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EM Immunogold Labeling Protocol - P(3)

时间:2006-01-22 15:57来源:ihcworld.com 作者:bioguider 点击: 1181次

10X Phosphate Buffered Saline (PBS):

To prepare 1 liter,

Na2HPO4 --------------------- 10.9 g

NaH2PO4 --------------------- 3.2 g

NaCl --------------------------- 90 g

Distilled water -------------- 1000 ml

Mix to dissolve and adjust pH to 7.4

 

0.2M Citric Acid:

To prepare 100 ml,

Citric acid (monohydrate) ------ 4.2 g

Distilled water -------------------- 100 ml

 

0.2M Sodium Citrate Buffer (fresh):

To prepare 100 ml,

Sodium citrate (dihydrate) ------ 5.88 g

Distilled water --------------------- 100 ml

Adjust pH to 7.4 with 0.2M citric acid (keeps refrigerated).

 

Fixative (4% Paraformaldehyde and 0.5% Glutaraldehyde in 0.1M PB):

To prepare 1 liter,

Paraformaldedyde -------------- 40 g

0.1M PB --------------------------- 1000 ml

Heat to 65 C. Add a few drops of 1N NaOH to clear the solution. Stir to dissolve. Filter the solution and add 10 ml of 50% glutaraldehyde.

 

Post-fixative (1% Osmium Tetroxide in 0.1M PB):

To prepare 20 ml,

4% osmium tetroxide ---------- 5 ml

0.2M PB --------------------------- 5 ml

0.1M PB -------------------------- 10 ml

 

2% Glutaraldehyde in PBS:

To prepare 100 ml,

50% glutaraldyhyde ------------ 4 ml

PBS -------------------------------- 100 ml

 

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