Antibody-Based Technologies => Antibodies Review => Antibody B => Topic started by: vivfly on February 10, 2006, 07:35:38 AM

Title: Did anybody use primary AB-anti-BrdU (VP-B209) from Vector?
Post by: vivfly on February 10, 2006, 07:35:38 AM
Hi, dear all: did anybody ever use the primary AB (anti-BrdU) from Vector lab? (VP-B209). I'm recently trying this AB for frozen sections. However, it never worked out. If you did successfully before,could you please share with me your experience? Thanks a lot!!!
Title: Did anybody use primary AB-anti-BrdU (VP-B209) from Vector?
Post by: richard03 on February 11, 2006, 10:51:22 AM
You got no staining, background staining or else?

How did you pretreat the sections? Can you post the protocol you were using?
Title: Did anybody use primary AB-anti-BrdU (VP-B209) from Vector?
Post by: vivfly on March 08, 2006, 03:57:44 AM
Hi, richard03, thanks for your attention. recently i got some positive results by using high concentraion of the primary AB. However, it's accompanied by high background also. certainly, this high background could be improved by reducing the concentration of secondary AB (biotinylated anti-mouse, rabbit), but then it leads to less specific staining again. I would like to post my protocol briefly for discussion: Wash by PBS--fix by acetone(4degree) 15min--0.5%H2O2/MeOH incubation--permeabilize by 0.1%Triton X-100 15min--2N HCL incubation 30min (37degree)--0.5M NaHCO3 10min--50mM Tris buffer 10min--1%BSA/3%HS 30min--primary AB 1:200 2h RT--Secondary AB (biotinylated anti-mouse, rabbit)  1:1000 1h RT--ABC kit--develop.
Title: Did anybody use primary AB-anti-BrdU (VP-B209) from Vector?
Post by: richard03 on March 08, 2006, 07:44:10 AM
Is this mouse tissue?
Title: Did anybody use primary AB-anti-BrdU (VP-B209) from Vector?
Post by: vivfly on March 08, 2006, 08:34:18 AM
Quote from: "richard03"
Is this mouse tissue?


yes, it is. Did you ever use this antibody before? Any suggestions?Thanks
Title: Did anybody use primary AB-anti-BrdU (VP-B209) from Vector?
Post by: richard03 on March 08, 2006, 06:25:43 PM
That is the problem.

Read this protocol, follow it strictly and you should be fine.

http://www.ihcworld.com/_protocols/general_IHC/immuno_mom.htm
Title: Did anybody use primary AB-anti-BrdU (VP-B209) from Vector?
Post by: vivfly on March 09, 2006, 04:48:37 AM
Thanks a lot! I would like to try it!
Title: anti-mouse BrdU
Post by: madeleine huey on October 09, 2006, 03:44:06 PM
Hello,
You can try Zymed/Invitrogen's BrdU kit for mouse tissues.  I used it before, and no background what so ever (there's a special blocking solution in this kit to help reduced the background.  I tried different condition, and still find their kit is better than mine).
Good luck!
M