[Fluoroimmunoassay and Magnetic Lateral Flow Immunoassay for the Detection of Ractopamine].

A fluoroimmunoassay based on quantum dots (QDs) and a lateral flow immunoassay system based on the magnetic beads (MB) were constructed to detect ractopamine (RAG) in urine samples. The monoclonal antibody (Ab1) against RAC was conjugated with QDs or MB as detector reagent, respectively. They apply a competitive format using an immobilized RAC conjugate and free RAC present in samples. That is to say, the concentration of RAC in the sample was negative related to the fluorescense intensity of QDs or the color density of MB. Results showed that the limit of detection (LOD) of fluorescence immunoassay method is 1 ng · mL⁻¹ and analysis time is 4 h, while the visual LOD was 10 ng · mL⁻¹ and analysis time was 15 min in magnetic lateral flow immunoassay system (MFLIS). Taken into consideration of the advantages and disadvantages of the two methods, it was suitable for the trace detection of RAC using fluoroimmunoassay while it was appropriate for point-of-care tesing of RAC by MFLIS.

[Fluorescent and Magnetic Relaxation Switch Immunosensor for the Detecting Foodborne Pathogen Salmonella enterica in Water Samples].

Fluoroimmunoassay based on quantum dots (QDs) and magnetic relaxation switch (MRS) immunoassay based on superparamagnetic nanoparticles (SMN) were constructed to detect Salmonella enterica (S. enterica) in water samples. In fluoroimmunoassay, magnetic beads was conjugated with S. enterica capture antibody (MB-Ab2) to enrich S. enterica from sample solution, then the QDs was conjugated with the S. enterica detection antibody (QDs-Ab1) to detect S. enterica based on sandwich immunoassay format. And the fluorescence intensity is positive related to the bacteria concentration of the sample. Results showed that the limit of detection (LOD) of this method was 102 cfu · mL⁻¹ and analysis time was 2 h. In MRS assay, magnetic nanoparticle-antibody conjugate (MN-Ab1) can switch their dispersed and aggregated state in the presence of the target. This state of change can modulate the spin-spin relaxation time (T2) of the neighboring water molecule. The change in T2(ΔT2) positively correlates with the amount of the target in the sample. Thus, AT can be used as a detection signal in MRS immunosensors. Results showed that LOD of MRS sensor for S. enterica was 10³ cfu · mL⁻¹ and analysis time was 0.5 h. Two methods were compared in terms of advantages and disadvantages in detecting S. enterica.

A boron-containing carbazole dimer: synthesis, photophysical properties and sensing properties.

A novel boron-containing π-conjugated compound has been synthesized by the introduction of electron-acceptors (dimesitylboron groups) at the 3,3'-positions of a carbazole dimer (electron-donor). The compound possesses excellent electrochemical properties and high fluorescence quantum yields. In addition, is a sensitive fluorescence sensor with remarkable colour changes and the results could be confirmed through theoretical calculations of the compounds and [(n)Bu(4)N](+)(2)[·(F)(2)](2-). Our studies indicate that could be used as an excellent optoelectronic material in OLED devices and a ratiometric fluorescent chemosensor.

A novel phenolphthalein-based fluorescent chemosensor for pyrophosphate detection via an Al3+ displacement approach in real samples and living cells.

Herein, we report a new phenolphthalein appended Schiff base (PASB) as reversible fluorescent sensor for the detection of pyrophosphate (PPi) ions through the metal displacement mechanism. PASB showed sensing exclusively toward Al3+ ions in DMF/H2O (v/v = 1/4, pH 5.5) solution, which resulted in a significant fluorescence enhancement at 540 nm. The 1: 2 binding stoichiometry for the complex formation between PASB and Al3+ was confirmed by Job's plot and mass spectroscopic studies. Moreover, a solution of the in situ formed PASB-Al3+ complex displayed a high selectivity to PPi. The addition of PPi to PASB-Al3+ ensemble significantly quenched its fluorescence. Thus, a dual response was established based on "Off-On-Off" strategy for detection of both Al3+ and PPi. The detection limit is 5.86 nM and 26 nM for Al3+ and PPi, respectively. On this basis, we use PASB to detect Al3+ in food samples. Furthermore, PASB was successfully applicable to detect Al3+ and PPi for intracellular imaging in Human liver cancer cells.

Dicyanoisophorone-based fluorescent probe with large Stokes shift for ratiometric detection and imaging of exogenous/endogenous hypochlorite in cell and zebrafish.

A novel dicyanoisophorone-based red-emissive fluorescence probe (YT) with large Stokes shift (230 nm) was synthesized for rapid (<20 s) and selective detection of hypochlorite ions in nearly 100% aqueous medium. YT responded to hypochlorite ions via the ClO--promoted oxidative deprotection of thioacetal, leading to a red shift in its fluorescence maximum from 590 nm to 640 nm accompanied by naked-eye color change from orange to red. The emission response of the probe toward ClO- presented a good linear relationship in the 5-160 µM concentration range, with the LOD of 4.64 µM. Further, the probe YT was successfully employed in exogenous and LPS-induced endogenous imaging of ClO- in live cells and zebrafish, demonstrating its potential applications in biological science.

A phenazine-imidazole based ratiometric fluorescent probe for Cd2+ ions and its application in in vivo imaging.

A new phenazine-imidazole based Schiff base (PIS) fluorescent probe has been developed for the ratiometric detection of Cd2+ ions in aqueous media at physiological pH. PIS upon binding with Cd2+ ions shows red shifted fluorescence and thereby, permits ratiometric detection of Cd(II) ions. A detection limit down to 2.10 × 10-8 M was determined for Cd2+ quantitation. Also, the accompanying apparent fluorescence color change (from yellow to orange red) is noticeable to the naked eye under a UV-lamp. The sensing mechanism could be attributed to the 1 : 1 PIS-Cd complexation, followed by extension of the conjugation due to better planarity and modulation of the charge transfer efficiency in the probe. This was complemented by solvatochromism and density functional theory calculations. Furthermore, PIS was used to detect Cd2+ in Oxya chinensis cells, zebrafish larvae and live tissues of Arabidopsis thaliana under a fluorescence microscope, showing great potential in analyzing living biosystems.

A phenolphthalein-based fluorescent probe for the sequential sensing of Al3+ and F- ions in aqueous medium and live cells.

A novel fluorescent probe, phenolphthalein­dialdehyde­(2­pyridyl) hydrazone (L), for sequentially detecting Al3+ and F- in almost 100% aqueous medium was successfully designed and synthesized. The probe offers two binding pockets for Al3+ to form a 1: 2 ligand/metal complex, leading to a significant fluorescence enhancement at 465 nm. Further, the in-situ formed L-Al complex acts as a secondary fluorescent chemosensor for F- by quenching the fluorescence of the complex with high selectivity. The detection limit for Al3+ and F- sensing is 2.28 nM and 0.13 µM, respectively, which are far below the World Health Organization (WHO) acceptable limits (7.41 µM for Al3+ ion and 79 µM for F-) in drinking water. The probe L was successfully applied to the detection of Al3+ and F- in cells using fluorescence microscopy.

A Label-free aptasensor based on Aptamer/NH2 Janus particles for ultrasensitive electrochemical detection of Ochratoxin A.

In this study, a novel aptasensor based on Aptamer/NH2 Janus particles is developed for the detection of Ochratoxin A(OTA). By coating gold on the hemispherical surface of the aminated polystyrene particles, Ochratoxin A aptamer is immobilized on the surface of the gold layer for selective identification and the other hemispherical able to bind to Glassy carbon electrode via peptide bond. Under optimum conditions, the sensor exhibited a wide dynamic range of OTA concentration from 1 × 10-5 nM to 10 nM, and the detection limit is 3.3 × 10-3 pM on condition of acceptable stability and reproducibility. The sensors were showed excellent performance in the detection of OTA in red wine sample with recoveries between 95.7% and 100.18%, which studied by the standard addition spiking technique. This work provides a new idea and method for preparing immune electrochemical sensors and is expected to be used for the OTA detection in red wine sample analysis.

A lysosome-targetable fluorescent probe for real-time imaging cysteine under oxidative stress in living cells.

As an effective lysosomal biomarker for oxidative stress status, cysteine (Cys) plays an important role in lysosomal proteolysis. Herein, we report the first lysosome-targetable fluorescence probe (MCAB) for Cys-selective detection based on nucleophilic addition reaction of sulfhydryl toward a α, ß-unsaturated ketone and demonstrate its application to lysosomal-targetable imaging. MCAB is designed based on a α, ß-unsaturated ethanoylcarbazole as the fluorophore and the thiols reaction site, and a methylcarbitol unit as a lysosome-targetable group. Upon reacting with Cys, this probe turns on highly specific fluorescence signals linearly proportional to Cys concentrations over the range of 0-300 µM. MCAB detects Cys with a rapid response time (within 12 min) and low limit of detection (0.38 µM). MCAB is highly selective to Cys over other similar biothiols including homocysteine (Hcy) and glutathione (GSH). Moreover, it also exhibits significant lysosomal-targetable ability, which is ideal for lysosomal Cys-selective imaging. Using MCAB, we have successfully visualized the fluctuation endogenous Cys in lysosomes under oxidative stress status in real-time.

Study on the supramolecular system of 5-(p-hydroxyphenyl)-10,15,20-tris-(4-chlorophenyl)porphyrin with cyclodextrins and its analytical characteristics.

The supramolecular systems of 5-(p-hydroxyphenyl)-10,15,20-tris-(4-chlorophenyl)porphyrin (p-HTClPP) with beta-cyclodextrin (beta-CD), heptakis(2,3,6-tri-O-methyl)-beta-CD (TM-beta-CD), carboxymethyl-beta-cyclodextrin (CM-beta-CD) and sulfurbutylether-beta-CD (SBE-beta-CD) have been investigated by means of absorption, fluorescence and (1)H NMR spectroscopy. The formation of inclusion complexes has been confirmed on the base of changes of spectroscopy properties. "The double reciprocal method" has been used to determine the stoichiometry and the inclusion constants of p-HTClPP with the four cyclodextrins (CDs). The results show that p-HTClPP can form 1:1 inclusion complexes with the four CDs. Compared with parent native beta-CD, the inclusion abilities of modified beta-CDs with p-HTClPP are stronger. It indicates that the hydrophobic effect plays an important role in the inclusion procedure. The mechanism of inclusion interaction was examined by (1)H NMR technique. During the study of p-HTClPP-TM-beta-CD supramolecular complex, an efficient enhancement of fluorescence intensity was observed. Based on this phenomenon, fluorometric method for the determination of p-HTClPP was developed. The relationship between fluorescence intensity and the concentration of p-HTClPP is linear from 1.0 x 10(-9) to 7.0 x 10(-6)mol L(-1). The limit of detection is 8.3 x 10(-10)mol L(-1) and the relative standard deviation (R.S.D.) is 1.3% (n=8). This research will provide useful information for further application of p-HTClPP.

[Study on the interaction between ICT fluorescence probe and bovine serum albumins].

The present article studied the interaction between intramolecular charge transfer fluorescence probe-1-keto-2-(p-dimethylaminobenzal)-tetrohydronaphthalene (KDTN) and bovine serum albumins (BSA). With the concentration of KDTN increasing, the fluorescence of BSA rapidly quenched and the fluorescence peak gradually blue-shifted. The result indicated that they were bound mainly by hydrophobic interaction. The binding sites is 0.94 (3 degrees C) and the equilibrium constant K is 3.27 x 10(4) L x mol(-1). Temperature increment is advantageous to the combination. It is a single static quenching process that the fluorescence of BSA quenches, which is induced by the combination of KDTN and BSA. Further study showed that different substances had different effects on the combination of KDTN and BSA.

Facile preparation of bright orange fluorescent carbon dots and the constructed biosensing platform for the detection of pH in living cells.

Long wavelength (i.e., orange- to red-light) fluorescence emission and functionality are critically longing for the development and applications of carbon dots (CDs) toward biosensor and bioimaging analysis. Herein, the N-doped carbon dots (N-CDs) with bright orange fluorescence emission at 592 nm were facilely synthesized via p-phenylenediamine as a carbon precursor by microwave method. The as-prepared N-CDs exhibit favorable biocompatibility, excellent water solubility, highly optical stabilities and display sensitive pH response behavior. When the pH is decreased from 9.45 to 2.45, its emission fluorescence intensity will be significantly enhanced. The pKa value of N-CDs is 5.80 and it shows linear response to the physiological pH range of 4.45-7.00. Moreover, the N-CDs also possess high selectivity of hydrogen ions over common metal ions and some bioactive molecules, excellent photostability and reversibility. Based on this, a fluorescence sensing platform is established for the detection of pH in the environment. The N-CDs were successfully used as the fluorescent probe for cellular imaging and applied to monitor pH fluctuations in live cells based on its biocompatibility and cell membrane permeability. It is also anticipated that the N-CDs are potential bio-nano-materials for real-time tracking of the intracellular pH especially under physiological conditions in the disease diagnosis, biosensing and biomedical fields.

Interaction of human serum albumin with bendroflumethiazide studied by fluorescence spectroscopy.

The interactions between bendroflumethiazide (BFTZ) and human serum albumin (HSA) have been studied by fluorescence spectroscopy. Binding constants for drug attachment to the various binding sites of HSA have been measured at different temperatures in physiological buffer solution. The effect of metal ions on BFTZ interaction with HSA was also investigated. The thermodynamic parameters, DeltaH and DeltaS, have been calculated to be 49.28kJmol(-1)>0, and 258.83Jmol(-1)K(-1)>0, respectively. The distance between HSA and BFTZ, r, was determined to be 1.47nm based on Förster's non-radiative energy transfer theory. The experimental results reveal that BFTZ has a strong ability to quench the intrinsic fluorescence of HSA through a static quenching mechanism. Furthermore, the binding constants between BFTZ and HSA are remarkably independent of temperature, and decrease in the presence of various ions, usually by about 30-55%. Hydrophobic interaction occurs between BFTZ and the sub-domain II A of HSA.

Study for luminescence performance of three methyl xanthine derivatives.

In this paper, the low-temperature phosphorescence (LTP), the low-temperature fluorescence (LTF), the paper substrate room-temperature phosphorescence (PS-RTP) and the room fluorescence (RTF) properties of caffeine (CF), theophylline (TP), and theobromine (TB) are investigated and compared, and some rules are found out: their maximal excitation wavelength and emission wavelength are in the range of 270-300 nm and 395-445 nm, respectively. And the PS-RTP characters of lifetime, polarization and quanta yield are also investigated and compared. It is found that their lifetimes of PS-RTP are all in the level of 0.1s. They belong to long-life phosphorescence and their PS-RTP spectra are incompletely polarized.

Down-regulation of aquaporin3 expression by lipopolysaccharide via p38/c-Jun N-terminal kinase signalling pathway in HT-29 human colon epithelial cells.

AIM: To investigate the influence of lipopolysaccharide (LPS) through the p38/c-Jun N-terminal kinase (JNK) signalling pathway on aquaporin 3 (AQP3) expression in HT-29 human colon epithelial cells. METHODS: HT-29 cells were treated with LPS, and then the membrane localisation of AQP3 was examined by immunofluorescence staining. The mRNA and protein expression of AQP3 with LPS exposure was measured by real-time reverse transcription-PCR and Western blot, respectively. Activation of p38 and JNK was evaluated by detection of phosphorylation of p38 and JNK using Western blot assay. AQP3 protein expression was determined by Western blot in cells after treatment with SB203580, a selective p38 MAPK inhibitor, or SP600125, a selective JNK inhibitor. RESULTS: In HT-29 cells, the transcription and protein expression of AQP3 were decreased by LPS in a dose- and time-dependent manner, the expression of AQP3 was significantly decreased with the increased concentration of LPS, and at a dose of 100 µg/mL LPS, AQP3 mRNA and protein levels were decreased by a maximum (P < 0.05) of 1.51-fold and 1.49-fold, respectively. When cells were treated with 100 µg/mL LPS for 0, 3, 6, 12, and 24 h, the AQP3 mRNA level was significantly decreased at an early time point of 3 h, and reached about 10% of the control level at 24 h post-treatment (P < 0.05). Down-regulation of AQP3 expression was significantly inhibited by the p38 inhibitor (SB203580) and JNK inhibitor (SP600125). CONCLUSION: p38 and JNK may be promising targets for the preservation of AQP3 expression and may be beneficial to the clinical management of diarrhoea.

Studies on the spectroscopic behavior of cryptotanshinone, tanshinone IIA, and tanshinone I.

A comparative study on the spectroscopic behavior of cryptotanshinone (CTan), tanshinone IIA (Tan IIA), and tanshinone I (Tan I) has been investigated, including UV-Vis absorption, low temperature phosphorescence (LTP), low temperature fluorescence (LTF), paper substrate-room temperature phosphorescence (PS-RTP), paper substrate-room temperature fluorescence (PS-RTF) and fluorescence in liquid (LF). The effect of pH on the luminescence intensity is discussed. Lifetime and polarization of the LTP and RTP have been examined with phosphorescence lifetime in the range of 0.6-0.9s and polarization in the range of 0.10-0.27. Analytical characteristics of LF, PS-RTF and PS-RTP of CTan, Tan IIA, and Tan I have been studied.

Study on inclusion complex of cyclodextrin with methyl xanthine derivatives by fluorimetry.

The inclusion complexes of beta-cyclodextrin (beta-CD) and HP-beta-cyclodextrin (HP-beta-CD) with caffeine, theophylline and theobromine were investigated by fluorimetry. Various factors affecting the formation of inclusion complexes were discussed in detail including forming time, pH effect and temperature. The results indicate that inclusion process was affected seriously by laying time and pH. The forming time of beta-CD inclusion complexes is much longer than that of HP-beta-CD. The optimum pH range is about 7-12 for caffeine, 8-10 for TP, 10.5-12 for TB. The intensities of their fluorescence increase with the decreasing of temperature. Their maximum excitation wavelengths are all in the range of 280-290 nm. The emission wavelength of caffeine and theophylline are both in the range of 340-360 nm, and that of theobromine is about 325 nm. The fluorescence signals are intensified with the increasing concentration of CD. The stoichiometry of the inclusion complexes of CD with these three methyl xanthine derivatives are all 1:1 and the formation constant are all calculated.

[Study of the effect of aliphatic alcohol on the inclusion interaction of qucertin-cyclodextrin complexes].

The inclusion interaction of qucertin with beta-cyclodextrin and its derivatives was studied by fluorimetry. The formation constants of their inclusion complexes were determined, and the stoichiometry ratio on the formation of the inclusion complexes was determined by equimolar continueous variation method. The effect of aliphatic alcohols with short chains on inclusion process was discussed. The results showed that the inclusion ability of beta-CD and its derivatives was in the order: HP-beta-CD>M-beta-CD>beta-CD. The effect of aliphatic alcohol on complexes was related to the carbon number and volume of alcohol molecules.

Study on the inclusion behavior of p-sulfonatocalix[6]arene with propranolol by spectrofluorometry.

The inclusion interaction between propranolol (PPL) and p-sulfonatocalix[6]arene (SCX6) was investigated by fluorescence and (1)H NMR spectroscopy. Influences of pH, temperature, ionic strength and the concentration of SCX6 were examined in detail. In phosphate buffer solution with pH 7.5, the fluorescence of PPL dramatically quenched upon addition of SCX6 revealing the formation of inclusion complexes between PPL and SCX6. The stoichiometric ratio was verified to be 1:1 by the continuous variation method. The inclusion constant of PPL-SCX6 complexes was calculated as 2.2×10(4)L/mol by the nonlinear curve fitting method. (1)H NMR titration spectra testified that the aliphatic chain of PPL may be partially penetrated into the hydrophobic cavity of SCX6. This was confirmed by molecular dynamics calculations.

Novel far-visible and near-infrared pH probes based on styrylcyanine for imaging intracellular pH in live cells.

Two novel vis-NIR pH probes based on styrylcyanine with acidic pH response are easily synthesized, which display large Stokes shift and high sensitivity. The significant colocalizations of two probes with LysoTracker Green DND-26 are achieved in C6 cells, suggesting potential application for imaging acidic organelles in live cells.