Microtiter plate assay for phosphate using a europium–tetracycline complex as a sensitive luminescent probe

A new luminescent europium probe is presented for the determination of phosphate (P) in microtiter plate format. The assay is based on the quenching of the luminescence of the europium–tetracycline (EuTc) 1:1 complex by phosphate using a reagent concentration of 20.8 μmol/L. The probe is excited at 400 nm and displays a large Stokes’ shift of 210 nm. The emission maximum is located at 616 nm. The system works best at neutral pH 7 and is therefore suitable for phosphate determination in biological and biochemical systems. The linear range of the calibration plot is from 5 × 10−6 mol/L to 7.5 × 10−4 mol/L of phosphate, and the limit of detection is 3 μmol/L

Anion-lnduced Fluorescence Quenching of a New Zwitterionic Biacridine Derivative

The effect of halides and different buffer anions on the quenching of the fluorescence of the new probe 10,10′-bis(3-sulfopropyl)-9,9′-biacridine (SPBA) has been studied using fluorescence and decay time measurements. The linearity of the Stern-Volmer plot indicates that fluorescence quenching by halides can be described reasonably well by a single-exponential decay with a K of 4.06 times 10⁶M⁻¹s⁻¹for chloride, 7.83 times 10⁶M⁻¹s⁻¹for bromide and 1.12 times 10⁷M⁻¹s⁻¹for iodide. We have found that SPBA is collisionally quenched also by the buffers 3-(N-mor-pholino)propanesulfonic acid (MOPS) and N-2-hydroxy-ethylpiperazine-N′-ethansulfonic acid (HEPES). The bi-molecular rate constants are 1.67 × 10⁶M⁻¹s⁻¹for HEPES and 1.44 times 10⁶M⁻¹s⁻¹for MOPS

Hypoxia in Leishmania major Skin Lesions Impairs the NO-Dependent Leishmanicidal Activity of Macrophages

Cure of infections with Leishmania major is critically dependent on the ability of macrophages to induce the type 2 nitic oxide (NO) synthase (NOS2) that produces high levels of NO in the presence of ample oxygen. Therefore, we analyzed the oxygen levels found in leishmanial skin lesions and their effect on the NOS2-dependent leishmanicidal activity of macrophages (MΦ). When L. major skin lesions of self-healing C57BL/6 mice reached their maximum size, the infected tissue displayed low oxygen levels (pO2~21Torr). MΦ activated under these oxygen tensions failed to produce sufficient amounts of NO to clear L. major. Nos2-deficient and hypoxic wild-type macrophages displayed a similar phenotype. Killing was restored when MΦ were reoxygenated or exposed to a NO donor. The resolution of the lesion in C57BL/6 mice was paralleled by an increase of lesional pO2. When mice were kept under normobaric hypoxia, this caused a persistent suppression of the lesional pO2 and a concurrent increase of the parasite load. In Nos2-deficient mice, there was no effect of atmospheric hypoxia. Low oxygen levels found at leishmanial skin lesions impaired the NOS2-dependent leishmanicidal activity of MΦ. Hence, tissue oxygenation represents an underestimated local milieu factor that participates in the persistence of Leishmania

A Fluorescent Probe for Diacetyl Detection

A water-soluble fluorescent probe, rhodamine B hydrazide (RBH), was prepared and its properties for recognition of diacetyl were studied. The method employs the reaction of diacetyl with RBH, a colorless and non-fluorescent rhodamine B spiro form derivative to give a pink-colored fluorescent substance. In weakly acidic media, RBH reacts more selectively with diacetyl than with other carbonyls, causing a large increase in fluorescence intensity and thereby providing an easy assay for the determination of diacetyl

Fluorescent chameleon labels for bioconjugation and imaging of proteins, nucleic acids, biogenic amines and surface amino groups. a review

Chameleon labels (ChLs) possess the unique property of changing (visible) color and fluorescence on binding to amino groups of biomolecules. MostChLs react with primary aliphatic amino groups such as those in lysine or with amino groups artificially introduced into polynucleic acids or saccharides, but someothers also react with secondary amino groups. Under controlled circumstances, the reactions are fairly specific. The review is subdivided into the following sections: (1) An introduction and classification of fluorescent labels; (2) pyrylium labels that undergo shortwave color changes upon labelling, typically from blue to red; (3) polymethine type of labels (that also undergo shortwave color changes, typically from green to blue; (4) various other (less common) chromogenic and fluorogenic systems; (5) hemicyanine labels that undergo longwave color changes, typically from yellow to purple; (6) the application of ChLs to labeling of proteins and oligonucleotides; (7) applications to fluorometric assays and sensing; (8) applications to fluorescence imaging of biomolecules; (9) applications in studies on affinity interactions (receptor-ligand binding); (10) applications in surface and interface chemistry; and (11) applications in chromatography, electrophoresis and isotachophoresis of biomolecules