Ohio EPA Drinking Water Certification

MMO-MUG Reagents

Colilert, Colilert-18, Colisure

How should MMO-MUG reagents be stored?

In the dark, from 2-6°C

How should chemical reagents be stored?

Manufacturer’s recommendations

How should dehydrated media be stored?

Manufacturer’s recommendations

How should prepared media be stored?

In the fridge

How should pH buffers be stored?

At room temperature

When do the MMO-MUG reagents expire?

Manufacturer’s expiration date

When do the chemical reagents expire?

Manufacturer’s expiration date

When does dehydrated media expire?

1 year after opening if stored in desiccator

When does the 10% sodium thiosulfate expire?

Manufacturer’s expiration date

When do the media performance check cultures expire?

3 months refrigerated

When do the pH buffers expire?

Manufacturer’s expiration date

Explain the procedure for the total coliform/E. coli positive quality control check

A 100 mL sterile bottle of water is to be inoculated with a known E. coli positive, reagent added, and incubated for the required time at 35.0 ± 0.5°C. The test needs to result in a positive.

How often does the total coliform/E. coli positive quality control check need to be performed?

Once per month per analyst

Explain the procedure for the sample bottle sterility quality control check

Add approximately 25 mL of prepared sterile Tryptic Soy Broth (TSB) per bottle tested. Mix to expose the entire interior of the bottle to the growth media.

Incubated at 35.0 ± 0.5°C and check after 24 hours for growth. Growth is indicated by any turbidity.

If any growth occurs, repeat the sterility check. If it remains positive, contact the manufacturer and do not use any bottles from the affected case.

How often should the sample bottle sterility quality control check be performed?

1% per lot received (maximum of 4 per lot)

Explain the procedure for the sample bottle fluorescence quality control check

Put an unused sample bottle under the UV light and check for auto-fluorescence

How often should the sample bottle fluorescence quality control check be performed?

1% per lot received (maximum of 4 per lot)

Explain the procedure for the media performance check

Inoculate one sterile 25 mL test tube of TSB with a known coliform culture. A second 25 mL tube of TSB serves as a control blank.

Incubate both tubes for 24 hours at 35.0 ± 0.5°C.

The inoculated tube must show cloudy growth while the control blank must not show cloudy growth.

Do not use media if growth is not indicated in the inoculated tube.

How often should the media performance quality control check be performed?

Once per batch

Explain the procedure for the MMO-MUG reagent check

Aseptically fill three test vessels with 100 mL sterile reagent water. Add a packet of the MMO-MUG reagent being checked to each test vessel and mix thoroughly to dissolve.

Inoculate each test vessel with one of three known cultures (Escherichia coli, Klebsiella

pneumoniae or Pseudomonas aeruginosa) and label each bottle with the bacterium used.

Incubate all test vessels at 35.0 ± 0.5°C for the required amount of time.

After incubation, check that the E. coli is positive for both total coliform and E coli, the Klebsiella pneumoniae is positive for total coliform and negative for E. coli, and the Pseudomonas aeruginosa is negative for both total coliform and E. coli.

Explain how to calibrate the equipment timers

Use an accurate stopwatch to time how long the equipment times. Note the correction and insure that it is less than 1 minute and 30 seconds for a 15 minute autoclave cycle and less than 5 minutes for a 45 minute cycle.

How often should the equipment timers calibration be performed?

Once every three months

Explain how to calibrate a pH meter

Use the 4.0, 7.0, and 10.0 to calibrate the pH meter. The linearity must be recorded each time the meter is calibrated; acceptance limits are 95 to 105% or -56 to -62 mV.

How often does the pH meter need to be calibrated?

Prior to use

Explain the balance calibration quality control check

All balances need an annual service contract and must be checked with certified weights. At least three weights that bracket the range of weights normally used in the lab. Place each mass on the balance and record the weight (reference weight). Add a test load (0.1 g) and record the reference weight plus the test load weight.

To what readability do the top loading balances have to be?

0.1 g

How often does the balance calibration check need to be performed?

Prior to use

What is needed for the refrigerator record?

The temperature needs to be between 2-6°C, the date, and the analyst’s initials.

How often does the refrigerator temperature need to be recorded?

Daily

What is needed for the incubator/water bath record?

The temperature needs to be between 35.0 ± 0.5°C, the date, time, and analyst’s initials.

How often does the incubator/water bath temperature need to be recorded?

Twice daily, per shelf (if applicable)

Explain the autoclave sterility check

Autoclave the biological indicator ampule for 15 minutes at 121°C. Allow to cool for 10 minutes, then crack the ampule and get a non-autoclaved ampule and also crack it. Incubate sterilized and unsterilized (control) ampules at 55 - 60°C for 24 hours. Growth is evident by a color change per manufacturer’s instructions. If color change occurs, corrective action for the autoclave is required.

How often is the autoclave sterility check performed?

Once every three months

Explain the alternative autoclave sterility check

Fill an Erlenmeyer flask with 25 to 50 mL of TSB/BHI, inoculate with a known

coliform culture, cover flask opening with aluminum foil and incubate at 35.0 ± 0.5°C for 24

hours. After incubation, when TSB/BHI shows growth, autoclave at 119 - 121°C for 12 to 15

minutes on slow exhaust. Allow to cool to room temperature. Fill a test vessel with

approximately 25 mL of TSB/BHI and inoculate the TSB/BHI with the “sterilized” culture from

the Erlenmeyer flask. Incubate test vessel at 35.0 ± 0.5°C for 24 hours. After the 24-hour

incubation period, remove the test vessel from the incubator. The inoculated test vessel must

not show growth. If growth is present in the inoculated test vessel, corrective action for the

autoclave is required.

How are positive samples to be disposed of?

All positives must be autoclaved before disposal

Explain the presence/absence test procedure

Shake the samples, pour into larger vessel if needed, pour off volume to 100 mL, tap down reagent packets, add to sample, shake to homogenize, if it is colilert-18 incubate in the water batch for 20 minutes, incubate for required time at 35.0 ± 0.5°C, check for color change and fluorescence

Explain the quantitray test procedure

Shake the samples, pour into larger vessel if needed, pour off volume to 100 mL, tap down reagent packets, add to sample, shake to homogenize, pour into quantitray, seal, incubate for required time at 35.0 ± 0.5°C, check for color change and fluorescence

When should you use colisure instead of colilert?

If the sample has a yellow background color