Compact Dry™ ETB

Cat. no. ETB100 Compact Dry™ ETB, 60x75mm Tray with 10x60mm Well 100 trays/box

INTENDED USE

Hardy Diagnostics Compact Dry™ ETB is a ready-to-use test method recommended for the isolation and enumeration of organisms belonging to the family Enterobacteriaceae in raw materials, finished products, or on environmental surfaces pertaining to food and related industries.

This product is not intended to be used for the diagnosis of human disease.

SUMMARY

The family Enterobacteriaceae is a large, diverse family of gram-negative bacteria that includes a wide variety of harmless symbionts and recognized pathogens. Of these, Salmonella, Escherichia coli, and Shigella are well-documented food-borne pathogens, and Salmonella alone was responsible for 35% of all food-borne illness-related hospitalizations and 28% of food-borne illness-related deaths in the United States in 2011.(1, 4) Thus, the detection of these potentially pathogenic species is essential in ensuring food safety for consumers.

Compact Dry™ ETB is a ready-to-use chromogenic medium for detecting and enumerating Enterobacteriaceae that contains dehydrated culture media and a cold water-soluble gelling agent in a non-woven cloth matrix. The medium is instantly hydrated when inoculated with a sample, and capillary action diffuses the sample evenly over the matrix to form a gel within seconds. Enterobacteriaceae colonies grown on Compact Dry™ ETB turn red or purple due to a chromogenic indicator included in the media.

Compact Dry™ ETB is comparable to other dry film test methods for the detection and enumeration of Enterobacteriaceae.(6) Compact Dry™ ETB is NordVal validated (NordVal no. 034) and MicroVal validated (no. MV0806-002LR). The ready-to-use trays save space and greatly reduce the time needed to perform microbiological testing. Compared to other commonly used culture systems, Compact Dry™ has a longer shelf life, can be stored at room temperature, does not require manual sample spreading, is rigid, stackable and easy to label, and allows for direct colony picking for further subculture.

FORMULA

Compact Dry™ ETB contains dehydrated culture media with glucose, a gelling agent, and a chromogenic indicator to facilitate the identification and differentiation of Enterobacteriaceae from other types of organisms.

Final pH 7.0 +/- 0.2 at 25°C.

STORAGE AND SHELF LIFE


PRECAUTIONS


PROCEDURE


Method of Use:

1. Remove the set of four trays from the foil pouch and separate each individual tray by gently bending along the connecting edge until each tray snaps free. Alternatively, if setting up a dilution series of the same sample, trays can be left connected to facilitate reading similar samples. Trays that are not used immediately should be resealed in the foil pouch. Refer to the "Storage and Shelf Life" section for proper storage of unused trays.

2. Remove the lid of the tray using two fingers to hold down one end of the lid and the thumb to lift the opposite end. Lids are easier to remove using a “peel back” method as opposed to a “pull off” method.

3. Inoculate by pipetting 1ml of sample directly to the center of a dry tray well, being careful not to touch the surface of the matrix with the pipet tip. Once dispensed, the sample will automatically diffuse across the surface by capillary action to form a gel; manual spreading of the inoculum is discouraged. Remember to account for the sample inoculum when calculating the dilution series.

4. Replace the lid and label the tray with appropriate information, including the sample dilution factor.

5. Invert the tray and incubate, upside down with the medium on top, at 35-37°C for 24 ± 2 hours. NOTE: Use the appropriate temperature/time designation according to the legal specification of the prescribed food analysis regulation.

6. Count colonies from the backside of the tray to calculate CFU/ml using the Mini Light Box II (Cat. no. 378642000) or comparable back lighting. If the colony count is high, use the 1cm x 1cm molded grid on the back of the tray to assist in colony counting. Use a sheet of white paper with gridded lines to diffuse the light if the molded grids in the tray are difficult to visualize with a light box.

Alternate Membrane Filter Method:

1. Remove the plate cap and pipet 1.0ml of sterile water into the middle of a dry Compact Dry™ ETB sheet. Set aside.

2. Using sterile forceps, pick up a sterile membrane filter.

3. Remove the funnel from a sterilized filtering device and set the sterilized membrane filter in place.

4. Set the funnel and pour the sample into the funnel. Filter the sample under reduced pressure.

5. After filtering, wash the inner surface of the funnel with 20-30ml of sterile water and filter. Repeat this step two or three times.

6. Detach the funnel and remove the membrane filter with sterile forceps. Place the filter--trap side up--on the pre-moistened Compact Dry ETB sheet, making sure to avoid trapping air bubbles between the filter and the plate.

7. Recap and invert the plate and incubate under prescribed conditions.

INTERPRETATION OF RESULTS

After incubation, read trays against a white background such as the Mini Light Box II (Cat. no. 378642000) or comparable back lighting.

Enterobacteriaceae colonies will be red or purple due to utilization of the chromogenic indicator in the medium. Most colonies grown will be red or purple. Un-pigmented or otherwise colored colonies are indicative of other gram-negative bacteria. Count all red or purple colonies to obtain the total Enterobacteriaceae count. Count all colonies, regardless of pigmentation, to obtain the total gram-negative count. The growth area is 20cm2 . If the colony count is high, the total count can be obtained by multiplying the average number of colonies observed in one 1cm x 1cm square grid by 20.

LIMITATIONS

During inoculation, do not touch the surface of medium and be careful to avoid any contamination by airborne microorganisms.

During incubation, keep cap tight on plates to avoid any possible dehydration.

A dilution may be needed when the sample has a dark color.

When the sample is viscous (thick), pipetting the sample on several points on a plate or an additional dilution may be needed for an even suspension.

When the sample contains an enzyme, it may react with the enzyme substrate in the dry sheet and affect the color.

If the nature of sample does affect the reaction of the medium, inoculate only after the factor is eliminated by means of dilution and other techniques (e.g. samples with high viscosity, colored, reactive with chromogenic substrate, and with a high or low pH).

It is recommended to use a stomacher and filter homogenized sample afterwards to eliminate carry over of tiny particles of foodstuff onto the surface of the medium.

Some colonies may not be clearly red or purple colored.

Counting colonies may be difficult against a dark background. For best results, count colonies with the tray held against a white background such as with the Mini Light Box II (Cat. no. 378642000) .

If using a light box, molded grid lines in the tray, or colonies, may be difficult to view due to excessive brightness. Diffuse the light using a sheet of white, gridded (1cm x 1cm) paper underneath the tray to facilitate colony counting.

Colonies are not distinguishable on trays if concentrations are above 100 CFU/ml, as high colony counts will result in the whole surface becoming colored. The sample should be diluted to a concentration of less than 100 CFU/ml for best use.


MATERIALS REQUIRED BUT NOT PROVIDED

Standard microbiological supplies and equipment such as loops, swabs such as EnviroTrans™ , applicator sticks, scoops, dilution buffers such as Dilu-Lok™ II , other culture media, a light box (Cat. no. 378642000), incinerators, and incubators, etc., as well as serological and biochemical reagents, are not provided.

QUALITY CONTROL

Test Organisms Inoculation Method* Incubation Results
Time Temperature Atmosphere
Escherichia coli
ATCC ® 25922
J 24±2hr 35-37°C Aerobic Growth; red colonies
Salmonella enterica var. Typhimurium
ATCC ® 14028
J 24±2hr 35-37°C Aerobic Growth; red colonies
Klebsiella pneumoniae
ATCC ® 13883
J 24±2hr 35-37°C Aerobic Growth; red colonies
Enterococcus faecalis
ATCC ® 29212
J 24±2hr 35-37°C Aerobic No growth

USER QUALITY CONTROL

PHYSICAL APPEARANCE

Compact Dry™ ETB should appear dry, free of particles, and light yellow to light pink in color.

E. coli growing on Blood Agar

Escherichia coli (ATCC ® 25922) colonies growing on Compact Dry ETB (Cat. no. ETB100). Incubated aerobically for 24 hours at 35°C.

REFERENCES

1. "CDC 2011 Estimates: Findings." CDC Estimates of Foodborne Illness in the United States. Centers for Disease Control and Prevention, 8 Jan. 2014. Web.

2. American Public Health Association. Standard Methods for the Examination of Dairy Products. APHA, Washington, D.C.

3. APHA Technical Committee on Microbiological Methods for Foods. Compendium of Methods for the Microbiological Examination of Foods. APHA, Washington, D.C.

4. U.S. Food and Drug Administration. Bacteriological Analytical Manual. Arlington, VA
http://www.fda.gov/Food/FoodScienceResearch/LaboratoryMethods/ucm2006949.htm

5. American Public Health Association. Standard Methods for the Examination of Water and Wastewater. APHA, Washington, D.C.

6. Teramura, Hajime, Shingo Mizuochi, and Hidemasa Kodaka. "Evaluation of the dry sheet medium (Compact Dry ETB (R)) for enumerating Enterobacteriaceae in meat samples." Biocontrol science 17.3 (2011): 149-154.


ATCC is a registered trademark of the American Type Culture Collection.
Compact Dry™ is a trademark of Nissui Pharmaceutical Co., Ltd.

MicroVal approval no. MV0806-002LR per ISO 16140:2003
NordVal approval no. 034

020416hh

Manufactured for and distributed by: