ERP Micro December 2019

(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.

(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.

E. Sample Enrichment (a) Prewarm mEHEC® broth to 42 C ± 1°C before use.

(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) 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. (d) carcass cloth: Use Fremonta MicroTally TM ( https://www.fremonta.com/microtally ) cloth or equivalent for sampling. Collect carcass cloth per FSIS (FSIS Directive 10,010.1 Rev. 4). Add 200

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