only yellow spots in microarray hybridization - troubleshooting (Oct/18/2006 )

Hi all,

I'm having trouble with my hybridisations. More spots than expected give signal and they are all yellow (even if I hybridise 2 different tissues against each other). Does anyone have an idea of why this happens? Any suggestions are welcome.

Thank's!

Hi,

If you didn't suspect your probes, ...
The problem could become of the slides, maybe, the washing during the spotting of your slide didn't work well, so you find a carry-over on all your slides and unfortunately the plates with your reporters of the spotting ( oligonucleotides, pcr product,... ) are contaminated !!
Another point of view is the hybridisation and the washing "post-hybridization" , indeed, if you didn't hybridize a the good temperature and if your washing didn't have the precise stringency, you didn't eliminate the non-specific hybridization, so you find a yellow slide.
Hope this help you..
regards

doc_lille

Hi all,

I'm having trouble with my hybridisations. More spots than expected give signal and they are all yellow (even if I hybridise 2 different tissues against each other). Does anyone have an idea of why this happens? Any suggestions are welcome.

Thank's!

hi annamaria

What are you doing? CGH, Gene expression, miRNA hybridisations?

are all the temperature setting correct in your protocol? your incubator etc are all right? maybe unspecific binding or something like that?