The fluorescent microscopy method for pollen viability determination
is a method with which it is possible to monitor pollen
germination on stigma and pollen tube growth through the style to
the ovary and egg cell. It was described in detail by Kho and Baër
(1968) and it has been used with certain modifications for different
plant species.
A few hours or days after pollination the pistil is fixed as a sample
for the analysis. Fixation lasts in most cases about 24 h on room
temperature. Most frequently used fixatives are Carnoy I (3:1; alcohol:
acetic acid) and F.A.A. (1:1:8; formalin: acetic acid: alcohol).
The fixed sample can be kept in refrigeration on +4 ◦C in 70% alcohol
until the sample preparation. Prior to the sample preparation
pistil is softened in a NaOH solution of various concentrations and
time intervals depending on the studied plant species. The staining
is made with 0.1% aniline blue (dissolved in 1 N K3PO4) for 12–24 h.
Some plant species require the usage of fluorochrome.