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Homogenising tissue, what do you rinse with? - (Dec/01/2009 )

Hi guys, rather basic question but... I'm currently looking into RNA expression in several major organs in my animal model and I'm using Trizol to isolate my RNA. During homogenisation, what should I use to rinse/wash the homogeniser to avoid cross contamination? I'm currently using 70% ethanol followed by a H2O wash.

-Xanthier-

Hi Xanthier,

Why don't you use autoclaved homogenizer tubes or mortar pestles? I autoclave them and wipe with 70% ethanol and RNaseZap before use

-gogreen-

I agree that autoclaving would be best. you could also soak them in dilute HCl followed by DEPC-treated water.

-gfischer-

gogreen on Dec 2 2009, 05:52 PM said:

Hi Xanthier,

Why don't you use autoclaved homogenizer tubes or mortar pestles? I autoclave them and wipe with 70% ethanol and RNaseZap before use


Hi, thanks for your suggestions. I'm not using the plastic tube homogenisers though :) I've only got access to a metallic blade one and unfortunately the design of the homogenizer means its rather difficult to take out the blades and autoclave them, (i'm not even sure if there are multiple blade bits!).

Looks like a smaller version of this: http://www.specialty-medicine.com/Images/homogenizer.jpg

-Xanthier-

Hi,

Have you tried running the homogenizer probe in 0.5% bleach and detergent for 20-30 sec.
Then few times with a few changes of water -> finally in clean water.
The bleach needs to be made fresh every day.

For years we used to do this routinely when processing tissues for diagnostic PCR for viruses with no cross contamination.

Now we use instead a Mixer Mill and 2 ml tubes containing 1.5 ml lysis buffer with 3 stainless steel ball bearings.
Saves time, next to zero risk of cross contamination and uses precious little sample.

-Falco79AD-