【求助】 电镜切片的制作
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谁能告诉我详细的电镜切片的制作过程阿
谢谢了
有篇外文的可是有些单词翻不出来
Microscopy
1. Rinse slices in 0.1M phosphate buffer pH 7.3.
2. Fix slices in 1.5% glutaraldehyde and 1.0% paraformaldehyde in 0.1M phosphate buffer ( pH 7.3) for 2H at 4°C.
3. Slice away membrane around culture*.
4. Rinse explant in phosphate buffer ( pH 7.3, cold solution) for 1H at 20°C in the dark.
5. Rinse 3 more times for 15 mins each in phosphate buffer.
6. Dehydrated samples through an ascending series of acetone concentrations ( 25, 50, 75, 95%) for 10 mins., each followed by 3 changes of 100% acetone for 20 minutes each.
7. Replace acetone with propylene-oxide (PO, 2 changes of 5 minutes each).
8. Infiltrate samples through graded PO:EPON mixtures ( 1:1, 1:3; 2H each).
9. Store overnight in 100% EPON.
Embedding
1. Embed slices flat in EPON between transparent foils for 48 Hr. At 600°C.
2. Remove olyester foils and the thin blocks of tissue re-embedded in EPON.
3. Polymerize at 60°C for 2 more days.
Staining
1. 1-2 µm thin sections can be stained in a solution of methylene blue and azur II in borax for light microscopy.
2. Ultra thin sections of 60-80 nm can be mounted on uncoated copper grids and stained with aqueous uranyl acetate and lead citrate.
3. Section can be examined with Philips EM300 electron Microscope at 80kV (or equivalent).
谢谢了
有篇外文的可是有些单词翻不出来
Microscopy
1. Rinse slices in 0.1M phosphate buffer pH 7.3.
2. Fix slices in 1.5% glutaraldehyde and 1.0% paraformaldehyde in 0.1M phosphate buffer ( pH 7.3) for 2H at 4°C.
3. Slice away membrane around culture*.
4. Rinse explant in phosphate buffer ( pH 7.3, cold solution) for 1H at 20°C in the dark.
5. Rinse 3 more times for 15 mins each in phosphate buffer.
6. Dehydrated samples through an ascending series of acetone concentrations ( 25, 50, 75, 95%) for 10 mins., each followed by 3 changes of 100% acetone for 20 minutes each.
7. Replace acetone with propylene-oxide (PO, 2 changes of 5 minutes each).
8. Infiltrate samples through graded PO:EPON mixtures ( 1:1, 1:3; 2H each).
9. Store overnight in 100% EPON.
Embedding
1. Embed slices flat in EPON between transparent foils for 48 Hr. At 600°C.
2. Remove olyester foils and the thin blocks of tissue re-embedded in EPON.
3. Polymerize at 60°C for 2 more days.
Staining
1. 1-2 µm thin sections can be stained in a solution of methylene blue and azur II in borax for light microscopy.
2. Ultra thin sections of 60-80 nm can be mounted on uncoated copper grids and stained with aqueous uranyl acetate and lead citrate.
3. Section can be examined with Philips EM300 electron Microscope at 80kV (or equivalent).
