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3.7 Tyndallisation of MMDs
In the event an autoclave or pressure cooker
is not available then the sterilisation of the
MMD containing the liquid MLSB media can
be carried out using a pan of boiling water.
This is a process called
Tyndallisation
.
When liquids are heated up to 100°C,
the heat will kill the bacterial cells but
the bacterial spores may survive.
Tyndallisation essentially consists of boiling
the substance for 15 minutes for three days
in a row. On the second day most of the
spores that survived the first day will have
germinated into bacterial cells. These cells
will be killed by the second day’s heating.
The third day kills bacterial cells from late-
germinating spores. During the waiting
periods over the three days, the substance
being sterilised is kept in warm, moist
conditions that are conducive to germination
of the spores. When the environment is
conducive to the formation of cells from
spores, the formation of spores from cells
does not occur.
The Tyndallisation procedure can be
summarised as follows:
•
DAY 1
: Place the MMD containing the
MLSB into a pan or pot of boiling water,
use a rack or stand if possible and try to
prevent the MMD coming into contact
with the base of the pan
• Boil for 15 minutes
• Leave the MLSB to cool and then stand
at room temperature for 24 hours
•
DAY 2
: Once again heat the MLSB
in a pan of boiling water for a further
15 minutes
• Leave the MLSB to cool and then stand
at room temperature for 24 hours
•
DAY 3
: Repeat the immersion in boiling
water for 15 minutes for a third and
final time
• The MLSB media should now be sterile
3.8 Storage of Liquid Media
The sterile MLSB should be stable for up
to 6 months if stored in a refrigerator. If no
refrigerator is available the media can be
stored for up to 3 months if kept in a dark
place away from extremes of heat and
moisture. However, if there are any signs of
contamination e.g. yellowing, cloudiness etc.,
then it has become contaminated. It should
be discarded and under no circumstances
used in a microbiological test.
3.9 Pre-Prepared Media
It is possible to use pre-prepared growth media
rather than prepare your own as described
previously. The main advantages these
options offer are that they save time and
reduce the amount of equipment required.
However it should be noted that they are
generally more expensive in terms of cost
per test, and their shelf-life is shorter.
This makes shipping to, and then using in,
remote locations more problematic. Some
of the pre-prepared media also have
specific storage requirements.
The most commonly used pre-prepared
media options are:
Ampoules
Sterile
Ampoules
containing 2ml of
dissolved media. Available in different
varieties for the testing of a wide range of
microorganisms. Simply unscrew the cap,
pour the media onto the absorbent pad
and discard the empty ampoule.
NutriDisks
For single use, a
Nutridisk
consists of a
sterile plastic petri dish which includes an
absorbent pad impregnated with dehydrated
growth media. This is re-hydrated using
sterile distilled water before use in the
microbiological test.
The
Nutridisks
are available in different
varieties for the testing of a wide range
of microorganisms.
3
Preparation
