Bio-Rad Aurum™ Total RNA Fatty and Fibrous Tissue Kit - Manual (Page 19)

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Warning: Processing larger amounts of starting material may lead to column
clogging and reduced RNA purity. It is crucial that the appropriate amount of
starting material be used. For samples that are known to be rich in RNA, it is
highly recommended that less than the maximum amount of starting material
be used so that the binding capacity of the column is not exceeded. In
addition, complete disruption and homogenization of the starting material is
critical to prevent column clogging and reduced RNA yields.
2. Disrupt and homogenize the sample. Below are recommended
procedures for disruption and homogenization.
Note: Incomplete disruption will clog the column in subsequent steps and
result in reduced yields of total RNA.
Fresh Tissue
Fresh tissues can be processed in PureZOL™ immediately after dissection.
Alternatively, freshly dissected tissue can be immediately frozen in liquid
nitrogen and processed using instructions for frozen tissue. Transfer up to
100 mg of freshly dissected tissue into a 2.0 ml microcentrifuge tube and
add 1 ml of PureZOL. Disrupt the sample for 30–60 sec using a rotor-stator
homogenizer or a bead mill homogenizer (refer to manufacturer instructions
for more details). Although not as effective, passing the tissue sample
through an 18-gauge needle and syringe can be used for sample
disruption if a homogenizer is not available. Pass the sample through the
needle and syringe until no more solid tissue is left in the lysate. The
sample volume should not exceed 10% of the volume of PureZOL used for
disruption. Proceed to step 3.
Frozen Tissue
Grind up the frozen tissues to fine powder with a mortar and pestle under
liquid nitrogen. Avoid thawing the sample by periodically adding liquid
nitrogen to the mortar. Weigh up to 100 mg of tissue and transfer the
sample into a 2.0 ml microcentrifuge tube. Add 1 ml of PureZOL and
disrupt for 30–60 sec using a rotor-stator homogenizer or a bead mill
homogenizer (refer to manufacturer's instructions for more details).
Although not as effective, passing the tissue sample through an 18-gauge
needle and syringe can be used for sample disruption if a homogenizer is
not available. Pass the sample through the needle and syringe until no
more solid tissue is left in the lysate. The sample volume should not exceed
10% of the volume of PureZOL used for disruption. Proceed to step 3.
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