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Immunostaining Protocol

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25 views

Immunostaining Protocol

Uploaded by

anton
Copyright
© © All Rights Reserved
We take content rights seriously. If you suspect this is your content, claim it here.
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Immunostaining protocol:

Procedure

1.    Wash the cells with PBS and fix the cells with ice cold methanol for 20 minutes.

2.    Wash 3 x 5 minutes with PBS-T

3.    Block and permeabilize the cells for 30 minutes with PBS containing 0.2% triton X-100 and 1%
BSA.

4.    Wash 3 x 5 minutes with PBS-T

5.    Incubate for an hour with anti γH2AX antibody

6.    Wash 3 x 5 minutes with PBS-T

7.    Incubate for 5 minutes with propidium iodide

8.    Wash 3 x 5 minutes with PBS

9.    Put a drop of mounting media on objective glass slide. Put inverted stained cover slip on top of the
mounting media. Prevent air bubbles.

10.  Seal the edge of cover slip with nail protector

11.  Observe your samples using fluorescence microscope.  Alexa Fluor 488 is observable at 488 nm.
Propidium iodide is observable at 488 nm or 514 nm.

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