional platelets and cross-species function evolution.

Emerging evidences indicate that blood platelets function in multiple biological processes including immune response, bone metastasis and liver regeneration in addition to their known roles in hemostasis and thrombosis. Global elucidation of platelet proteome will provide the molecular base of these platelet functions. Here, we set up a high-throughput platform for maximum exploration of the rat/human platelet proteome using integrated proteomic technologies, and then applied to identify the largest number of the proteins expressed in both rat and human platelets. After stringent statistical filtration, a total of 837 unique proteins matched with at least two unique peptides were precisely identified, making it the first comprehensive protein database so far for rat platelets. Meanwhile, quantitative analyses of the thrombin-stimulated platelets offered great insights into the biological functions of platelet proteins and therefore confirmed our global profiling data. A comparative proteomic analysis between rat and human platelets was also conducted, which revealed not only a significant similarity, but also an across-species evolutionary link that the orthologous proteins representing "core proteome", and the "evolutionary proteome" is actually a relatively static proteome.

Modulation of TLR signaling by multiple MyD88-interacting partners including leucine-rich repeat Fli-I-interacting proteins.

Emerging evidences suggest TLR-mediated signaling is tightly regulated by a specific chain of intracellular protein-protein interactions, some of which are yet to be identified. Previously we utilized a dual-tagging quantitative proteomics approach to uncover MyD88 interactions in LPS-stimulated cells and described the function of Fliih, a leucine-rich repeat (LRR) protein that negatively regulates NF-kappaB activity. Here we characterize two distinct LRR-binding MyD88 interactors, LRRFIP2 and Flap-1, and found that both are positive regulators of NF-kappaB activity. Upon LPS stimulation, LRRFIP2 was also found to positively regulate cytokine production in macrophages, suggesting a functional role in TLR4-mediated inflammatory response. Furthermore, we observed that immediately following LPS stimulation both LRRFIP2 and Flap-1 compete with Fliih for interacting with MyD88 to activate the signaling. By using a novel multiplex quantitative proteomic approach, we found that at endogenous levels these positive and negative regulators interact with MyD88 in a timely and orderly manner to differentially mediate the NF-kappaB activity through the course of signaling from initiation to prolongation, and to repression. Based on these data, we describe a mechanistic model in which selective modulation of TLR signaling is achieved by temporal and dynamic interactions of MyD88 with its regulators.

Preparation of Fe3O4@ZrO2 core-shell microspheres as affinity probes for selective enrichment and direct determination of phosphopeptides using matrix-assisted laser desorption ionization mass spectrometry.

Fe3O4@ZrO2 microspheres with well-defined core-shell structure were prepared and applied for the highly selective enrichment of phosphopeptides from tryptic digest product of proteins. To successfully coat iron oxide microspheres with uniform zirconia shell, magnetic Fe3O4 microspheres were first synthesized via a solvothermal reaction, followed by being coated with a thin layer of carbon by polymerization and carbonization of glucose through hydrothermal reaction. Finally, with the use of the Fe3O4@C microspheres as templates, zirconium isopropoxide was prehydrolyzed and absorbed onto the microspheres and eventually converted into zirconia by calcinations. The as-prepared Fe3O4@ZrO2 core-shell microspheres were used as affinity probes to selectively concentrate phosphopeptides from tryptic digest of beta-casein, casein, and five protein mixtures to exemplify their selective enrichment ability of phosphopeptides from complex protein samples. In only 0.5 min, phosphopeptides sufficient for characterization by MALDI-MS could be enriched by the Fe3O4@ZrO2 microspheres. The results demonstrate that Fe3O4@ZrO2 microspheres have the excellent selective enrichment capacity for phosphopeptides from complex samples. The performance of the Fe3O4@ZrO2 microspheres was further compared with commercial IMAC beads for the enrichment of peptides originating from tryptic digestion of beta-casein and bovine serum albumin (BSA) with a molar ratio of 1:50, and the results proved a stronger selective ability of Fe3O4@ZrO2 microspheres over IMAC beads. Finally, the Fe3O4@ZrO2 microspheres were successfully utilized for enrichment of phosphopeptides from human blood serum without any other purification procedures.