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How to get rid of nuclear stain (unspecific)?

change blocking
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change fixation
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anything else?
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Offline Sona

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Unspecific nuclear stain
« on: February 25, 2010, 08:26:52 AM »
Hi All,

Do any of you know how to get rid of nuclear stainning (unspecific). I am trying to stain primary cells for vinculin. I can see a specific vinculin stainning in the focal adhesions but ... there is also an unspecific nuclear stain! I have tried different fixatives (methanol,acetone, paraformaldehye) and different blocking serum(goat, casein, BSA).... also tried low cross buffer...  but could not get rid of the nuclear stain.

Please let me know if any of you have a solution for this.

Thanks
Sona.

Unspecific nuclear stain
« on: February 25, 2010, 08:26:52 AM »

Offline richard03

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Re: Unspecific nuclear stain
« Reply #1 on: March 07, 2010, 10:30:06 PM »
Try higher dilution of primary antibody and sometimes it will be sufficient to remove unspecific staining.

Offline gula

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Re: Unspecific nuclear stain
« Reply #2 on: March 08, 2010, 10:14:22 AM »
Have you informed the antibody vendor about this unspecific staining? Sometimes ab have unwanted reactions and are not as specific as they are described. Then a change would be recommended.
gula

Offline Sona

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Re: Unspecific nuclear stain
« Reply #3 on: July 02, 2010, 12:55:56 PM »
Appologies for the delay.... Thanks a lot for your suggestions. I almost gave up vinculin staining. Unfortunately, the higher dilutions didnt work :( I will contact the company asap.

Offline Yao

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Re: Unspecific nuclear stain
« Reply #4 on: July 05, 2010, 06:40:50 AM »
is it so important? if it doesn't affect your story, some unspecific staining is OK, although not perfect.

Re: Unspecific nuclear stain
« Reply #4 on: July 05, 2010, 06:40:50 AM »