1. Remove sample (filter, cubitainer, or sample bottle) from refrigerator or 

 cooler just prior to shipping. Wrap sample in bubble wrap to prevent freezing

2. Place sample in cooler lined with two large plastic trash bags

3. Pack cooler with sealed ice bags or the equivalent number of gel ice packs 

 sufficient to maintain the sample between 1° and 10°C during shipment. Use 

 the amount of coolant appropriate for the season, geographic location, and 

 cooler-type used (follow laboratory recommendations). NEVER USE DRY ICE.

4. Knot and seal the two large plastic trash bags

5. Sign and date sample collection form, place in a water-tight bag in cooler

6. Seal and secure cooler with tape

7. Complete airbill and attach to cooler, retain shipper’s copy

8. Ship sample(s) to the laboratory via overnight delivery 

• Thoroughly clean all reused influent equipment prior to the capsule/filter

 housing/bulk water container with a warm detergent solution and scrubbing

 with a brush (new equipment should not need cleaning)

• Expose equipment to hypochlorite solution (Add 25 mL of 5% household 

 bleach for every 1 gallon of pH 7 water) for at least 30 minutes at 

 room temperature

• Thoroughly rinse equipment with reagent grade water that is free of analytes

 and interfering substances. Dry equipment in an area free of potential

 Cryptosporidium contamination

*Recommended Sampling Equipment Cleaning Procedure

(or alternatively follow the manufacturer’s cleaning instructions)

Not for sterilization of E. coli sample bottles

10-L Matrix Spike Sample

Follow instructions for collecting a 

10-L bulk water sample given on 

previous panel.

Matrix Spike Samples Greater 

than 10-L

Either ship the entire matrix spike volume 

as a bulk sample to the laboratory to be spiked and analyzed, OR filter all 

but 10 L of the matrix spike sample at your utility and collect the remaining 

10 L as a bulk sample. Chill both the filter and bulk sample as soon as possible

to between 1° and 10°C before shipment. Clearly label both the filter and

bulk sample as comprising 2 parts of a single sample. 

LT2 Rule Cryptosporidium & E. coli Sample Collection Recommendations: A Pocket Guide

E. coli 

Sample Collection

Packing and Shipping

Cryptosporidium

10-L Bulk Water Collection

Cryptosporidium 

Matrix Spike Sample Collection

Complete the sample collection form including utility and sample 

ID information, date, time, turbidity, pH, and temperature.

Thoroughly clean* all reused collection equipment and containers (new 

hoses, cubitainers, etc. should not need cleaning), and wear gloves 

whenever needed to prevent sample contamination throughout the 

collection procedure. Prior to sample collection, thoroughly flush stagnant

water and debris in the sample line until the temperature and turbidity

stabilize (approx. 2-3 mins). 

Send an extra 10 L bulk sample for spiking at the Cryptosporidium laboratory 

concurrent with the 1st and 21st routine monitoring sample collections and

again with every 20 samples if more than 40 source water samples are

collected. Coordinate the collection of matrix spike samples with your laboratory.

Matrix spike samples should be collected from the same location and at the

same time as the associated routine sample. The matrix spike sample and

routine sample should be the same volume. Label the cubitainer with the

following information using a waterproof pen: PWSID, facility name, and sample

collection date. Chill the sample as soon as possible to between 1° and 10°C

before shipment.

Pocket Guide

LT2 Rule 

Cryptosporidium

& 

E. coli

Sample Collection

Recommendations

1. Fill the 10-L cubitainer from a pressurized 

 sample tap or manually collect sample using 

 hose, pump, funnel, etc. from the location 

 described to the State.

 2. Once the sample is collected, 

 immediately place the cap on the

 cubitainer and tighten

 3. Chill the sample as soon as possible to

 between 1° and 10°C before shipment

Wear gloves whenever needed to prevent 

sample contamination throughout the 

collection procedure.

Complete the sample collection form 

including utility and sample ID 

information, date and time, etc.

1. Water taps used for sampling 

 should be free of aerators, hose 

 attachments, etc.

2. Prior to sample collection, thoroughly 

 flush stagnant water and debris 

 in the sample line until the

 temperature and turbidity

 stabilize (approx. 2-3 mins):

 DO NOT rinse the sample

 bottle

3. Record the turbidity, 

 pH, and temperature

4. Aseptically fill the sterile

 E. coli sample bottle 

 from a pressurized 

 sample tap or manually 

 collect sample using, 

 hose, pump, funnel, 

 etc. from the location 

 described to the State

5. Leave at least one inch of 

 head space in the E. coli sample 

 bottle, if possible. Collect at least 100 mL of sample for analysis

6. Immediately close the sample bottle

7. Chill sample as soon as possible to between 0° and 10°C until ready

 for immediate shipment

* Refer to the cleaning procedure located on the Packing and

 Shipping panel of this pocket guide
Cryptosporidium

Pressurized Sample Filtration

Cryptosporidium

Unpressurized Sample Filtration

EnvirochekTM or EnvirochekTM HV Capsules

 1. Install filter capsule

 2. Record start time and initial meter reading

 3. Slowly open sample tap, bleed out air, and adjust up to 

 the following maximum values:

 • 100 psig and 2 L/min 

 4. When at least 10 L has passed through the filter, turn off sample tap

 allowing pressure to decrease until water stops

 5. Record stop time and final meter reading

 6. Hold inlet pointing up, remove outlet tubing and allow water to drain

 7. Open bleed valve to speed the draining process and disconnect inlet tubing

 8. Seal the capsule with vinyl end caps, close the bleed valve, and place in 

 gallon-size zippered plastic bag

 9. Chill the sample as soon as possible to between 1° and 10°C

 until ready for shipment 

1. Install foam filter inside filter housing

2. Record start time and initial meter reading

3. Slowly open sample tap and adjust up to the following recommended values:

 • 72.5 psig and 3-4 L/min 

4. When at least 10 L has passed through the filter, turn off sample tap allowing 

 pressure to decrease until water stops

5. Record stop time and final meter reading

6. Hold inlet pointing up, remove the outlet tubing and allow water to drain

7. Disconnect the inlet tubing

8. Seal filter inside filter housing with rubber stoppers and place into a 

 gallon-size zippered plastic bag

9. Chill the sample as soon as possible to between 1° and 10°C

 until ready for shipment

 1. Install filter capsule and turn on pump 

 2. Record start time and initial meter reading

 3. Bleed out air and adjust flow rate up to a maximum of 2 L/min

 4. When at least 10 L has passed through the filter, turn off pump

 5. Record stop time and final meter reading

 6. Hold inlet pointing up, remove outlet tubing and allow water to drain

 7. Open bleed valve to speed the draining process and disconnect inlet tubing

 8. Seal the capsule with vinyl end caps, close the bleed valve, and place in 

 gallon-size zippered plastic bag

 9. Chill the sample as soon as possible to between 1° and 10°C

 until ready for shipment

Filta-Max® Filter

Pressure

regulator Flow totalizer Filta-Max®

filter housing

Flow control 

valve may be 

used in place 

of flow rate 

meter

Tap pressure >72.5 psig

Tap pressure 1-100 psig

Tap pressure >100 psig

Flow totalizer

Filta-Max® filter housing 

Tap pressure 7.5 - 72.5 psig

 

EnvirochekTM capsule 

1. Install foam filter inside filter housing and 

 turn on pump

2. Record start time and initial meter reading

3. Adjust flow rate up to a maximum of 3-4 L/min

4. When at least 10 L has passed through the filter, turn off pump

5. Record stop time and final meter reading

6. Hold inlet pointing up, remove outlet tubing and allow water to drain

7. Disconnect inlet tubing

8. Seal filter inside filter housing with rubber stoppers and place into a 

 gallon-size zippered plastic bag

9. Chill the sample as soon as possible to between 1° and 10°C until ready

 for shipment

EnvirochekTM or EnvirochekTM HV Capsules

Centrifugal pump Flow totalizer

Flow rate meter

with valve

Flow rate meter

with valve

Flow rate meter

with valve

Flow totalizer Pressure

regulator

Pressure

gauge

Pressure

gauge

EnvirochekTM

capsule Flow control 

valve may be 

used in place 

of flow rate 

meter

Office of Water (4601M)

EPA 817-K-06-002

April 2006 

This pocket guide is intended as an outline and supplement to the 

Source Water Monitoring Guidance Manual for Public Water Systems 

and the on-line Sample Collection Training Module. This guide is

intended to provide useful information and recommendations, but

does not substitute for the LT2 Rule.

The guidance manual is available for download and study at: 

http://www.epa.gov/safewater/disinfection/lt2/pdfs/guide_lt2_

swmonitoringguidance.pdf

The on-line training module can be accessed at: 

http://www.epa.gov/safewater/lt2/training/modules.html

Copies of this pocket guide can be ordered by using the document 

number listed below and contacting the National Service Center 

for Environmental Publications at 1-800-490-9198

Cryptosporidium

Preparation for Sample Filtration

Thoroughly clean* all reused collection equipment and containers (new tubing, 

fittings, etc. should not need cleaning), and wear gloves whenever needed to 

prevent sample contamination throughout the collection and filtration procedure. 

Before connecting the sampling unit to the tap, thoroughly flush stagnant water

and debris in the sample line until the temperature and turbidity stabilize

(approx. 2-3 mins).

Complete the sample collection form including utility and 

sample ID information, date, turbidity, pH and temperature.

Pressurized source

1. Determine sample line water pressure at the location described to the State

2. Connect assembled sampling unit, without capsule/foam filter, to sample tap

3. Flush the sampling unit for 2-3 minutes and test for leaks

Unpressurized source

1. Fill 30+ L carboy from a pressurized sample tap or manually collect the sample 

 using hose, pump, funnel, etc. from the location described to the State

2. Chill the full carboy to between 1° and 10°C if filtration is not performed 

 immediately after filling the carboy

3. Assemble sampling unit, without capsule/foam filter, and insert influent tube 

 into the center of the carboy

4. Turn on pump to flush the sampling unit for 2-3 minutes and test for leaks

* Refer to the cleaning procedure located on the Packing and Shipping panel

 of this pocket guide

LT2 Rule Cryptosporidium & E. coli Sample Collection Recommendations: A Pocket Guide

Flow control 

valve may be 

used in place of 

flow rate meter

Filta-Max® filter housing Flow totalizer

Peristaltic pump

Filta-Max® Filter

Flow rate meter with valve

Flow control 

valve may be 

used in place of 

flow rate meter

Flow control 

valve may be 

used in place of 

flow rate meter

Flow rate meter Flow totalizer with valve

EnvirochekTM capsule 

Flow control 

valve may be 

used in place of 

flow rate meter

Flow rate meter

with valve