CryptTest™ membrane filter cartridge

> CrypTest membrane filter cartridge

The CrypTest Cartridge captures protozoan cysts and oocysts from sample water pumped through the disposable cartridge contained in a re-usable housing. The CrypTest contains a 1.0µm pore size Nuclepore® polycarbonate track etch membrane.

Protozoan cysts and oocysts are recovered from the membrane cartridge by using a simple backwashing procedure. The eluant, containing cysts and oocysts, is decanted from the housing into a collection vessel.

Further concentration, purification and separation by centrifugation, immunomagnetic separation or flow cytometry/cell sorting is then performed. Enumeration and confirmation of the cysts and oocysts is done using epifluorescence and DIC procedures.

Features and Benefits

• Nuclepore polycarbonate track etch membrane provides total surface capture
• Simple backwashing procedure yields consistently high recovery
• Convenient for field sampling-sample water pumped through disposable cartridge contained in durable, reusable housing
• Simple to use

Materials and Equipment

• CrypTest Cartridge - Catalogue Number 610064
• Cartridge housing, AMETEK 5" clear polycarbonate - Catalogue Number 71503
• 1/2" male pipe threads with male hose connectors (2)
• Laboratory tubing, Tygon® formula R-3603, or equivalent, to which cysts/oocysts will not adhere
• Flow control valve - 2 litres/minute (0.5 gallons/minute)
• Water flow meter
• Peristaltic pump
• Ultrasonic bath

Elution Buffer

• NaCl - 8.0g
• KH2PO4 - 0.2g
• Na2HPO4 (12H2O) - 2.9g
• KCl - 0.2g
• Sodium lauryl sulphate (SDS) - 0.2g
• Tween 80 - 0.2ml
• Antifoam A* - 0.02ml
• Adjust volume to 1 litre with reagent water.
• Adjust pH to 7.4 with 1 N NaOH or HCl.

*Sigma Chemical Co - Catalogue Number A5758.

> Sampling from Carboy

Sample Filtration

Key
A - Sample
B - Filter Inlet
C - Filter Outlet
D - Peristaltic Pump
E - Flowmeter
F - To Waste

1. Clean and rinse cartridge housing
2. Attach male pipe threads to inlet and outlet of cartridge housing cap.
3. Attach new or well-cleaned tubing to male hose connectors on inlet and outlet of cartridge housing. Secure both with band clamps.
4. Install the cartridge filter in the housing.
5. When filtering sample water from a carboy, thread the outlet tubing through the peristaltic pump and immerse the open end of the inlet tubing in the carboy. Alternatively attach open end of inlet tubing directly to feed line.
6 Turn on pump and adjust the flow rate to 2 litres/min.
7 Turn off the pump when the desired sample volume has been filtered.

> Sampling from Feed Line

Key
A - Sample Tap
B - Pressure Regulator & Gauge
C - Filter in Housing
D - Flow Totalizer
E - Flow Control Valve
F - Drain Hose

Backwashing the Cartridge to Elute Cysts/Oocysts

1 Detach tubing from housing inlet and decant any water remaining in the inlet side of the housing into a 1 litre sample bottle.
2 Attach the housing outlet to the discharge tubing of the peristaltic pump. Place the free end of this tubing into a beaker of elution buffer.
3 Start the pump and run at approximately 400ml/min until the filter outlet core and the housing inlet cavity are filled (approximately 300ml).
4 Cap the housing inlet and outlet and place the housing in the ultrasonic bath for 2 minutes.
5 Remove the inlet cap and decant the liquid from the inlet side into the collection bottle.
6 Repeat steps 3, 4 and 5 and then proceed to step 7.
7 Connect housing outlet to regulated compressed air source and gradually increase pressure to 5 to 10psi as necessary to drive remaining elution buffer from the outlet side to the inlet side and into the collection bottle.
After backwashing the cartridge and collecting the eluate, further purification, detection and enumeration of cysts/oocysts may be performed using EPA recommended methods.<

> Backwash

Key
A - Eluting Solution
B - Peristaltic Pump
C - Filter Outlet
D - Filter Inlet

Cleaning the Cartridge Housing

The following procedure is recommended for cleaning the cartridge housing between samples.

1. Wash thoroughly with water and laboratory detergent, using a brush to clean all crevices.
2. Soak in 6% sodium hypochlorite solution for 30 minutes.
3. Rinse thoroughly with tap water.
4. Rinse with laboratory water.