The solution is no longer usable when it turns a dull, dark orange color. For Arabidopsis apices, add about 500 ul staining solution per apex in the vial. Larger samples may require longer staining times. After the staining procedure, the tissue may appear slightly orange. Rinse with 0.1 M L-arginine buffer, with the pH adjusted to 8.0 with HCl.
Shorter changes result in incomplete clearing.
Leaving the tissue longer than 2 hours in any of the xylene series solutions does not harm the sample. Place a series of 4 individual drops of immersion oil on a clear standard microscope slide. The first drop should be large enough to contain the entire sample. Using fine forceps, place the tissue sample in the first drop, and begin dissection under a dissecting scope. In the case of the shoot apex, hold the stem with one forceps, and break off the older flowers by pushing them toward the base of the stem.