Fluorescence Differentiation of ATP-related Multiple Enzymatic Activities in Synovial Fluid as a Marker of Calcium Pyrophosphate Deposition Disease using Kyoto Green.

Calcium pyrophosphate deposition disease (CPPD) is a crystal induced inflammation in joints, and causes severe pain in elderly people. The accumulation of pyrophosphate (PPi) in synovial fluid (SF) results from several enzymatic reactions, especially the highly activated e-NPPs, which catalyze the conversion of ATP to PPi. This study demonstrates the detection of relative catalytic activity of 3 enzymes-ecto-nucleotide pyrophosphatase/phosphodiesterases (e-NPPs), tissue nonspecific alkaline phosphatase (TNAP), and ecto-nucleoside triphosphate diphosphohydrolases (e-NTPDases)-using a single molecular sensor called Kyoto Green. Kyoto Green exhibits excellent performance in sensing the catalytic activity of the commercial representatives of the e-NPPs, TNAP, and e-NTPDases, which are ENPP1, PPase, and apyrase, respectively, in both single-enzyme and multi-enzyme assays. Analysis of SF enzymes in 19 SF samples from human and swine revealed moderate activity of e-NPPs, high activity of e-NTPDases, and low activity of TNAP. Our newly developed method for analysis of multiple enzymatic activities using Kyoto Green in biological SF will assist improvement in accuracy of the CPPD prognosis/diagnosis, which will minimize unnecessary medical procedures.

Arsenic toxicity in the water weed Wolffia arrhiza measured using Pulse Amplitude Modulation Fluorometry (PAM) measurements of photosynthesis.

Accumulation of arsenic in plants is a serious South-east Asian environmental problem. Photosynthesis in the small aquatic angiosperm Wolffia arrhiza is very sensitive to arsenic toxicity, particularly in water below pH 7 where arsenite (As (OH)3) (AsIII) is the dominant form; at pH >7 AsO4(2-) (As(V) predominates). A blue-diode PAM (Pulse Amplitude Fluorometer) machine was used to monitor photosynthesis in Wolffia. Maximum gross photosynthesis (Pgmax) and not maximum yield (Ymax) is the most reliable indicator of arsenic toxicity. The toxicity of arsenite As(III) and arsenate (H2AsO4(2-)) As(V) vary with pH. As(V) was less toxic than As(III) at both pH 5 and pH 8 but both forms of arsenic were toxic (>90% inhibition) at below 0.1molm(-3) when incubated in arsenic for 24h. Arsenite toxicity was apparent after 1h based on Pgmax and gradually increased over 7h but there was no apparent effect on Ymax or photosynthetic efficiency (α0).

Mesoporous Silica Nanoparticles-Enhanced Microarray Technology for Highly Sensitive Simultaneous Detection of Multiplex Foodborne Pathogens.

Ensuring food safety is paramount for the food industry and global health concerns. In this study, we have developed a method for the detection of prevalent foodborne pathogenic bacteria, including Escherichia coli, Salmonella spp., Listeria spp., Shigella spp., Campylobacter spp., Clostridium spp., and Vibrio spp., utilizing antibody-aptamer arrays. To enhance the fluorescence signals on the microarray, the mesoporous silica nanoparticles (MSNs) conjugated with fluorescein, streptavidin, and seven detection antibodies-biotin were employed, forming fluorescein doped mesoporous silica nanoparticles conjugated with detection antibodies (MSNs-Flu-SA-Abs) complexes. The array pattern was designed for easy readability and enabled the simultaneous detection of all seven foodborne pathogens, referred to as the 7FP-biochip. Following the optimization of MSNs-Flu-SA-Abs complexes attachment and enhancement of the detection signal in fluorescent immunoassays, a high level of sensitivity was achieved. The detection limits for the seven pathogens in both buffer and food samples were 102 CFU/mL through visual screening, with fluorescent intensity quantification achieving levels as low as 20-34 CFU/g were achieved on the antibody-aptamer arrays. Our antibody-aptamer array offers several advantages, including significantly reduced nonspecific binding with no cross-reaction between bacteria. Importantly, our platform detection exhibited no cross-reactivity among the tested bacteria in this study. The multiplex detection of foodborne pathogens in canned tuna samples with spiked bacteria was successfully demonstrated in real food measurements. In conclusion, our study presents a promising method for detecting multiple foodborne pathogens simultaneously. With its high sensitivity and specificity, the developed antibody-aptamer array holds great potential for enhancing food safety and public health.

Masking Phosphate with Rare-Earth Elements Enables Selective Detection of Arsenate by Dipycolylamine-ZnII Chemosensor.

Functional reassessment of the phosphate-specific chemosensors revealed their potential as arsenate detectors. A series of dipicolylamine (Dpa)-ZnII chemosensors were screened, among which acridine Dpa-ZnII chemosensor showed the highest capability in sensing arsenate. The presence of excess ZnII improved sensitivity and strengthened the binding between acridine Dpa-ZnII complex to arsenate as well as phosphate. However, due to their response to phosphate, these sensors are not suited for arsenate detection when phosphate is also present. This study demonstrated for the first time that rare-earth elements could effectively mask phosphate, allowing the specific fluorescence detection of arsenate in phosphate-arsenate coexisting systems. In addition, detection of arsenate contamination in the real river water samples and soil samples was performed to prove its practical use. This sensor was further employed for the visualization of arsenate and phosphate uptake in vegetables and flowering plants for the first time, as well as in the evaluation of a potent inhibitor of arsenate/phosphate uptake.

Measurement of photosynthesis using PAM technology in a purple sulfur bacterium Thermochromatium tepidum (Chromatiaceae).

We demonstrate that Blue-diode-based pulse amplitude modulation (PAM) technology can be used to measure the photosynthetic electron transport rate (ETR) of purple sulfur bacteria (Thermochromatium tepidum, Chromatiaceae). Previous studies showed that PAM technology could be used to estimate photosynthesis in purple nonsulfur bacteria and so PAM technology can be used to estimate photosynthesis of both kinds of purple photosynthetic bacteria. The absorptance of Thermochromatium films on glass fiber disks was measured and used to calculate actual ETR. ETR vs Irradiance (P vs E) curves fitted the waiting-in-line model (ETR = (ETRmax × E/Eopt) × exp (1−E/Eopt)). Yield (Y) was only ≈ 0.3­0.4. Thermochromatium saturates at 325 ± 13.8 µmol photons m(−2) s(−1) or ≈15% sunlight and shows photoinhibition at high irradiances. A pond of Thermochromatium would exhibit classic surface inhibition. Photosynthesis is extremely low in the absence of an electron source: ETR increases in the presence of acetate (5 mol m(−3)) provided as an organic carbon source and also increases in the presence of sulfite (3 mol m(−3)) but not sulfide and is only marginally increased by the presence of Fe(2+). Nonphotochemical quenching does occur in Thermochromatium but at very low levels compared to oxygenic photo-organisms or Rhodopseudomonads.