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Membrane Filtration Protocol

The membrane filtration protocol is used to isolate bacteria from the water samples
collected. Essentially, the water is filtered through a cellulose membrane (pore size
0.2µm) which should capture the maority of bacteria from the sample. !articularly
turbid samples re"uire pre#filtration with a larger membrane (pore size 0.$% µm).
&oth of these protocols are completed using the 'icrosart ( e.et )artorius membrane
filtration unit.
*ne litre of water from each site will be filtered using this unit.
!rotocol
+. ,sing -') clean the lab bench
2. )et up the membrane filtration unit i.e. plug in, turn on and ma.e sure the
outflow is in a suitable location
/. !osition the sterile filtration apparatus so that the 0al0es are in the horizontal
off position.
$. 1emo0e the funnel from the filtration unit.
%. 2lean forceps (ethanol or flame), then use the forceps to remo0e a filter
(0.2µm or 0.$%µm) from the pac.et.
3. !lace the filter, shiny side up, on the porous dis. of the filtration unit.
4. ,sing a syringe put a few drops of distilled water onto the filter to secure it in
position. The ideal positioning for a filter is5 central o0er the porous dis. with
a /mm diameter boarder
6. 1inse the funnel with distilled water twice the clamp it into place
7. !our the sample (in 2%0ml loads) into the funnel and switch on the 0acuum
+0. 8hen filtration is complete, remo0e the funnel and ta.e the membrane from
the based using sterile forceps
++. -f the filter is being used for direct growth on agar 9 place the membrane onto
the agar with the shiny side in contact with the medium (i.e. it is in0erted)
+2. -f the filter is being used for further in0estigate (i.e. :;< e=traction etc) place
in a small (%0mm), labelled !etri dish with the shiny side up and store in cold
storage ($>2).
)amples? 'embrane @iltration
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