[Histonet] ISH preparation

[Emily Brown]

That may be what you have to do, as the first day of ISH needs to be RNase
free!
When I do them, it is 10 slides at a time and we have dishes and a lab
bench set aside for RNase free procedures only.  Obviously this is not
feasible in all workplaces!
I would just think about it logically.  Use gloves when you can, always.
 Can you keep the hybridizing part RNase free? I don't know what machines
you use for this--we do process the slides manually in glass dishes and
then hybridize in a slide container and oven kept for RNase free purposes.
 After hybridization, you don't really need to be too careful.

Now I'm wondering if there's some sort of magical machine that does ISH!

Emily

"By bitching and bitching and bitching, they could exhaust the drama of
their own horror stories. Grow bored. Only then could they accept a new
story for their lives. Move forward."

-Chuck Palahniuk, "Haunted"


On Mon, Jun 23, 2014 at 2:06 PM, <Jennifer.Johnson <@t> genzyme.com> wrote:

> Hi Folks!
> I was hoping to get a little input about preparing samples for ISH.
> We currently RNAse-away our tools at the time of collection, toss the
> tissues into formalin, process normally and then cut them using RNAse away
> cleaned microtomes/forceps on a bath of DEPC treated water (in a cleaned
> water bath).
>
> How do other labs prepare paraffin embedded samples?
>
> Do you use special reagents - formalin?  Do you clean the processor and
> use treated reagents there too?  Do you wash the slides in DEPC treated
> water?
>
> I am trying to figure out how far we need to go here.  We are a high
> throughput lab and we deal with all kinds of tissues and I don't want to
> have to take down a processor and keep it RNAse free if I don't have to!
> I will run the experiments and test different methods, but I was hoping
> some of you could share your wisdom and give me a place to start.
> Thanks,
> JJ
>
> Jennifer Johnson
> Staff Scientist
> Genzyme Corp.
> Department of Pathology
> 5 Mountain Road
> Framingham, MA 01701-9322
> Phn - 508-271-3610
> Fax - 508-872-9080
>
>
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