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Author Topic: 8 OHDG troubles  (Read 3261 times)

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Offline labgal

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8 OHDG troubles
« on: February 16, 2009, 11:12:44 AM »
Hello,

i was wondering if anyone had a good working protocol for 8 ohdg in human, we are trying to stain human prostate biopsies and we are getting a ton of non specific stain and background. We have the protocol working beautifully in rat but its really ugly in human.

currently our protocol is:

antigen retrieval in .62 ZnSo4 in a steamer for 20 mins
2HCL incubation at 37degrees
Block with a normal horse serum at 37degrees
Avidin/Biotin block (vector)
Primary 1:60 over night
Incubate with Biotinylated antimouse IGG (vector vectastain ABC kit)
Preoxidase block
Incubate with Avidin-biotin complex (vector vectastain ABC Kit)
DAB 2 mins


Does anyone have any idea's or suggestions? Please help if you can.

8 OHDG troubles
« on: February 16, 2009, 11:12:44 AM »

Offline richard03

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Re: 8 OHDG troubles
« Reply #1 on: February 17, 2009, 07:36:47 AM »
You will need a peroxidase blocking step using H2O2.

Offline labgal

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Re: 8 OHDG troubles
« Reply #2 on: February 17, 2009, 11:19:51 AM »
i do one the second day so i do not damage the dna before i put the primary antibody on

Re: 8 OHDG troubles
« Reply #2 on: February 17, 2009, 11:19:51 AM »