3.3.
OSMIUM TETROXIDE FIXATIVES
39
Fixative:
(Continued)
0.1 N HC1
11
ml
Distilled water to make
50
ml
Osmium tetroxide (dry)
0.5 gm
The fixative should have a pH of 7.2-7.4, and its tonicity is estimated to
be 0.34M.
No adequate comparison of the Caulfield and Zetterqvist fixatives has
been made. There probably would be no detectable difference in the
deeper cells of most tissue blocks. Cells exposed at a surface, however,
experience the effect of an osmotic imbalance with extreme rapidity, and
are particularly sensitive to damage. Investigators who have most successfully dealt with tissue culture cells usually have added salts to the Palade
fixative, and thus have used formulas much like the Zetterqvist one.
Tahmisian (1964), who has studied the freezing-point depression of
Veronal-buffered osmium tetroxide mixtures, indicates that only 1 ml of
9% NaCl need be introduced as part of the formulation of each 25 ml of
Palade fixative to achieve equivalance with 9% (0.154 M) NaCl.
Palade has had such a profound influence on tissue preservation that
his least pronouncements often have carried the weight of divine authority. Thus it was that Palade (1952) indicated that he deliberately avoided
phosphate buffers because of the possibility that insoluble salts might
form within cells as a result of reactions with bivalent cations. Thus, subsequent investigators who occasionally did use phosphate-buffered osmium tetroxide fixatives tended to be ignored. However, Millonig (1961,
1962), while working at the Rockefeller Institute and at Harvard University, presented convincing demonstrations that phosphate buffers were at
least sometimes superior to Veronal. Some cytomembrane systems were
preserved with a detail that was lacking under other circumstances. Some
inclusions, notably glycogen, apparently were retained intact. The present
writer is happy to acknowledge that he has never seen any better preserved material than Millonig has shown, and certainly it has not commonly been equalled. The present writer has adopted the Millonig fixative for his own laboratory since its inception. We have had over two
years experience with it with very favorable results. It has proved to be
consistently reliable.
Millonig's buffer depends upon the equilibrium which forms between
monosodium phosphate and disodium phosphate when sodium hydroxide
is added to an isotonic solution of the former. By varying the amount of
OSMIUM TETROXIDE FIXATIVES
39
Fixative:
(Continued)
0.1 N HC1
11
ml
Distilled water to make
50
ml
Osmium tetroxide (dry)
0.5 gm
The fixative should have a pH of 7.2-7.4, and its tonicity is estimated to
be 0.34M.
No adequate comparison of the Caulfield and Zetterqvist fixatives has
been made. There probably would be no detectable difference in the
deeper cells of most tissue blocks. Cells exposed at a surface, however,
experience the effect of an osmotic imbalance with extreme rapidity, and
are particularly sensitive to damage. Investigators who have most successfully dealt with tissue culture cells usually have added salts to the Palade
fixative, and thus have used formulas much like the Zetterqvist one.
Tahmisian (1964), who has studied the freezing-point depression of
Veronal-buffered osmium tetroxide mixtures, indicates that only 1 ml of
9% NaCl need be introduced as part of the formulation of each 25 ml of
Palade fixative to achieve equivalance with 9% (0.154 M) NaCl.
Palade has had such a profound influence on tissue preservation that
his least pronouncements often have carried the weight of divine authority. Thus it was that Palade (1952) indicated that he deliberately avoided
phosphate buffers because of the possibility that insoluble salts might
form within cells as a result of reactions with bivalent cations. Thus, subsequent investigators who occasionally did use phosphate-buffered osmium tetroxide fixatives tended to be ignored. However, Millonig (1961,
1962), while working at the Rockefeller Institute and at Harvard University, presented convincing demonstrations that phosphate buffers were at
least sometimes superior to Veronal. Some cytomembrane systems were
preserved with a detail that was lacking under other circumstances. Some
inclusions, notably glycogen, apparently were retained intact. The present
writer is happy to acknowledge that he has never seen any better preserved material than Millonig has shown, and certainly it has not commonly been equalled. The present writer has adopted the Millonig fixative for his own laboratory since its inception. We have had over two
years experience with it with very favorable results. It has proved to be
consistently reliable.
Millonig's buffer depends upon the equilibrium which forms between
monosodium phosphate and disodium phosphate when sodium hydroxide
is added to an isotonic solution of the former. By varying the amount of