Rig Filtration of SPOT 0m Water
Soak all small rig components in 10 % acid for a minimum of 30 minutes .
Rinse all components thoroughly using MilliQ ( hold disks on edges ) .
Place on rig once rinsed .
Rinse red tubes and large plastic Erlenmeyer flask with MilliQ .
Assemble pump to rig using red tubes and large plastic Erlenmeyer flask .
Attach tube to right side of pump for vacuum .
Right knob on pump controls vacuum , loosen all the way .
Remove filter top component and place upside down on bench .
Remove micropore forceps from ethanol and rinse well with MilliQ .
Hold in a new sterile Falcon tube during use .
Using micropore forceps remove 0.8μm backing disk from between blue disks and place on each rig filter .
Again using the micropore forceps remove 0.2μm filter disk from between blue disks and place carefully on top of each 0.8μm disk .
Make sure they are centered and do not have holes or rips .
Place top component back on to each filter and twist on .
Open each rig filter by turning orange knob to verticle position .
Using a serological pipet and sterile glass pipet tip remove desired amount from bucket and pipet directly onto disks .
There should be 6 filtrations / filters of 10ml and 1 each of 20ml , 30ml , 40ml , 50ml , 60ml and 100ml .
Once desired amount of sample water is placed on filter and knobs are vertical , turn on pump .
Adujst top right knob on pump so that the arrow on the top right guage is slightly above 0 .
Allow vacuum to work until most of the water has passed through the filters .
Once all water has passed through a filter , turn orange knob closed to the horizontal position .
Once all filtration has finished and all knobs are horizontal , turn off pump .
Twist off top component of rig filter and place upside down on bench .
Using the micropore forceps carefully remove only the top 0.2μm filter disk and place on a labeled TCBS Agar plate .
Use a sweeping or dragging motion to place filter flat on plate .
Repeat Step 12 for all 6 filters onto separate plates .
Repeat Steps 7 - 13 for remaining volumes of water .
The 0.8μm filter disks may remain on the filters , only new 0.2μm filter disks are needed each time .
Leave plates right side up at room temperature on bench for at least 10 minutes .
Then place upside down in drawer to grow for at least 24 hours .
