A novel fluorescence method for detection of phosphate anions based on porphyrin metalation.

In this work, a novel fluorescence method for the detection of phosphate anions (PO43-) was developed based on porphyrin metalation. Through catalysis by G-quadruplex (G4), Cu2+ could insert into the porphyrins to quench their fluorescence. G4 simultaneously improved the fluorescence of the porphyrins but not that of Cu2+-porphyrin. In the absence of PO43-, the porphyrins were metallized by Cu2+, and no fluorescence was observed. In the presence of PO43-, PO43- could coordinate with Cu2+ to prevent porphyrin metalation. Free porphyrin could bind with G4 to emit strong fluorescence. By comparing four common porphyrins, we found that G4 had the greatest effect on increasing the fluorescence intensity of N-methylmesoporphyrin IX (NMM). Thus, NMM/G4 was chosen for the design of a biosensor. Under optimal experimental conditions, this method showed high sensitivity and satisfactory selectivity for PO43- with a detection limit of 44 nM in a linear range of 0.01-1.0 µM. The recovery experiments showed recovery rates of 93.75-106.00%, suggesting a great potential for measuring PO43- in real samples.

Determination of Kanamycin by High Performance Liquid Chromatography.

Kanamycin is an aminoglycoside antibiotic widely used in treating animal diseases caused by Gram-negative and Gram-positive infections. Kanamycin has a relatively narrow therapeutic index, and can accumulate in the human body through the food chain. The abuse of kanamycin can have serious side-effects. Therefore, it was necessary to develop a sensitive and selective analysis method to detect kanamycin residue in food to ensure public health. There are many analytical methods to determine kanamycin concentration, among which high performance liquid chromatography (HPLC) is a common and practical tool. This paper presents a review of the application of HPLC analysis of kanamycin in different sample matrices. The different detectors coupled with HPLC, including Ultraviolet (UV)/Fluorescence, Evaporative Light Scattering Detector (ELSD)/Pulsed Electrochemical Detection (PED), and Mass Spectrometry, are discussed. Meanwhile, the strengths and weaknesses of each method are compared. The pre-treatment methods of food samples, including protein precipitation, liquid-liquid extraction (LLE), and solid-phase extraction (SPE) are also summarized in this paper.

Simple and Label-Free Fluorescent Detection of Melamine Based on Melamine⁻Thymine Recognition.

In the past few years, melamine has been illegally added into dairy products to increase the apparent crude protein levels. If humans or animals drink the milk adulteration of melamine, it can form insoluble melamine⁻cyanurate crystals in their kidneys which causes kidney damage or even death. In the present work, we constructed a simple and label-free fluorescent method for melamine detection based on melamine-thymine recognition. SYBR Green I was utilized as a reporter for this method as it did not require any modification or expensive equipment. In the absence of melamine, polythymine DNA was digested by Exo I, which caused a decrease in the fluorescence signal. In the presence of melamine, the polythymine DNA was able to fold into a double chain structure, however this was done with the help of T-melamine-T mismatches to prevent degradation. Then, the SYBR Green I combined with the double-stranded DNA to result in an intense fluorescence signal. The limit of detection in this method was 1.58 µM, which satisfied the FDA standards. This method also had a good linear relationship within the range of 10⁻200 µM. In addition, this new method has a good selectivity to distinguish melamine from the component of milk. As a result, we developed a simple and highly selectivity method for melamine detection.

A Label-free "Turn-on" Fluorescence Method for Detecting Mercury Ion in Aqueous Solution Based on N-Methyl Mesoporphyrin IX (NMM)/G-quadruplex DNA.

In the present work, a simple design of "turn-on" fluorescence method for mercury ion was developed and explored based on specific T-Hg-T mismatches as a recognizer and N-methyl mesoporphyrin IX (NMM)/G-quadruplex DNA system as a reporter. The titration experiment showed that the mercury-ion concentration and the fluorescence intensity signal change exhibited a consistent linear correlation within the 50 to 500 nM range with a detection limit down to 12.9 nM. In a selectivity experiment, our method showed obvious selectivity against other metal ions, being consistent with other reported detection methods of mercury ion based on the specific reactivity of T-Hg-T mismatches. Our method was then successfully employed to detect mercury ion in pond water with excellent reliability.

Decreased peripheral natural killer cells activity in the immune activated stage of chronic hepatitis B.

BACKGROUND & AIMS: The natural course of chronic hepatitis B virus (HBV) infection is characterized by different immune responses, ranging from immune tolerant (IT) to immune activated (IA) stages. In our study, we investigated the natural killer (NK) cells activity in patients at different immunological stages of chronic HBV infection. METHODS: Blood samples obtained from 57 HBeAg positive patients with chronic hepatitis B (CHB), including 15 patients in the immune tolerant (IT) stage, 42 patients in the immune activated (IA) stage, and 18 healthy individuals (HI). The analyses included flow cytometry to detect NK cells, the determination of cytokine levels as well as of surface receptor expression and cytotoxicity. RESULTS: NK cells in peripheral blood were significantly lower in patients in the IA stage of CHB compared to HI (p<0.05). Patients in the IA stage of CHB had lower levels of NK cells activating receptor NKp30 and NKG2D expression, cytokine interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α) production, as compared to patients in the IT stage and HI, respectively (p<0.05). Cytotoxicity of NK cells was lower in patients in the IA stage of CHB compared to patients in the IT stage and HI, respectively (p<0.05). The level of IFN-γ but not level of TNF-α and cytotoxicity of NK cells was inversely correlated with serum HBV load in patients with CHB. Peripheral NK cells activity did not correlate with ALT level. CONCLUSION: NK cells activity was lower in CHB patients, especially in those in the IA stage.