For routine use:

• tetramethylrhodamine dextran, lysine fixable MW 10,000 (Molecular Probes D-1817) - orange/red fluorescence
• fluorescein dextran, lysine fixable MW 10,000 (Molecular Probes D-1820) - green fluorescence [use a mildly basic buffer environment (pH9-10) for best fluorescence emission]

Additional useful dextran tracers:

• biotinylated dextran, amine fixable MW 10,000 (Molecular Probes D-1956) - to be further labeled with conjugated avidin/streptavidin or antibody for detection
• Cascade Blue dextran, lysine fixable MW 10,000 (Molecular Probes D-1976) - blue fluorescence
• Texas Red dextran, lysine fixable MW 10,000 (Molecular Probe D-1863) - near red fluorescence, not suitable for use in combination with tetramethylrhodamine
• Alexa Fluor 647 dextran, amine fixable MW 10,000 (Molecular Probes D-22914) - far red fluorescence, invisible to human eyes

Spectral variants of Alexa Fluor dye conjugated fixable dextran MW 10,000 (AF488 D-22910, AF546 D-22911, AF568 D-22912, AF594 D-22913) are available from Molecular Probes. They are more expensive and without a significant advantage over tetramethylrhodamine and fluorescein for this particular application.

Dextran tracer stock solution (except Alexa Fluor dextran) is 100 mg/ml in nuclease-free water or 0.2M KCl, filtered through Ultrafree MC GV 0.22 μm filter (Millipore), aliquoted into 4 μl each tube, and store in -80°C.

Alexa Fluor dextran stock solution is 20 mg/ml in nuclease-free water or 0.2M KCl, filtered through Ultrafree MC GV 0.22 μm filter (Millipore), aliquoted into 2 μl each tube, and store at -80°C.

For pressure injection, dilute into 5-50 mg/ml (non-Alexa Fluor dextran) or 1-10 mg/ml (Alexa Fluor dextran) with 1% Fast Green (Sigma) in 0.2M KCl or distilled water, depending on the purpose and the fluorophore. For visualizing late-stage pattern elements with tetramethylrhodamine dextran and/or fluorescein dextran, use 25-50 mg/ml. Alexa Fluor dyes have a better quantum efficiency and, they can be used at a lower concentration (5-10 mg/ml). 0.5 μl is enough to load a single micropipette. Store the unused portion in -20°C. This is good for a few weeks; avoid long-term storage in -20°C (the main problem is the loss of water by freeze-drying effect).

Fast Green stock solution: 4% in 0.2M KCl (made with UltraPure water), Whatman paper filtered, stored at 4°C.; 0.22 μm filter (syringe filter or Ultrafree MC GV centrifuge filter) before use.

Priority in dye selection: tetramethylrhodamine > fluorescein > Alexa Fluor 647 > Cascade Blue.

Avidin and streptavidin bind to nephridia and are thus not a good choice for labeling biotinylated dextran in late-stage embryos.