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In the present work, the density distribution of rhodamine 6G ions (R6G) in the gas phase and the droplets of an electrospray plume was studied by spatial and spectral imaging. The intention is to contribute to the fundamental understanding of the release mechanism of gaseous R6G in the electrospray ionization (ESI) process. Furthermore, the influence of ESI-parameters on the release efficiency of R6G, e. g. solvent flow, R6G and salt concentration were examined via direct fluorescence imaging of R6G. A solvent-shift of the fluorescence maximum,lambda(max) = 555 nm in methanolic solution and lambda(max) = 505 nm in gas phase, allows the discrimination between solvated and gaseous R6G. Two experimental setups were used for our measurements. In the first experiment, the R6G fluorescence and the light scattered from the spray plume were imaged in two spatial dimensions using a tunable wavelength filter. The second experiment was designed for obtaining 1-dimensional spatially resolved emission spectra of the spray. Here, the intensity distribution of solvated and gaseous R6G as well as scattered light (lambda = 355 nm) were measured simultaneously.
The results show the distribution of gaseous R6G in the plane, orthogonal to the ESI capillary, decreasing slightly towards the spray center and showing maxima at the cone margins. The distribution of gaseous R6G confirms the preferred release of gaseous ions from nano-droplets, indicating the ion evaporation model (IEM) to be the dominating release mechanism. Up to now, only a few fluorescence spectra of ionic compounds in the gas phase were published because the measurement of emission spectra of mass-selected ions in an ion trap is experimentally challenging. The fluorescence spectrum of gaseous lucigenin at atmospheric pressure is reported for the first time. This spectrum of lucigenin in the gas phase exhibits a blue shift of about Delta lambda = 10 nm in comparison to the corresponding spectrum in methanol.
Lahn M, Dosche C, Hille C. Two-photon microscopy and fluorescence lifetime imaging reveal stimulus-induced intracellular Na+ and Cl- changes in cockroach salivary acinar cells. Am J Physiol Cell Physiol 300: C1323-C1336, 2011. First published February 23, 2011; doi: 10.1152/ajpcell.00320.2010.-The intracellular ion homeostasis in cockroach salivary acinar cells during salivation is not satisfactorily understood. This is mainly due to technical problems regarding strong tissue autofluorescence and ineffective ion concentration quantification. For minimizing these problems, we describe the successful application of two-photon (2P) microscopy partly in combination with fluorescence lifetime imaging microscopy (FLIM) to record intracellular Na+ and Cl- concentrations ([Na+](i), [Cl-](i)) in cockroach salivary acinar cells. Quantitative 2P-FLIM Cl- measurements with the dye N-(ethoxycarbonylmethyl)-6-methoxy-quinolinium bromide indicate that the resting [Cl-](i) is 1.6 times above the Cl- electrochemical equilibrium but is not influenced by pharmacological inhibition of the Na+-K+-2Cl(-) cotransporter (NKCC) and anion exchanger using bumetanide and 4,4'-diisothiocyanatodihydrostilbene-2,2'-disulfonic acid disodium salt. In contrast, rapid Cl- reuptake after extracellular Cl- removal is almost totally NKCC mediated both in the absence and presence of dopamine. However, in physiological saline [Cl-](i) does not change during dopamine stimulation although dopamine stimulates fluid secretion in these glands. On the other hand, dopamine causes a decrease in the sodium-binding benzofuran isophthalate tetra-ammonium salt (SBFI) fluorescence and an increase in the Sodium Green fluorescence after 2P excitation. This opposite behavior of both dyes suggests a dopamine-induced [Na+](i) rise in the acinar cells, which is supported by the determined 2P-action cross sections of SBFI. The [Na+](i) rise is Cl- dependent and inhibited by bumetanide. The Ca2+-ionophore ionomycin also causes a bumetanide-sensitive [Na+](i) rise. We propose that a Ca2+-mediated NKCC activity in acinar peripheral cells attributable to dopamine stimulation serves for basolateral Na+ uptake during saliva secretion and that the concomitantly transported Cl- is recycled back to the bath.