Re: cyclin D1

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From:"ANN MARUSKA" <amarusk1@FAIRVIEW.ORG> (by way of histonet)
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I have to agree with you....BioCare's DR1BC2 works the best.....or I should
say DID work the best.  They have improved their DCS-6 (those guys must
always be cookin' in the kitchen!) and it really is great.  You still have
to work hard to get it to work, but it finally gives the stain you want.  I
use the DCS-6 predilute, incubate at 3hrs room temp. and then run on the
NexES with an enhanced kit (and BioCare's link).  My pretreatment is
decloak for 3 min. in EDTA.  Try it - you'll like it!!

Ann Maruska
Fairview-University Med Ctr.
Mpls. MN  55454

>>> Phyllis Davie <> 04/28/00 11:59AM >>>
     I'll add a dissenting opinion here.  I have tried 5 different clones
of cyclin D1, including the Novocastra one (clone  P2D11F11), but my
favorite one is clone DR1BC2 (I purchase it from Biocare Medical, phone
800-799-9499, cat# CM-090D, but it may be available from other sources as
well.)  HOWEVER, I must add that ALL of the clones I tried (DR1BC2,
P2D11F11, DCS-6, 5D4, and AM29) do work, but that NONE of them is what I
would call a stellar performer, or an easy antibody to work with.  I
prefer the DR1BC2 clone because it gives me the most consistent
performance, and generally the strongest signal of any I have tried.
     For the rest, I do epitope retrieval in a steamer for 40 minutes in
pH 10 Tris, followed by a 20 minute cooldown.  I incubate in my primary
antibody @ 1:50 for 1 hour at room temperature, followed by secondary,
tertiary, DAB, etc. as per usual.
     And finally, I agree 100% with Zenobia Haffajee's final line, "Give
it a try, GOOD LUCK....wave your magic IHC wand before you start."

Phyllis Davie

>Hi Deb
>We use the CyclinD1-GM from Novocastra, found it to be the best one so far.
>CyclinD-1 is a tricky unstable antibody....likes to give problems. So donot
>feel alone; going bald, well donot do it for the cyclind-1!
> Yes, sometimes it works great and at other times a bit iffy..
>We microwave in an alkaline pH ( Trisbase at pH 10) for 20 mins on high in
>a domestic microwave, allow to cool for 20 mins. Incubate with primary at a
>1:25 dilution overnight at rt and use the ABC detection ; biotinylated goat
>anti-mouse 1:200 for 30 mins and Streptavidin peroxidase at 1:500 for 30
>mins at rt. DAB as the chromagen. Give it a try, GOOD LUCK....wave your
>magic IHC wand before you start.
>Zenobia Haffajee
>HAPS, Newcastle, Australia
>At 07:36 PM 11/04/00 -0500, you wrote:
>>          Hi,
>>               Would anyone who has Cyclin D-1 working reliably---and often
>>please share all your info--including products used, times for all process,
>>kit used etc-- and where is everyone finding mantle cell lymphoma for
>>controls as that is what we are insterested in staining for????
>>                     Thanks-Deb   I'm tearing my hair out on this one!

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