Fig. 3. Analysis of fluorescent fluid samples collected in optically flat hollow rectangle glass capillaries. (A) X–Y image of a sample of 50 µmol l^–1 Texas Red collected under paraffin oil in a hollow rectangle glass capillary. The width of the image is 250 µm. (B) X–Z image of a sample of 125 µmol l^–1 Texas Red. The solid line was positioned at the apparent midline of the bright band of fluorescence, corresponding to the Z-axis midpoint. For comparison, the upper and lower dotted lines indicate positions 10 µm from the apparent Z-axis midpoint. The dotted lines are clearly well away from the midline of the bright band in the X–Z image and are shown to indicate that the Z-axis midpoint can be reliably judged by eye. (C) Fluorescence intensity of X–Y images collected for the same sample at the Z-positions indicated. The curve shows that fluorescent intensity is nearly constant when the optical slice is positioned within 5 µm of the apparent Z-axis midpoint. (D) Representative calibration curve relating fluorescence intensity to the concentration of Texas Red in fluid samples collected in optically flat glass capillaries. Texas Red concentrations are plotted on the Y-axis so that the equation of the curve fit to the data by non-linear regression analysis allows calculation of the dye concentration of experimental samples from the measured fluorescence intensities. Measured fluorescence intensities were divided by 100 for convenience of the curve-fitting calculations. For the curve shown, CLSM detector gain was adjusted so that a near maximal signal (3900 on a scale of 0–4095) was obtained from the highest concentration (250 µmol l^–1) of Texas Red in this series of four calibration samples.