Sampling Protocols

Surface Sampling

A. Swab Sampling

Materials Included:

Sterile swabs
Sterile, plastic test tubes
Sterile water in plastic test tubes
Sharpie Marker
Chain of Custody Sample Identification Form

Procedure :

Sterile materials should remain sterile (free of bacteria and mold) until you use them to collect a sample. Wearing gloves is helpful but not necessary. If visible mold is present or suspected, please use caution when collecting the sample; a face mask or other protective respiratory gear is recommended to minimize mold spore inhalation during collection.

1) Fill out the chain of custody form with all necessary information including descriptions of the sample location, material sample will be taken from, surface area tested, etc.

2) At the sample site, remove a swab, stick end first, from the package. There are two swabs in each package but only one will be needed per sample (the second one can be used for a second sample). Do not put the exposed swab down before or after you’ve collected the sample and hold the swab by the very end of the stick.

3) Open the sterile water tube (the cap should be placed face down on a clean surface) and dip the swab into the water. Recap the sterile water tube immediately since you may be using it to collect other samples and the water needs to remain sterile.

4) Collect the sample with the wetted swab by lightly brushing the swab on the surface. Molds on kitchen wallboard will almost certainly be different from molds on the basement floor joists, therefore, different swab samples should be collected on each surface type containing visible mold. At each site, swab different types of molds identified by different colors or textures.

5) Open an empty, clearly labeled (by you), sterile test tube and place the cap face down on a clean surface. Place the swab into the tube and gently snap the end of the swab stick off on the edge of the tube. Replace the cap.

6) When comparing contamination conditions, collect samples consistently at each site. In other words, at each site, for example, swab a 3″X3″ area of the material in question.

7) Store samples in a cool place (4°C) and send them to the lab as soon as possible for analysis.

B. Bulk Sampling

Label the collection vessel and record sample information on the Chain of Custody

Collect a known size/ piece of suspect material using a clean collection tool or gloved hand

Place the sample material in the sterile or clean collection vessel. Use a ziplock type bag for larger samples.

Return samples to the lab

C. Carpet Sampling

Materials needed:

Sterile bacteriological cassettes (Millipore)
Staplex EC-1 Air Sampler (vacuum pump)
Ruler or template for sampling area guide
Water insoluble labeling marker (Sharpie brand works great)

Procedure :

1) At the chosen sample site, remove the red cassette plug and attach the pump tubing (or vacuum hose) securely.

2) Remove the top of the cassette (blue plug side); a quarter can pry the top off gently (like a paint can, you’ll need to go around the top a few times loosening as you go). Place the cassette top down on a clean surface.

3) Have your template or ruler on the site to be sampled and turn the pump on to begin sampling.

4) The cassette acts like a vacuum attachment and, to get a good sample, it is best to sweep the cassette over the area at a slight angle. Be consistent with the way in which you hold the cassette.

5) The sample size depends on how soiled the rugs are. It’s a good idea to have a standard template that you can use for consistent sweeping. The template can be a piece of thin cardboard with a specific area cut out of the center (e.g. 3″X3″). If you suspect heavily soiled rugs sweep the cut-out surface once. If the rugs are expected to be clean and free of contaminants, sweep the cut-out surface two or three times. Just be consistent! If you are trying to compare rugs in different rooms, choose one sample size (one sweep, for example) and use that size for all samples.

6) Timing is also important. Even though you are sweeping the cassette over a known and consistent area, if you pass the cassette quickly over one area and more slowly over another, this will result in inconsistent sampling. Timing the sweeps can be done with a stopwatch or second hand on a wristwatch. Eventually, when you’ve done a lot of sampling, you will become familiar with how long each sweep takes and may not need the timer.

7) Once you have collected the sample, replace the top part of the cassette and press down firmly to close it securely. Remove the pump tubing from the bottom and replace the plug. Be sure the cassette is labeled clearly with a water insoluble marker and that this label corresponds with that you have described on the Chain of Custody/Sample Submission Form that you send along with the samples.

8) Store samples at room temperature and send them to the lab as soon as possible. A bubble-wrap envelope is perfect for shipping. Or, wrap the cassettes in newspaper, etc. and send in a padded envelope.<

Air Sampling

Ambient Air Sampling

Materials and Equipment:

• Staplex EC-1 Vacuum Pump
• Culture Plates
• Permanent Markers
• Calibrated Rotameter
• Single stage Bioaerosol Impactor (400 holed)
• Ultrasonic Cleaner
• Alcohol Swabs
• Stopwatch
• Tripod for sampler

Procedure :

• Design sampling scheme and completely fill out the Chain of Custody.
• Label culture plates containing growth medium with project name, date, and location. The label should be on the bottom of the culture plate (the side that contains the growth medium) and in small printing around the edge of the dish. Water-insoluble markers (i.e. Sharpie brand) work well.
• Calibrate the air pump flow rate to 28.5 liters per minute (lpm) using a calibrated flow meter. A range from 26 lpm-30lpm will effectively collect bioaerosols; please record flow rate on the Chain of Custody.
• Disassemble the single stage bioaerosol impactor into its three pieces. Both sides of the top two pieces should be wiped with an alcohol swab and allowed to thoroughly dry before each sample collection. It is good practice to sterilize the impactor in the sample collection location.
• Remove the cover of the culture plate and place it face down on a clean surface. Place the bottom of the plate containing the growth medium on the impactor base with the open side facing up. The plate must be centered on the impactor base to achieve a good seal and successful sample collection.
• Place the 400-holed middle piece of the impactor over the open culture plate being sure that nothing comes in contact with the medium. The top piece can then be fitted on top with the three holes lined up with the spring closures on the base of the impactor. Close the sampler by attaching the three springs into the holes on the top piece of the impactor.
• Samples should be collected at least two feet off the ground unless circumstances require otherwise. A consistent height should be used for sample comparison; a photographic tripod works well and many impactors have a threaded hole on the bottom for this purpose.
• Simultaneously turn on the air pump and a stopwatch or other timing device. Time the sample for three minutes. If the time varies from the three minutes, be sure it is clearly recorded on the Chain of Custody.
• Turn off the air pump, disassemble the impactor, remove the culture plate, which should have a 400 hole concentric circle pattern in the medium, and cover the culture plate immediately. Place the culture plate upside down (medium/labeled side up) and secure with a tightly fitting elastic band.
• Repeat all of the above steps for each subsequent sample.
• Place all samples in a clean, sealed ‘Ziploc’-type bag and ship to the lab via an overnight delivery service.

Tips

• Summer time/warm weather sampling requires that culture plates be stored in a cooler on ice (4°C) until samples are shipped. Watch for undesirable condensation.
• If testing is conducted on a Friday or weekend/holiday day, please place all samples in a refrigerator until they can be shipped overnight to arrive at the lab Monday through Friday.
• Proper shipping maintaining sample integrity includes culture plates securely closed with elastic bands and placed upside down in a clean, sealed bag. Sufficient packing materials (bubble-wrap, Styrofoam peanuts, etc.) should be placed in the shipping container around the sample bag.
• Wall cavity samples are set up somewhat differently than room air samples and require adapters for the impactor. If wall cavity samples are to be collected, please contact the lab for specific instructions and sampling guidelines.
• When collecting air samples, do not agitate surrounding surfaces prior to sample collection. Collect surface samples after air samples are collected.
• Windows and doors leading to the outside should be closed as much as possible at least 24 hours before testing. Outside air can significantly influence/skew indoor air sample results.
• If possible, collect a representative sample in the location where the mold is observed or suspected and collect an “indoor control” sample in a location separated from the contaminated area. Additionally, collect an outdoor sample for comparison purposes. The outdoor sample data may not be significant in some cases, but it is better to have collected the sample at the time of the initial investigation rather than going back when conditions may have changed.
• Sharpie or other water-insoluble markers are recommended for sample labels. If these are not available, a small piece of tape may be placed on the bottom of the culture plate and labeled.
• Culture plates should always be stored upside down, whether they are sterile, unused plates or sample plates. This prevents condensation from collecting on the cover and dripping onto the growth medium; this excess liquid can carry spores and bacterial cells around the plate skewing the results.
• Inspect culture plates before use and always take extra plates to the testing location. The media becomes dehydrated over time and may be too thin to use for sampling. Do not use culture plates that have microbial growth on them; contamination of unused plates is rare, but possible.
• Finally, please feel free to call the lab for sampling guidance or with questions that may arise prior to or after mold investigations.

Cavity Air Sampling

Materials and Equipment

• Staplex EC-1 Vacuum Pump
• Culture Plates
• Permanent Markers
• Calibrated Rotameter
• Single stage Bioaerosol Impactor (400 holed)
• Ultrasonic Cleaner
• Alcohol Swabs
• Stopwatch
• Tripod for sampler
• GML created adapters
• Sterilized cavity tubing

Procedure :

Follow air sampling procedures above in addition to the instruction below.
Prior to each sample collection, sterilize the drill bit, Andersen tubing adapter, and Andersen sampler

• Open and insert a new, individually wrapped, sterile piece of tubing
• Measure the flow rate of sampler using calibrated rotameter and record on P1571 Chain of Custody.

LOCATION FOR SAMPLE COLLECTION

• Collect cavity samples via holes drilled by GML in undisturbed areas or through existing holes; e.g. plumbing openings, electrical outlets, etc.

COLLECTING CAVITY SAMPLES

• Insert the open end of the tubing that is attached to the adapter on the Bioaerosol Impacter through the hole into the cavity.
• Collect cavity air onto prepared culture plates for three minutes at a rate of approximately 28.5 liters per minute; the rate is specifically determined with the rotameter.

COLLECTING CONTROL SAMPLES

• Collect control cavity air samples in areas not affected by moisture in order to provide a comparison sample, and are used as interpretive guides.

RECORDING INFORMATION

• Record location and other collection information on chain of custody. Label samples consistent with chain of custody