A Miniaturized and Highly Sensitive "Windmill" Three-Channel Fluorescence Detector for Simultaneous Detection of Various Mycotoxins.

A novel "windmill" three-channel light-emitting diode induced fluorescence detector (LED-IF) was proposed to maximize the excitation efficiency and fluorescence collection efficiency. Compared with the typical collinear arrangement, the fluorescence intensity of the three channels was increased by 7.85, 3.88, and 2.94 times, respectively. The compact shaping optical path was designed to obtain higher excitation efficiency and a lower background stray light effect caused by high divergence angle high-power ultraviolet (UV)-LEDs simultaneously, which increased the sensitivity of three channels by 4.6 to 5.7 times. It was found that using a photodiode (PD) with a flat window and a larger photosensitive surface can collect the Lambertian emission fluorescence in the flow cell more efficiently, increasing the signal-to-noise ratio of each channel 1.3 to 1.8 times. The limits of detection (LODs, 3 times peak-peak noise) of aflatoxin B2 (AFB2), ochratoxin (OTA), and zearalenone (ZEN) were 0.33, 1.80, and 28.2 ng/L, respectively. Finally, six mycotoxins were analyzed simultaneously by the detector coupling with HPLC. The results showed that the sensitivity of the detector was at the best level to date, which was better than that of the top commercial fluorescence detectors (FLDs). The developed detector has the advantages of having small volume, low cost, and long lifetime and being robust, which has wide application and market prospects.

A Simple and Low-cost Preliminary Quantification of Target Membrane Protein in Single Cells.

To study the heterogeneity of target membrane proteins in single cells with cellular integrity, we proposed a simple and low-cost method to obtain the copy number of the membrane proteins. HeLa cells were labeled by FITC affinity bodies specifically targeting HER2 membrane proteins. The immunolabeled HeLa cells were quantified by a laboratory-built laser induced fluorescence detector. A series of fluorescent microspheres with known number of FITC molecules on the surface were used to establish the calibration curve, instead of the standard fluorescent solutions, because the morphology of the microspheres was similar to the cells, and the distribution of FITC on the spheres were similar to the distribution of HER2 on the HeLa. The fluorescence intensity of the cells was converted to the molecule number of HER2 by the calibration curve. A capillary electrophoresis system was used to drive the microspheres and cells through the detection window. The copy number of HER2 in HeLa cells ranged from 4,036 to 1,224,920 ± 100 (2.5-97.5%), and the median of copy numbers were 104,438 ± 100 per cell. This method for measuring low-abundance membrane proteins can be utilized for the initial exploration of proteomics in ordinary laboratories.

Online micro solid phase extraction coupled with ultra-performance liquid chromatography-tandem mass spectrometry for trace analysis of endogenous plant hormones in Ulva linza.

OBJECTIVE: Ulva linza (L.) is a species of green algae widely distributed in China. We aimed to establish a sensitive online analytical method for quantification of endogenous phytohormones in fresh minute seaweed samples. METHOD: The method for quantification of endogenous plant hormones in fresh minute samples was developed based on a homemade online micro solid phase extraction (m-SPE) system coupled with an ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) platform. The online m-SPE instrument injected the eluent of m-SPE directly onto the LC separation column, improving the utilization of samples and saving time. The m-SPE column, of which the effective size was 9.6 × 2 mm i.d., was filled with 19 mg of C18 (10 µm). RESULTS: Under optimized conditions, the limits of detection were 0.002-0.060 ng ml-1 for five plant hormones. The actual sample recoveries of phytohormones were 76.4-103.4% and the coefficients of variance were below 14.1%. The temporal distribution of these endogenous plant hormones of U. linza during different growth periods is described. CONCLUSION: The proposed online m-SPE method was successfully applied to quantification of endogenous acidic and alkaline plant hormones in U. linza. It provides important information for the further study of the physiological and ecological effects of plant hormones in lower algal species.

Enhancement of Chemiluminescence Intensity of S2* in Non-premixed Hydrogen Microjet Flame in the Photometric Detector for Sulfur Detection.

A transparent quartz rod (q) placed vertically on top of a non-premixed hydrogen microjet flame in a flame photometric detector (qFPD) was developed and evaluated for sulfur detection. The microjet flame burned around the quartz rod because of Coanda effect, forming an extended downstream flame zone with a relatively low temperature between 550 and 650 °C, which is favorable to the formation of S2*. The emission intensity of S2* and the signal-to-noise ratio (SNR) of sulfur response were enhanced 2.6- and 2.1-fold, respectively. It was found that the quartz rod of diameter 4 mm with a tip shape of semicircle placed 6 mm above the nozzle yielded the highest SNR. The limits of detection (LOD) for seven kinds of tested sulfur-containing compounds of qFPD were 0.3-0.5 pg S s-1, which is 5-7 times better than that of commercially available FPD detectors (LOD: 1.6-2.8 pg S s-1). The selectivity of sulfur over carbon was 105 on qFPD when the SNR for the mass flow rate of S and C atoms was ∼3 times. It was the first time that a quartz rod was used vertically on top of a microjet hydrogen-rich flame in FPD to enhance the chemiluminescence of S2* and improve the LOD down to 0.3-0.5 pg S s-1.

Spherical Dichroic Reflector Improves Limit of Detection in Laser-Induced Fluorescence Detection.

A miniature laser-induced fluorescence (mLIF) detector utilizing a novel spherical dichroic reflector (SDR), an unconventional long working distance high magnification objective, an uncommon broadband emission-matched excitation filter pair, and a silicon-based photodiode detector assembly instead of a photomultiplier tube was developed and evaluated. The detection cell was placed at the spherical center of the SDR instead of the regular focus, yielding a 1.8× signal-to-noise ratio (SNR) improvement. Different from previous works, the use of a 40× objective with a long working distance of 5.38 mm and a broadband BP 527-70 nm emission filter with matched BP 450-30 nm excitation filter improved SNR to 4.6× and 1.9×, respectively. By flow injection analysis (FIA) evaluation, the limit of detection (LOD; 3σ method) for fluorescein sodium was 1.5 × 10-13 M or 8.9 fluorescein molecules in 98 pL detection volume, which was the lowest level of LIFs evaluated by FIA mode. The analysis of three kinds of amino acids with LODs at sub pM to fM level (the lowest levels, hundreds of times lower than previous works using normal capillary) demonstrated the potential of the mLIF in ultratrace analysis of biological and environmental samples, including low copy molecules in a single cell.

Cationic metal-organic frameworks as an efficient adsorbent for the removal of 2,4-dichlorophenoxyacetic acid from aqueous solutions.

Metal-organic frameworks (MOFs) material with high surface area, good chemical stability and multi-functionality, has become an emerging adsorbent for water treatment. A novel kind of quaternary amine anionic-exchange MOFs UiO-66 namely UiO-66-NMe3+ was firstly synthesized for adsorptive removal of a widely used toxic herbicide 2,4-dichlorophenoxyacetic acid (2,4-D) from aqueous solutions. The well-prepared UiO-66-NMe3+ MOFs were fully characterized, and then the main parameters affecting the adsorption process including solution pH, adsorbent dosage and coexisting anions were systematically investigated. The maximum adsorption capacity of UiO-66-NMe3+ toward 2,4-D reached as high as 279 mg g-1, much higher than that of pristine UiO-66 and aminated UiO-66. The adsorption mechanism could be attributed to the electrostatic interactions efficiently enhanced by the functionalization of quaternary amine groups, combining with the π-π conjugations between the linkers in MOFs and 2,4-D molecules, leading to the better adsorption performance of UiO-66-NMe3+. Additionally, the UiO-66-NMe3+ could be well regenerated by simple solvent washing and exhibited a slight decline of adsorption capacity after seven successive recycle. Furthermore, satisfactory adsorption capacity and reusability of the MOFs in environmental water samples were attained. Comparing with reported activated carbon and resin materials, the UiO-66-NMe3+ MOFs possessed higher adsorption capacity and shorter equilibrium time, as well as good reusability and practicality. The developed ion-exchange functionalized MOFs provided an ideal alternative for efficient adsorptive-removal of 2,4-D from complicated aqueous environment.

Quantification of Low Copy Number Proteins in Single Cells.

We have developed an ultrasensitive and highly selective method to quantify low copy number intracellular proteins in a single cell using a low-cost laser-induced fluorescence (LIF) detector and a BV605 fluorescent probe. Active caspase3 proteins in cells were labeled by corresponding antibody-BV605 fluorescent binding, and a cell was injected into a 20 cm × 50 µm i.d. capillary column, followed by in situ lysis and capillary electrophoresis (CE)-LIF analysis. About seven active caspase3 protein molecules in a detection volume of 91 pL could be detected. In our method, cross-bounding proteins other than active caspase3 could be separated and distinguished by differences of retention time. By using Si photodiode assembly as a fluorescent detector instead of PMT, the dynamic range of the LIF is over 4 orders of magnitude. In this experiment, we found that the number of active caspase3 molecules in 98 single Jurkat cells were from 629 to 12171, reflecting significant heterogeneity among the cells although they were from the same batch. For extended application, it could also be applied to quantify other types of low copy number proteins in a single cell as long as the corresponding antibodies are provided. This high-sensitive method could also be a promising tool for earlier cancer diagnosis and related disease pathway research which is relevant to low copy number proteins. In addition, this low-cost system could also be easily expanded to an array system for high-throughput quantitation of low copy proteins in single cells.

Spatial Profiling of Gibberellins in a Single Leaf Based on Microscale Matrix Solid-Phase Dispersion and Precolumn Derivatization Coupled with Ultraperformance Liquid Chromatography-Tandem Mass Spectrometry.

A spatial-resolved analysis method for profiling of gibberellins (GAs) in a single leaf was developed on the basis of microscale sample preparation and precolumn derivatization coupled with ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The proposed microscale sample preparation was based on modified matrix solid-phase dispersion (MSPD) method, in which the plant sample (<1 mg) and C18 sorbent were ground together in one microcentrifuge tube, and then extraction solvent was added followed by centrifugation. In this protocol, the grinding, extraction, and purification were performed in one microcentrifuge tube without any sample transfer step, resulting in an obvious decrease in sample loss. Moreover, a new derivatization reagent, 3-bromopropyltrimethylammonium bromide (BPTAB), was used to further enhance the signal intensities of GAs on MS by 3-4 orders of magnitude, which was much higher than the reported derivatization reagents for GAs such as bromocholine bromide and 3-bromoactonyltrimethylammonium bromide. The present method showed high sensitivity (minimum detectable amount (MDA) of 10.1-72.3 amol for eight GAs) and low sample consumption (down to 0.30 mg FW). Under the optimized conditions, the distribution of GA19 in a single Arabidopsis thaliana leaf was profiled with a spatial resolution of 2 × 2 mm2.

Sheathless interface to match flow rate of capillary electrophoresis with electrospray mass spectrometry using regular-sized capillary.

RATIONALE: The flow rate match has been a great challenge when coupling capillary electrophoresis (CE) with electrospray ionization mass spectrometry (ESI-MS). Conventional CE-ESI-MS interfaces used liquid sheath flow, narrowed capillary or additional pressure to meet this requirement; sacrifice of either capillary inner diameter (i.d.) or separation efficiency is often inevitable. Thus, a regular-sized capillary-based sheathless interface would be attractive for flow rate match in CE-MS. METHODS: The regular-sized capillary-based CE-MS interface was achieved by coupling CE with induced electrospray ionization (iESI) which was stimulated by the fact that the iESI could both achieve flow rate down to 0.2 µL/min and retain ionization efficiency. The CE-iESI-MS interface was completed with an intact separation capillary, outside the outlet end of which a metal electrode was attached for the application of alternating current (ac) high voltage (HV). RESULTS: The feasibility of this CE-iESI-MS interface was demonstrated through the stable total ion chromatograms obtained by continuous CE infusion of tripropylamine with regular-sized capillaries. Tripropylamine and atenolol were separated and detected successfully in phosphate buffer solution (PBS) by CE-iESI-MS using a 50 or 75 µm i.d. capillary. Furthermore, this new interface showed a better signal-to-noise (S/N) of 3 to 7 times enhancement compared with another sheathless CE-ESI-MS interface that using one high voltage for both separation and electrospray when analyzing the mixture of tripropylamine and proline in NH4 OAc buffer. In addition, the reproducibility of this interface gave satisfactory results with relative standard deviation (RSD) in retention time in the range between 1% and 3%. CONCLUSIONS: The novel sheathless CE-MS interface introduced here could match conventional electroosmotic flow (EOF) with electrospray which could also preserve the separation efficiency and sensitivity of CE-MS. This newly developed CE-iESI-MS interface was also demonstrated to be effective for different buffers, PBS and NH4 OAc, without any additives such as methanol and acetic acid. Hence, we believe that this sheathless CE-MS interface could be operated with other nonvolatile and volatile buffers. Copyright © 2016 John Wiley & Sons, Ltd.

Further investigation of a peptide extraction method with mesoporous silica using high-performance liquid chromatography coupled with tandem mass spectrometry.

Mobil Composition of Matter No. 41 (MCM-41) was the most frequently used mesoporous silica material to extract peptides from complex biological samples. However, there were confusing extraction conditions and large extraction efficiency variance among related reports, which resulted from unclear understanding about the interaction between the material and peptides. In this study, the extraction mechanism was investigated with one set of tryptic peptides by using high-performance liquid chromatography coupled with triple quadrupole mass spectrometry. Generally, hydrophobic interaction and electrostatic attraction were two major driving forces for extraction of peptides, while electrostatic repulsion greatly weakened the interaction between the material and peptides with isoelectric points below the pH. With most peptides positively charged and MCM-41 slightly negatively charged, most efficient extraction was obtained at pH 3, and it was proved that electrostatic and hydrophobic interaction acted in synergy for extraction of all the peptides. A mixed solution of acetonitrile with buffers of high pH or ion strength was demonstrated to be favorable for elution, which performed much better than the commonly used eluate (mixture of acetonitrile with 0.1% trifluoroacetic acid). Finally, under optimum conditions, it was found that extraction efficiency of MCM-41 for protein digest and human serum was greatly improved.

Functionalization of Carbonaceous Nanodots from Mn(II) -Coordinating Functional Knots.

Transition metal (TM)-induced functionalization has been expected to be powerful for the modification of carbonaceous nanodots (CDs) for many years. Although some attempts have been made, relevant research evolutions are at a standstill. In this work, a coordination-carbonization protocol to gain Mn(II) -functionalized CDs (MCDs) was innovatively employed, and the modification of CDs based on metal-coordinating functional knots was realized. By investigating the structural and photophysical properties, coordinating conformation of functional knots was confirmed and considered to be correlated with excitation-independent photoluminescence as well as large Stokes shift of MCDs. Moreover, potential applications of MCDs have been extended from common bioimaging and fluorescent inks to new areas, such as chemosensors for volatile organic compounds and anticorrosion fluorescent films.

Quantitative evaluation of peptide-extraction methods by HPLC-triple-quad MS-MS.

In this study, the efficiency of five peptide-extraction methods­acetonitrile (ACN) precipitation, ultrafiltration, C18 solid-phase extraction (SPE), dispersed SPE with mesoporous carbon CMK-3, and mesoporous silica MCM-41­was quantitatively investigated. With 28 tryptic peptides as target analytes, these methods were evaluated on the basis of recovery and reproducibility by using high-performance liquid chromatography-triple-quad tandem mass spectrometry in selected-reaction-monitoring mode. Because of the distinct extraction mechanisms of the methods, their preferences for extracting peptides of different properties were revealed to be quite different, usually depending on the pI values or hydrophobicity of peptides. When target peptides were spiked in bovine serum albumin (BSA) solution, the extraction efficiency of all the methods except ACN precipitation changed significantly. The binding of BSA with target peptides and nonspecific adsorption on adsorbents were believed to be the ways through which BSA affected the extraction behavior. When spiked in plasma, the performance of all five methods deteriorated substantially, with the number of peptides having recoveries exceeding 70% being 15 for ACN precipitation, and none for the other methods. Finally, the methods were evaluated in terms of the number of identified peptides for extraction of endogenous plasma peptides. Only ultrafiltration and CMK-3 dispersed SPE performed differently from the quantitative results with target peptides, and the wider distribution of the properties of endogenous peptides was believed to be the main reason.

Hybrid titania-zirconia nanoparticles coated adsorbent for highly selective capture of nucleosides from human urine in physiological condition.

Modified nucleosides are important biomarkers of cancers. For their analysis, boronate adsorbents were widely used to selectively capture them from urine, but often suffered from serious secondary hydrophobic interaction and harsh alkaline extraction condition. In this work, the hybrid titania-zirconia nanoparticles coated on porous silica spheres (TiO2-ZrO2/SiO2) were developed for the first time as a selective adsorbent for nucleosides under neutral conditions based on specific recognition of its Lewis acid sites to the cis-diol group. It was found here that TiO2-ZrO2 has higher binding constants than pure TiO2 or ZrO2, and a significant improvement of binding efficiencies was obtained by decreasing calcination temperature to 400 °C. Moreover, physiological pH of urine (pH 6-7) was found optimal to adsorb nucleosides and resist other Lewis base interferences. By self-assembly of TiO2-ZrO2 nanoparticles on silica, unprecedentedly high binding capacity (35 mg/g) for nucleosides was obtained due to high surface area (350 m(2)/g) and abundant Lewis acid sites on the surface. Due to efficient reduction of secondary hydrophobic interaction on the inorganic surface, cis-diol nucleosides could be captured from 500-fold non-cis-diol interferences. In the real sample application, nine nucleosides have been quantified with relative recoveries in 83%-126%, and 42 ribosylated metabolites had been identified with only 100 µL of urine at physiological pH. Among them, two nucleosides have never been identified in most previous studies using boronate adsorbents for capture.

Postage stamp-sized array sensor for the sensitive screening test of heavy-metal ions.

The sensitive determination of heavy-metal ions has been widely investigated in recent years due to their threat to the environment and to human health. Among various analytical detection techniques, inexpensive colorimetric testing papers/strips play a very important role. The limitation, however, is also clear: the sensitivity is usually low and the selectivity is poor. In this work, we have developed a postage stamp-sized array sensor composed of nine commercially available heterocyclic azo indicators. Combining filtration-based enrichment with an array of technologies-based pattern-recognition, we have obtained the discrimination capability for seven heavy-metal ions (Hg(2+), Pb(2+), Ag(+), Ni(2+), Cu(2+), Zn(2+), and Co(2+)) at their Chinese wastewater discharge standard concentrations. The allowable detection level of Hg(2+) was down to 0.05 mg L(-1). The heavy-metal ions screening test was readily achieved using a standard chemometric approach. And the array sensor applied well in real water samples.

An SPE-assisted BODIPY fluorometric paper sensor for the highly selective and sensitive determination of Cd²âº in complex sample: rice.

By using sensing technology, the individual component analysis at trace level in complex samples remains problematic simply because of various interfering species. For example, the determination of Cd(2+) in rice is difficult due to the co-existing interfering metal cations at thousands or even millions of times higher concentrations. In this study, a heavy-metal ion sensitive BODIPY (4,4-difluoro-4-bora-3a,4a-diaza-s-indacene)-based fluorometric paper sensor with assistance of solid phase extraction (SPE) was developed for the highly selective and sensitive determination of trace Cd(2+) in rice. SPE column packed with prepared sulfonated PS-DVB microspheres was employed to enrich trace Cd(2+) and meanwhile remove most interfering heavy-metal ions in simulated complex rice sample with oxalic acid as eluent, which was theoretically selected on the basis of f values. Mn(2+), as a major coexistent heavy-metal ion, could not be easily removed by SPE, but showed little fluorescent response for BODIPY fluorometric paper sensor even in excess amounts. Combining the separation and enrichment capability of SPE column with the selectivity of BODIPY-based fluorometric paper sensor, we were able to detect trace Cd(2+) in complex samples. The response of fluorometric paper sensor was linearly related with Cd(2+) concentrations in the range of 0.5-4 µM, with a detection limit of 0.5 µM. Twelve real rice samples spiked with Cd(2+) were analysed using this method and the results are in good agreement with ICP-MS measurements.

Clinical observation of Otitis Media Secretory during Covid-19.

OBJECTIVE: This study aims to analyze the onset of otitis media secretory, the peak period of infection with the Omicron strain of SARS-CoV-2 virus, and the time of transmigration during a pandemic of the Omicron strain. Additionally, the study aims to investigate to study the presence of SARS-CoV-2 virus in the middle ear cavity of patients with otitis media secretory and the survival time through a new method for detecting SARS-CoV-2 virus antigen in middle ear effusion. METHODS: Retrospective comparison of the incidence of otitis media secretory during infection with SARS-CoV-2 virus Omicron strain from December 15, 2022, to January 15, 2023, versus the noninfection period from December 15, 2021, to January 15, 2022. We used a questionnaire star application to investigate the demographic and epidemiological characteristics of the 40 patients with otitis media secretory who participated in this study were investigated. A novel coronavirus (2019-nCoV) antigen detection kit (colloidal gold method) was used to detect middle ear effusion in patients with otitis media secretory. The data were statistically analyzed using SPSS 29.0 software. The measurement data are expressed as x ± s , the count data are expressed as the number of cases (%), and the data were compared using the χ 2 test. p < 0.05 indicated a statistically significant difference. RESULTS: During the SARS-CoV-2 virus Omicron strain pandemic, the incidence of otitis media secretory increased by 15% compared with the noninfection period. The peak infection period for the SARS-CoV-2 virus Omicron strain was December 25, 2022, and December 15, 2022, and the peak time of conversion was 7 to 9 days after the infection. Middle ear effusion SARS-CoV-2 virus antigen testing was performed in patients with otitis media secretory after conversion; 5 patients (12%) were positive, and 35 patients (88%) were negative. The disease duration in patients with negative results was more than 3 weeks. CONCLUSIONS: Otitis media secretory is one of the most common ear complications after infection with the Omicron strain of SARS-CoV-2 virus, and the significantly higher incidence is associated with middle ear viral infection. Middle ear effusion SARS-CoV-2 virus antigen test detected the virus, which survived longer in the middle ear effusion than in the nasal cavity. The middle ear effusion test can detect SARS-CoV-2 virus antigen and determine whether the organism contains virus residue.

In vivo fast equilibrium microextraction by stable and biocompatible nanofiber membrane sandwiched in microfluidic device.

In vivo analysis poses higher requirements about the biocompatibility, selectivity and speed of analytical method. In this study, an in vivo fast equilibrium microextraction method was developed with a biocompatible core-sheath electrospun nanofiber membrane sandwiched within a microfluidic unit. The polystyrene/collagen core-sheath nanofiber membrane was coaxially electrospun and strengthened with in situ glutaraldehyde cross-linking. This membrane not only kept high mass transfer rate, large extraction capacity and biomatrix resistance as our previously proposed membrane (Anal. Chem. 2013, 85 (12), 5924-5932), but also got much better mechanical strength and stability in water. The microfluidic device was designed to sandwich the membrane, and the blood in vivo can be introduced into it and get contact with the membrane repetitively. With this membrane and device, a 2-min equilibrium in vivo extraction method was established, validated in a simulated blood circulation system, and was used to monitor the pharmacokinetic profiles of desipramine in rabbits. The free and total concentration of desipramine in vivo was monitored with 10-min interval almost without rabbit blood consumed. The results met well with those of in vitro extraction, and a correlation factor of 0.99 was obtained.

Fast equilibrium micro-extraction from biological fluids with biocompatible core-sheath electrospun nanofibers.

Sample preparation methods with high temporal resolution and matrix resistance will benefit fast direct analysis of analytes in a complex matrix, such as drug monitoring in biofluids. In this work, the core-sheath biocompatible electrospun nanofiber was fabricated as a micro-solid phase extraction material. With the poly(N-isopropylacrylamide) (PNIPAAm) as sheath polymer and polystyrene (PS) as core polymer, the fiber membrane was highly hydrophilic and exhibited good antifouling ability to proteins and cells. Its complete expansion in aqueous solution and its nanoscale fiber (100-200 nm) structure offered high mass transfer rate of analytes between liquid and solid phases. The equilibration time of microextraction with this membrane was all shorter than 2 min for eight drugs tested, and the linear ranges covered more than 3 orders of magnitude for most of them. This membrane could be applied to monitor free drugs in plasma and their protein binding kinetics by equilibrium-microextraction with a 2 min temporal resolution. The results showed that the core-sheath electrospun nanofiber membrane would be a better alternative of solid phase material for microextraction with good matrix-resistance ability and high temporal resolution.

Detection of glutathione in vitro and in cells by the controlled self-assembly of nanorings.

Taking advantage of a reduction-controlled biocompatible condensation reaction and self-assembly, we have developed a new method for the determination of glutathione (GSH) concentration in vitro and in HepG2 human liver cancer cells. Upon reduction by GSH under physiological conditions (pH 7.4 in buffer), the small molecule CBT-Cys(SEt) condenses and self-assembles into nanorings, increasing the UV absorbance at 380 nm (with significant linear correlation in the 0-87 µM GSH range and a limit of detection of 1 µM). This method is also selective to GSH rather than cysteine in biological samples. Through the use of added internal standards, we successfully determined the concentration of GSH in HepG2 cells to be 14.96 µM (2.99 fmol/cell). To better understand the mechanism of nanoring self-assembly, the condensation product of CBT-Cys(SEt) formed using different concentrations of GSH and different reaction times were characterized by transmission electron microscopy (TEM).

An integrated micro-volume fiber-optic sensor for oxygen determination in exhaled breath based on iridium(III) complexes immobilized in fluorinated xerogels.

A novel integrated fiber-optic sensor with micro detection volume is developed and evaluated for O(2) determination on a breath-by-breath basis in human health monitoring applications. The sensing element was fabricated by dip-coating an uncladded optical fiber with [Ir(piq)(2)(acac)]-doped hybrid fluorinated ORMOSIL (organically modified silicate) film, which was prepared from 3,3,3-trifluoropropyltrimethoxysilane (TFP-TriMOS) and n-propyltrimethoxysilane (n-propyl-TriMOS). The sensor was then constructed by inserting the prepared optical fiber into a transparent capillary. A microchannel formed between the optical fiber and the capillary inner wall acted as a flow cell for the sample flowing through. The evanescent wave (EW) field produced on the fiber core surface can excite the O(2)-sensitive fluorophores of [Ir(piq)(2)(acac)] to produce emission fluorescence. O(2) can be sensed by its quenching effect on the emission fluorescence intensity. Spectroscopic properties have been characterized by FTIR and fluorescence measurements. Stern-Volmer and Demas models were both employed to analyse the sensor sensitivity, which is 13.0 with the LOD = 0.009% (3σ) and the response time is about 1 s. By integrating the sensing and detection elements on the optical fiber, the novel configuration showed advantages of easy fabrication and low cost. Parameters of sensitivity, response time, repeatability, humidity effect and temperature effect were discussed in detail. The proposed sensor showed potential for practical in-breath O(2) analysis application due to its advantages of easy fabrication, low cost, fast response, excellent hydrophobicity, negligible temperature interference and suitable sensitivity.