Re: Re: [Histonet] subbing slides
tifei <@t> foxmail.com
Tue Dec 30 20:17:18 CST 2008
If you are talking about very thick brain sections, I have used 40-50 um sections only.
However my collegues noticed that 0.25% gel coated slides are much better than 0.5% gel coated slides.
We are performing BrdU staining-including the step of citrate buffer heat at 95 degree for 30 min.
The 0.25% gel stands out from this test.
发件人： Susan Bachus
发送时间： 2008-12-31 07:42:23
收件人： Neil Fournier; histonet <@t> lists.utsouthwestern.edu
主题： Re: [Histonet] subbing slides
Well, the attachment seems to have been deleted from what I sent, so here it
is pasted below:
Gelatin-Subbed slides WEAR GLOVES! (not to protect your hands, but to
protect the slides from RNase from your skin)
a.. Place slides in racks and soak in hot tap water and soap (e.g.
dishwashing soap, or Alconox) for at least 1 hr, but not more than a few
hours (as soap could leave residue).
b.. Rinse thoroughly in hot tap water until suds are completely gone.
c.. Let sit in 80% ethanol for at least one hr. This solution can be
re-used until it turns cloudy. Slides can be left overnight in this
d.. Rinse in a few changes of deionized water.
e.. Subbing solution: Dissolve 3.0 g of gelatin (300 bloom swine) in 150
ml deionized H2O by stirring while heating to 50 degrees C. Add 450 ml
deionized water to cool, and then dissolve 0.3 g CrK(SO4)2*12H2O ("chrom
alum") in the solution.
f.. Filter this solution through Whatman Type I filter paper to remove any
air bubbles. This solution is only good for the few hrs it will take for
this batch of slides, as it will quickly solidify.
g.. Dip the slides into the subbing solution, drain the slides onto a
paper towel and allow to air dry for 1 hr.
h.. Dip the slides into the subbing solution again. Drain and cover
loosely with bench paper and allow to dry for a few hours.
i.. When thoroughly dry, heat in oven at ~ 50 degrees C for a few hours,
then store the slides in slide boxes in zip-lock bags. As long as they are
kept dry, they should be good for years.
----- Original Message -----
From: "Susan Bachus" <susanbachus <@t> verizon.net>
To: "Neil Fournier" <nfournier <@t> sasktel.net>;
<histonet <@t> lists.utsouthwestern.edu>
Sent: Tuesday, December 30, 2008 6:31 PM
Subject: Re: [Histonet] subbing slides
> This always works like a charm for us and costs ~10x less than buying
> pre-subbed or electrostatic or whatever fancy expensive store-bought
> Have fun! Susan
> ----- Original Message -----
> From: "Neil Fournier" <nfournier <@t> sasktel.net>
> To: <histonet <@t> lists.utsouthwestern.edu>
> Sent: Tuesday, December 30, 2008 6:21 PM
> Subject: [Histonet] subbing slides
>> Hi Everyone,
>> Could someone share with me their procedure for cleaning and subbing
>> slides with gelantin? I am trying to work out a concentration of gelantin
>> and chromium potassium sulphate that will be optimal for adhering fairly
>> thick rat brain sections (30 to 120 um range depending on applications)
>> Much appreciated
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