7.3.
ALKALINE LEAD HYDROXIDE STAINS
221
dinary potentiality. It was also immediately apparent that there were
major difficulties in handling it. Lead ions are notorious for forming
insoluble crystals with most anions. One need only recall that lead carbonate is such an insoluble salt that it long served as the principal white
paint pigment of commerce. Crystals of lead carbonate happen to be
extremely hydrophobic. A scum of such crystals forms almost immediately
at the surface of any solution containing lead ions as soon as it comes into
contact with the carbon dioxide of air. This seriously contaminates any
grid that then comes in contact with the interface (Fig. 10). Even when
more or less heroic means are employed to prevent a surface scum from
forming, or to avoid passing grids through it, dissolved carbon dioxide
is a hazard of serious proportions. Even the carbonate contamination of
stock bottles of hydroxide used to prepare the stains represent a source
of trouble (Fig. 10). Thus it is that it has taken about 5 years for scientific
evolution to modify the original stain so that these hazards are reduced
to easily manageable proportions, and to devise simple and practical
operational techniques.
The stain that certainly is safest to use is the formulation of Reynolds
(1963), which incorporates citrate as a chelating agent. Mounted sections
can be stained with this in a reliable and simple way. In our own experience, however, this stain is not always as vigorous as the original Watson
formulation (nor do any of the other available variants seem to act quite
as powerfully). Thus, it seems as though at times there may be good
reason to continue to use the original Watson stain, so it is my intention
to describe this, as well as a relatively simple way of handling the stain
which has proved reliable in our own laboratory. In what follows the
Reynolds stain is presented first as being the most generally useful stain,
then the Watson stain will be described, and finally other variants which
have enjoyed popularity.
A number of the micrographs illustrated in this book are of tissues
stained with one or another of the alkaline lead hydroxide formulations.
These include Figs. 3, 10, 12, 28(a), and 39. It should also be noted that
tissue heavily stained with lead is quite sensitive to "beam damage" if
excessive illumination is used. Sufficient heat can be generated to liquify
local areas complexed with the lead, and coalescence occurs as tiny droplets. This is well illustrated in Fig. 25.
with lead are particularly vulnerable to this artifact, apparently because the leaded
areas easily can become heated to liquefaction, whereupon surface tension causes
coalescence, and produces fine granular deposits, (a) Liver, (b) myelin.
ALKALINE LEAD HYDROXIDE STAINS
221
dinary potentiality. It was also immediately apparent that there were
major difficulties in handling it. Lead ions are notorious for forming
insoluble crystals with most anions. One need only recall that lead carbonate is such an insoluble salt that it long served as the principal white
paint pigment of commerce. Crystals of lead carbonate happen to be
extremely hydrophobic. A scum of such crystals forms almost immediately
at the surface of any solution containing lead ions as soon as it comes into
contact with the carbon dioxide of air. This seriously contaminates any
grid that then comes in contact with the interface (Fig. 10). Even when
more or less heroic means are employed to prevent a surface scum from
forming, or to avoid passing grids through it, dissolved carbon dioxide
is a hazard of serious proportions. Even the carbonate contamination of
stock bottles of hydroxide used to prepare the stains represent a source
of trouble (Fig. 10). Thus it is that it has taken about 5 years for scientific
evolution to modify the original stain so that these hazards are reduced
to easily manageable proportions, and to devise simple and practical
operational techniques.
The stain that certainly is safest to use is the formulation of Reynolds
(1963), which incorporates citrate as a chelating agent. Mounted sections
can be stained with this in a reliable and simple way. In our own experience, however, this stain is not always as vigorous as the original Watson
formulation (nor do any of the other available variants seem to act quite
as powerfully). Thus, it seems as though at times there may be good
reason to continue to use the original Watson stain, so it is my intention
to describe this, as well as a relatively simple way of handling the stain
which has proved reliable in our own laboratory. In what follows the
Reynolds stain is presented first as being the most generally useful stain,
then the Watson stain will be described, and finally other variants which
have enjoyed popularity.
A number of the micrographs illustrated in this book are of tissues
stained with one or another of the alkaline lead hydroxide formulations.
These include Figs. 3, 10, 12, 28(a), and 39. It should also be noted that
tissue heavily stained with lead is quite sensitive to "beam damage" if
excessive illumination is used. Sufficient heat can be generated to liquify
local areas complexed with the lead, and coalescence occurs as tiny droplets. This is well illustrated in Fig. 25.
with lead are particularly vulnerable to this artifact, apparently because the leaded
areas easily can become heated to liquefaction, whereupon surface tension causes
coalescence, and produces fine granular deposits, (a) Liver, (b) myelin.