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For scanning electron microscopy, formalin-fixed lung tissues were trimmed into 2-4-mm pieces, postfixed for 1 h in 1% osmium tetroxide, and rinsed for 30 min in 0.
1M PBS, postfixed with 2 percent osmium tetroxide in water for 2 hours, and finally washed again with 0.
After fixation and an overnight wash in sodium cacodylate buffer, the specimens were postfixed with l per cent osmium tetroxide in sodium cacodylate buffer for 1 h at 4 [degrees]C, dehydrated in alcohol and embedded in araldite resin.
The remaining tissue was postfixed in 4% paraformaldehyde for immunohistochemistry and stereologic counts.
Four beetles were prepared for Transmission Electron Microscopy (TEM) similarly, except they were postfixed in 2% osmium, then washed and dehydrated.
Before FISH, slides were rehydrated, postfixed in 10 g/L paraformaldehyde in phosphate-buffered saline (pH 7.
0) for 16 h at 4[degrees]C, postfixed in 1% osmium tetroxide for 1h, embedded, sectioned, double stained with uranyl acetate and lead citrate, and examined in a Hitachi H-600 (Tokyo, Japan) at 50 KV at a maginification of 20,000 x (Yu et al.
The other samples were immediately postfixed in 1% osmium tetroxide in the same buffer and stored in 70% ethanol.
1 M phosphate buffer, postfixed in 1% osmium tetroxide, and embedded in epoxy resin.
The pellets were postfixed in 1 per cent Osmium tetroxide for 30 min, dehydrated in ascending grades of acetone, infiltrated and embedded in araldite CY 212 (TAAB, UK).
1 M cacodylate, postfixed in 1% S-collidine-buffered osmium tetroxide (1 hr), and then dehydrated in graded ethanol washes.
Electron microscopy was performed with fresh tissues fixed in glutaraldehyde and postfixed in osmium tetroxide.