LaMotte EZ-3-0034 User Manual

ColiQuant EZ

Mott

Code 3-0034

Quantity Contents

10 bottles Coliscan Easygel® Easygel®, 20 mL

10 Petri dishes, pretreated, with lids, sterile

10 Collection bottles, sterile

10 Droppers, 3 mL, sterile

1 ColiQuant EZ Manual

5 ColiQuant EZ Colony Color Guide

Coliscan Easygel is a registered trademark of Micrology Laboratories LLC

Storage Con di tions

Coliscan Easygel should be placed in the freezer (2 – 6°C) as soon as it is received. Coliscan Easygel can be refrozen if it has been thawed and not been used. Coliscan Easygel should be thawed at room temperature overnight. Bottles may also be thawed for same day use by standing the bottles in warm water until the contents become liquid. Coliscan Easygel will keep for 6 weeks in the refrigerator or 12 months in the freezer.

Safety

• Coliscan Easygel is non-toxic. Additional information for all LaMotte reagents is available in the United States, Canada, Puerto Rico, and the US Virgin Islands from Chem-Tel by calling 1-800-255-3924. For other areas, call 813-248-0585 collect to contact Chem-Tel’s International access number.

• Read the entire manual before performing any tests.

• Be sure that students understand the danger of treating reagents casually or endangering others through “horseplay”.

• Wear safety glasses and protective gloves when appropriate.

• Wash hands or use a hand sanitizer after performing tests. Avoid placing hands in contact with eyes or mouth. Do not eat or drink or apply cosmetics during testing or clean up.

• Wipe tabletops and work areas with disinfectant, such as a 1:10 dilution of household bleach, after each use.

• Follow the general safety guidelines for your school.

• Ensure a safe field-testing experience by using the following: safety glasses for

Smaller diameter

Contains Clear

Media Film

Lid

Bottom

each student, clean pail or bucket for washing hands, jug of clean water for washing hands, soap (biodegradable if possible), towels, protective gloves, eye wash bottle, first aid kit.

• Keep used Petri dishes taped shut and out of the reach of small children and animals.

• Use one of the recommended disposal methods in procedure.

Coliscan Easygel EZ Method

In order for coliforms to ferment lactose, they must produce certain enzymes, which can be identified and used to verify the presence of coliforms and E. coli. General coliforms produce the enzyme galactosidase from lactose fermentation whereas, E. coli produces the enzyme glucuronidase, in addition to galactosidase Coliscan Easygel incorporates two special chromogenic substrates, which are acted upon by the enzymes galactosidase and glucuronidase, to produce pigments of different colors. General coliforms will produce the enzyme galactosidase and the colonies that will grow will be a pink color. E. coli will produce galactosidase and glucuronidase, and will grow as blue/purple colonies. The sum of the E. coli and general coliforms is the total coliform number.

Col lecting a Wa ter Sam ple

1. Remove the cap of a collection bottle. Do not touch the neck of the bottle or the

inside of the cap.

2. If you are testing stream water, face upstream while holding the bottle 5 to 6 inches below the surface.

3. When the bottle has filled, replace the cap.

4. The test can be done at the site or water samples may be collected in the sterile

collection bottle and transported back to the testing site. Water samples kept for more than one hour before testing should be kept on ice or refrigerated.

5. Do not reuse the sample collection bottle.

Pro ce dure

1. Use a permanent marker or wax pencil to label a Petri

dish and a bottle of Coliscan Easygel with the sample information. Tape the dish shut with two pieces of clear tape. Use one piece of tape as the hinge and the other as a latch.

2. Unwrap a sterile dropper. Handle the dropper by the bulb end. Do not touch the tip of the dropper with anything but the water sample.

3. Thoroughly mix the water sample. Squeeze the bulb of the dropper. Insert the tip of the dropper 2 – 3 inches below the surface of the water. Slowly release the

bulb to draw the water into the dropper. Carefully squeeze the bulb to expel excess water until the dropper is filled to the desired sample volume.

Use these gen eral guide lines for de ter min ing sam ple vol ume to be tested:

• If the coliform level is unknown, begin with 1 mL. This sample size is

recommended for sources such as rivers, ponds, streams, lakes, ditches, etc. These samples often have moderate amounts of coliforms. If counts are too low, increase the sample size.

• If counts are too high for water samples with extremely high coliform levels, the

sample size can be reduced to 0.25, 0.5 or 0.75 mL.

• If the sample is suspected of having very low levels of coliform, such as

drinking water, use a 5.0 mL sample size.

4. Remove the cap from one bottle of Coliscan Easygel. Do not touch the neck of the bottle. Do not touch the inside of the cap or put the cap down where it could become contaminated.

5. Add the water sample from the dropper to the bottle of Coliscan Easygel. Hold the bottle and dropper at an angle to prevent spills. Carefully squirt the sample into the bottle.

6. Cap the bottle and swirl to distribute the contents. Do not shake or create air bubbles.

Note: After the addition of the water sample to the Coliscan Easygel bottle, samples that can not be plated within 10 minutes should be kept on ice or refrigerated. Keep the bottles out of direct sunlight.

7. Remove the tape from one side of the dish and lift up the lid. Pour the mixture into the Petri dish bottom.

8. Replace the lid on the Petri dish. Carefully swirl the poured dish until the bottom of the dish is completely covered with liquid. Do not splash the liquid over the side of the dish or onto the lid. Tape the dish shut.

9. While the contents of the dish are still liquid, place the dish with the lid side up in a warm level spot or in an incubator. The liquid will solidify in about 30 - 45 minutes.

10. When the medium has solidified. Turn the dish upside down (with the lid side down). Incubate Petri dish for 48-72 hours at room temperature (68-74°F or 20 – 24°C) or 24-48 hours in a preheated incubator at 95°F (35-37°C). See Incubation Hints. Note: A liquid may form during incubation. Avoid spilling the liquid. If liquid spills, observe safety procedures; disinfect the spill area and wash hands thoroughly.

11. Colonies will begin to appear after 24 hours. The best time to count colonies is

Total Coliform Colonies per mL (CFU/mL)

# Colonies

Amount of sample used

Total Coliform Colonies per 100mL (CFU/100mL)

# Colonies x 100

Amount of sample used

E. Coli or Fecal Coliform Colonies per mL (CFU/mL)

# Colonies

Amount of sample used

E. Coli or Fecal Coliform Colonies per 100mL (CFU/100mL)

# Colonies x 100

Amount of sample used

after 30 hours of incubation. Colonies should not be counted after 72 hours. Count the colonies with the dish upside down. Colonies are normally reported as Colony Forming Units (CFU) per 100 milliliter of water. Colonies may appear inside the medium. Count all colonies growing on the surface and within the medium.

12. To determine the result as E. coli or Fecal Coliform count the dark blue to purple colonies. Disregard any light-blue, blue-green or white colonies. Refer to

the ColiQuant EZ Colony Color Guide.

• To report the result in terms of E. coli or Fecal Coliform per mL of water: Divide

the number of colonies by the sample size

• To report the results in terms of E. coli or Fecal Coliform per 100 mL of water

(Most state agencies use this measurement): Multiply the number of colonies by 100. Then divide by the sample size.

13. To determine the result as Total Coliforms (E. coli + non-fecal coliforms) count the number of pink and purple colonies.

• To report the result in terms of Total Coliforms per mL of water: Divide the

number of colonies by the sample size

• To report the results in terms of Total Coliforms per 100 mL of water: Multiply

the number of colonies by 100. Then divide by the sample size.

14. If there are more than 300 CFU (colonies) of one color, the result should be recorded as Too Numerous To Count (TNTC). If the E. coli purple colony count is less than 300, even though there are more than 300 total colonies of all colors and types, the purple colonies can be counted and a result given for E. coli. Record TNTC for coliforms or non-coliforms that are present at greater than 300 CFU.

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