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The volume of each sample for fluorescence measurement is 400 µL in 20 mM Tris-HCl buffer containing 100 mM NaCl, 5 mM KCl, and 15 mM MgCl2 (pH: 7.4) if not specified.
Men were eligible for this substudy if they had an available semen sample for fluorescence in situ hybridization (FISH) analysis.
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The samples for fluorescence measurements were prepared by spin-coating the solution of silica nanoparticles onto a clean microscope cover slip.
LH2 samples for fluorescence and fluorescence excitation studies had a maximum absorbance of ~ 0.15 (1 cm path cuvettes) in the largest absorption features (Soret and Qy).
Samples for fluorescence microscopy were prepared by incubating MOF crystals in vacuum-distilled furfuryl alcohol for 18 24 h at 353 K.
Samples for fluorescence compensation were prepared in which either the secondary antibody (R-phycoerythrin conjugate) or Draq 5 was omitted from the procedure.
For sludge samples, fixation in 50%% ethanol (final concentration) was recommended, which is the same as sample fixation for fluorescence in situ hybridization (FISH) (Xia et al. 2007).
Sample preparation for fluorescence microscopy was done as per [ 26] with modifications.
The sample for the fluorescence measurement was dissolved in the dry toluene, filtered, transferred to a long quartz cell, and then capped and bubbled with high pure argon (without O2and moisture) for at least 15 min before measurement.
We analyzed at least 10,000 individual sperm per sample for FITC fluorescence emissions.
Finally, a culture sample for furanocoumarin fluorescence determination was taken after 120 h of cultivation time [ 36].
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