4.13.
POLYESTER VESTOPAL W EMBEDDING
117
tion well. The problems with this embedment relate to achieving uniform
infiltration. When the tissue specimen has altogether different physical
properties than the surrounding empty plastic, one must suspect that
there was a non-uniform distribution of the plastic mixture components
when polymerized. Freeman and Spurlock originally admitted encountering this sort of trouble, and suggested cutting the tissue out of the
first embedment, and re-embedding and re-curing in a fresh mixture. In
the later paper they emphasize that all alcohol must be eliminated from
the specimen by two changes of propylene oxide before infiltrating with
the resin mixture. Finally, they soak the tissue block overnight in the
Maraglas mixture at 10°C to insure internal equilibrium.
The difficulties with impregnation probably result from using Cardolite in the formulation, for this is known to be a very large molecule
which would not be expected to penetrate tissues well. However, it may
be that dibutyl phthalate is the villain for similar problems were encountered in the early days of the use of Araldite when efforts were made to
control its final hardness with this plasticizer.
Advantages claimed for Maraglas include a substantially greater transparency than Araldite, and an easier tissue "stainability." There are some
special hazards in sectioning Epon which sometimes disturb the surface
of sections leading to a "mottling" (Chapter 4.11, Fig. 11). Maraglas is
said not to be as prone to this effect as Epon, but some "mottling" is
visible in the figures the original authors have published so any advantage may be only a relative one. It is also claimed that Maraglas does not
show the grain which is often visible in Epon sections. We will need further experience to assess the value of this embedment in relation to others
of the epoxy group.
4.13. Polyester Vestopal W Embedding
In 1956 Kellenberger et al. demonstrated that polyesters might be used
as embedding media. Early results admittedly were inconsistent, but later
Ryter and Kellenberger (1958) demonstrated that Vestopal W was a
better choice than the resin originally used, and it has received wide
acceptance since. Figure 12 illustrates its use.
FIG. 12. Nerve cell cytoplasm, fixed first by perfusion with buffered formaldehyde,
secondarily fixed with Millonig's buffered osmium tetroxide, embedded in Vestopal W,
stained with alkaline lead hydroxide. Cytomembrane systems and protoplasmic proteins
reasonably well preserved and darkly stained, ribosomes extremely dense. In the upper
right is cytoplasm of an adjacent astrocyte in which densely stained granular masses are
thought to be glycogen.
POLYESTER VESTOPAL W EMBEDDING
117
tion well. The problems with this embedment relate to achieving uniform
infiltration. When the tissue specimen has altogether different physical
properties than the surrounding empty plastic, one must suspect that
there was a non-uniform distribution of the plastic mixture components
when polymerized. Freeman and Spurlock originally admitted encountering this sort of trouble, and suggested cutting the tissue out of the
first embedment, and re-embedding and re-curing in a fresh mixture. In
the later paper they emphasize that all alcohol must be eliminated from
the specimen by two changes of propylene oxide before infiltrating with
the resin mixture. Finally, they soak the tissue block overnight in the
Maraglas mixture at 10°C to insure internal equilibrium.
The difficulties with impregnation probably result from using Cardolite in the formulation, for this is known to be a very large molecule
which would not be expected to penetrate tissues well. However, it may
be that dibutyl phthalate is the villain for similar problems were encountered in the early days of the use of Araldite when efforts were made to
control its final hardness with this plasticizer.
Advantages claimed for Maraglas include a substantially greater transparency than Araldite, and an easier tissue "stainability." There are some
special hazards in sectioning Epon which sometimes disturb the surface
of sections leading to a "mottling" (Chapter 4.11, Fig. 11). Maraglas is
said not to be as prone to this effect as Epon, but some "mottling" is
visible in the figures the original authors have published so any advantage may be only a relative one. It is also claimed that Maraglas does not
show the grain which is often visible in Epon sections. We will need further experience to assess the value of this embedment in relation to others
of the epoxy group.
4.13. Polyester Vestopal W Embedding
In 1956 Kellenberger et al. demonstrated that polyesters might be used
as embedding media. Early results admittedly were inconsistent, but later
Ryter and Kellenberger (1958) demonstrated that Vestopal W was a
better choice than the resin originally used, and it has received wide
acceptance since. Figure 12 illustrates its use.
FIG. 12. Nerve cell cytoplasm, fixed first by perfusion with buffered formaldehyde,
secondarily fixed with Millonig's buffered osmium tetroxide, embedded in Vestopal W,
stained with alkaline lead hydroxide. Cytomembrane systems and protoplasmic proteins
reasonably well preserved and darkly stained, ribosomes extremely dense. In the upper
right is cytoplasm of an adjacent astrocyte in which densely stained granular masses are
thought to be glycogen.