ERP Micro December 2019

so may lead to inaccurate results. (c) When incubation indicates 42˚C, the allowed incubation range is 42 ± 1˚C.

potentially infectious materials. Decontaminate and dispose of materials in accordance with good laboratory practices and in accordance with local, state, and federal regulations. All enrichment broths should be sterilized following any culture- based confirmatory steps.

Safety Precautions

General Preparation

Assurance® GDS for E . coli O157:H7 Tq kit

(a) Use aseptic techniques. (b) Use filter laboratory bags during enrichment to minimize particulates. (c) Separate work areas for the following: media preparation, sample preparation, and pathogen detection. (d) Clean the work stations and lab equipment with a disinfectant of choice before and after use. (Sodium hypochlorite solution, phenol solution, Quaternary ammonium solution, etc.) (e) Do not reuse kit disposables.

This product is not intended for human or veterinary use. Assurance® GDS for E . coli O157:H7 Tq must be used as described in the package insert. Contents of the test may be harmful if swallowed or taken internally. If possible, maintain separate work zones and dedicated equipment and supplies for sample preparation and amplification and detection. It is recommended to utilize both positive and negative control samples. Do not use the test kit beyond expiration date on the product box label. Decontaminate and dispose of materials in accordance with good laboratory practices and in accordance with local, state, and federal regulations. Do not open or autoclave used Amplification tubes. After run is complete, place used Amplification Tubes into a sealed container with 10% bleach solution for a minimum of 15 min or else place the Amplification tubes into a double bag and dispose outside of the lab. Assurance® GDS Rotor-Gene® Improper use of the Assurance® GDS Rotor-Gene® thermocycler may cause personal injuries or damage to the instrument. Some components may pose a risk of personal injury due to excessive heat if improperly handled. For safe use, the instrument must only be operated by qualified laboratory personnel who have been appropriately trained. Servicing of instrument must only be performed by MilliporeSigma Service Engineers. Sample Enrichment The user should read, understand and follow all safety information in the instructions for the Assurance® GDS E . coli O157:H7 Tq Kit. Retain the safety instructions for future reference. To reduce the risks associated with exposure to chemicals and biohazards, perform pathogen testing in a properly equipped laboratory under the control of trained personnel. Always follow standard laboratory safety practices, including wearing appropriate protective apparel and eye protection while handling reagents and contaminated samples. Avoid contact with the contents of the enrichment media and reagent tubes after amplification. Dispose of enriched samples according to current industry standards. E . coli O157:H7 Precautions E . coli O157:H7 is a biosafety level 2 organism. Biological samples, such as enrichments, have the potential to transmit infectious diseases. Follow all applicable local, state/provincial, and/or national regulations on disposal of biological wastes. Wear appropriate protective equipment which includes but is not limited to: protective eyewear, face shield, clothing/lab coat, and gloves. All work should be conducted in properly equipped facilities utilizing the appropriate safety equipment (for example, physical containment devices). Individuals should be trained in accordance with applicable regulatory and company/institution requirements before working with

(f) Change pipette tips in between samples. (g) Wear personal protective equipment (PPE).

DNA Extraction and Purification

(a) Change pipette tips in between samples.

DNA Amplification

(a) Use aseptic technique. (b) Change pipette tips between samples. (c) Use gloves and protective laboratory wear. (d) Do not touch any PCR equipment and supplies without wearing gloves. (e) Avoid bubble formation. General Preparation (a) Ensure the gel cooling block has changed from pink (warm) to purple (frozen) in color and has been stored in the freezer (-20 ± 5 °C) for a minimum of 6 h. (b) Verify that the Assurance® GDS Rotor-Gene® program set up and data entry has been completed prior to transferring the samples. D. Sample Enrichment (a) (a) Prewarm mEHEC® broth to 42 C ± 1°C before use. (b) (b) 25 g sample: Aseptically weigh 25 g sample into 225 mL prewarmed mEHEC® broth. Stomach or homogenize for 2 min or mix well by hand. Incubate samples for 6.5–18 h at 42 ±1°C. For green onions or sprout irrigation water, incubate for 8 – 16 h. (c) (c) 375 g sample (raw beef, finely textured beef and leafy vegetables): Aseptically weigh 375 g sample into 1500 mL prewarmed mEHEC® broth. Stomach or homogenize for 2 min or mix well by hand. Incubate samples for 8–18 h at 42 ±1°C. For frozen finely textured beef, incubate for 10 – 18 h. (a) (d) carcass cloth: Use Fremonta MicroTallyTM (https://www.fremonta.com/microtally) cloth or equivalent for sampling. Collect carcass cloth per FSIS (FSIS Directive 10,010.1 Rev. 4). Add 200 mL pre-warmed mEHEC® broth to cloth contained in sample bag, assuring that cloth is completely

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