3.  Molecular Laboratory must have an ongoing Bio-safety SOP and also quality improvement program to monitor and evaluate objectively and systematically the quality and appropriateness of the information/service it provides

4. Why do we need safety  Safe handling of hazard microorganisms, Chemicals and radiations  Safe conditions for the work to keep the tests in right results

5. Bio-Safety for the Personnel  Hazard Microorganisms (Risk Group 1 to 4)  Chemicals  Radiations  General Bio-Safety Rules  Specific Considerations Ethidium Bromide, Acrylamide, UV radiation (Door Glasses)

6. Specimens Specimens are to be handled according to SOPs, with special attention to any safety precautions ( Laboratory General Bio-Safety)

7. Hazard Microorganisms Each detectable organisms must be classified on its Bio-Safety Level Risk of Lab. Infections must be determined for each organisms according to applied diagnostic tests Blood-borne and respiratory pathogens

8. Hazard Microorganisms Continue  If a dead air UV hood is used, turn on the UV lamp at least 15 minutes prior to working  When using a biological safety cabinet (BSC), it must be on 1/2 hour prior to specimen preparation procedures.  Working surfaces must be decontaminated prior to specimen preparation with 10% bleach solution (freshly made daily) followed by 70% ethanol rinse  Gloves must be worn at all times for your safety when handling specimens

9. Safe conditions for the work to keep the tests in right results

10. Specimen preparation area Defined Flow work and Lab. Design Awareness to the Applied Lab. Procedures (provide necessary SOP)

11.  Specimens In general, specimens should be received and accessioned in an area of the facility, that is isolated from testing areas.  separation and aliquoting of original specimens  Back up specimens should be kept frozen at -20 o C to - 70 o C. and avoided of freeze-thawing  Use of sterile container Patient Specimens and Genomic Materials should be stored separately than reagents Receiving samples:

12. Safe conditions for the work Continue  Gloves are to be changed frequently, anytime you contaminate them, whenever a specific procedure SOP dictates, and never be worn outside Post Amplification Area, especially in Pre Amplification Area  Aerosol barriers tips are used for pipetting into amplification tubes and pipetting amplification product

13. Other routes of contamination Contamination of negative specimens with control specimens False negative results due to inhibitors transferring from environment (Such as Latex gloves….) and inefficient extraction methods (Accreditation Protocols)

14. Molecular Lab.  Specific flow work and Lab design Depends on applied Technique Separation of work areas:  Pre-Amplification area,  Post-Amplification area,  Storing space  Specific instruments, materials and other condition for each area

15. How contamination can be distributed Routs of contamination depend on the technique  target amplification  signal amplification

16. Decontamination Post-amplification products from previous analysis can be distributed  Decontamination and inactivation protocols for the test and environment  Work-benches and sinks should be decontaminated each day of use  Products must be discard in a safe way

17. General Consideration in Designing Molecular Lab.  Each sample test contains 10 10 -10 12 amplicons, distributed easily by hand, hair, Lab coat, samplers,…….  Reagents, materials and instrument can be contaminated easily  Complete separation between pre & post amplification areas must be considered

18. Continue  Air flow should be from clean area to dirty area  Special program must be followed if different PCR sets are being used  Additional materials recommended to divide in small sections and stored in separate space from PCR Lab

19. General Rules  Each Working space must have positive pressure in Pre-Amplification Area  Transferring reagents, materials, and any instrument must be avoided from dirty area to clean area  Lab coat and gloves must be replaced before entering to the clean area  Necessary material must be kept at each lab  Each lab must have UV, Bleach (10%)

20. Preparation Lab  Preparation Lab is the center of the PCR Lab  Each solution must be checked carefully  Lab stocks must be divided to small amount and used one by one in the lab  Just mixture reaction will be prepared in this lab without any DNA samples  Preparation Lab must have its own specific materials, samplers, buffers, and tube samples

21. Extraction room  Any extraction must be carried out under safety cabinet  Necessary buffers, and materials must be taken out from freeze and kept at the room temperature before work  It is recommended of two gloves with working of hazardous specimens  10% bleach are recommended to used for any probable contamination of centrifuge, safety cabinet, and so on  Sampler tips must be un-reusable and plug tips recommended

22. Amplification room  Thermocycler must be kept in a small clean area  Different control negative must be considered in order to insure the quality of the test (buffers, primers, dNTP, …)

23. Detection Lab.  Test tubes can be opened after amplification just in Detection room  To prevent the distribution of amplified products 10% bleach and UV should be available  It is recommended not to keep the remaining products after any experiment  Lab coats and gloves must be removed