M1: INTRODUCTION TO MOLECULAR BIOLOGY LABORATORY Flashcards
An ideal laboratory consists of three completely independent rooms or areas
reagent preparation room
sample preparation room
PCR room
This part of the laboratory is identified as clean rooms
Pre-amplification rooms
This part of the laboratory is identified as dirty rooms
Post-amplification rooms
This room should be under positive air pressure
Pre amplification room
What does positive air pressure do ?
impedes the entrance of any airborne contaminant into the pre-amplification room when the door is opened
What does negative air pressure do ?
impedes anything from coming out of this laboratory and contaminating the surrounding environment
This is referred to as the PCR room
Post amplification room
Pre-amplification rooms include
reagent preparation room
sample preparation room
Describe the unidirectional workflow of a molecular biology laboratory
reagent prep -> sample prep -> amplification -> post amplification
The construction of this into each laboratory is an inexpensive way of creating differential pressure for the laboratory
anteroom
Safety precautions for anterooms
door leading into the anteroom from the outside and the door to the inside should never be opened at the same time
adding sticky mats at the entrance of each anteroom laboratory
This pertains to containment principles, technologies and practices that are implemented to prevent the unintentional exposure to biological material or their accidental release
biosafety
Pertains to the protection, control and accountability for biological agents and toxins within facilities in order to prevent their loss, theft, misuse, diversion, unauthorized access, or intentional unauthorized release
biosecurity
Safety concerns on inactivation
virulence and amount of pathogens sense and mobility limitations of wearing full PPE user hygiene and health status spills and aerosol-generating procedures harsh chemicals exposure to extreme heat and cold glassware
Safety concerns on Nucleic acid extraction
check if procedures have prior inactivation irritating chemicals spills and aerosol generating procedures harsh chemicals exposure to extreme heat and cold glassware
Safety concerns on PCR and other nucleic acid tests (Pathogen is inactivated)
Disposal is still in biohazard bags for environmental concerns
Cleanliness maintained for proper detection
Hazards from equipment (heat, entrapment, moving parts)
Safety concerns on Post-PCR (Pathogen is inactivated)
contamination from liquids with amplified nucleic acid
hazards from electricity, UV, chemicals, heat, sharps
This type of waste includes all utensils, containers, tissue paper, etc that have been in contact with biohazardous material
biohazard waste
This type of waste is usually segregated in strong yellow coloured bags
biohazard waste
This type of waste is usually collected in yellow bench-top bins
sharps
document that gives detailed information about a material and about any hazards associated with the material
Material Safety Data Sheet (MSDS)
written guideline stating the policies, procedures, and responsibilities that serve to protect laboratory personnel from the health hazards associated with the hazardous chemicals
Chemical Hygiene Plan (CHP)
Why should you never use a metal spatula when working with peroxides ?
because metals will decompose explosively with peroxides
used to track chemicals from their time of purchase to when they are used and discarded
Chemical tracking system
Patient specimens from suspected or confirmed cases should be transported as
UN3373, “Biological. Substance, Category B”
Viral cultures or isolates should be transported as
Category A, UN2814, “infectious substance, affecting humans”
What is a HEPA filter
High Efficiency Particulate Air Filter
This biosafety cabinet is also known as open front BSC
BSC 1
This BSC does not protect the sample/ product from possible contamination
BSC 1
This BSC protects the worker from potentially infectious aerosols; Also capable of protecting the sample from possible contamination
BSC 2
What is the principle of BSC 2 ?
Air passes through HEPA filter BEFORE flowing over the materials within [Protects the specimen from contamination]
Air to be exhausted first passes through HEPA filter [Protects the worker and the environment from potentially infectious aerosols]
What type of BSC 2 has 70% of air is recirculated to the working area; 30% of air is exhausted ?
BSC Class IIA
What type of BSC 2 has 30% of air is recirculated to the working area; 70% of air is exhausted ?
BSC Class IIB1
What type of BSC 2 has no recirculation of air; Total exhaust of air through an exhaust HEPA filter ?
BSC Cass IIB2
What type of BSC is completely enclosed and are equipped with glove ports; Infectious material within is handled with rubber gloves that are attached and sealed
BSC 3
Trained laboratory staff or certified contractors should perform tests or check on Adjustable and fixed-volume automatic pipettors
Check for volumetric accuracy and reproducibility, and recalibrate as necessary before placing in service initially and at specific defined intervals, at least once every 6 months.
Pipettor inaccuracy is one of the most common sources of laboratory error.
Trained laboratory staff or certified contractors should perform tests or check on Thermometers
Test against a standardized thermometric device annually
Trained laboratory staff or certified contractors should perform tests or check on Refrigerators and freezers
Establish tolerance limits for temperatures. The refrigerator’s temperature tolerance limits
must reflect the most stringent needs of all reagents, supplies, and specimens stored within it.
Maintain daily records of temperatures and appropriate documentation of corrective action for out-of-range temperatures.
Trained laboratory staff or certified contractors should perform tests or check on Incubators and water bath
Establish tolerance limits for temperatures, carbon dioxide level, and humidity as applicable.
Maintain daily record of temperatures and appropriate documentation of corrective action for out-of-range
temperatures.
Trained laboratory staff or certified contractors should perform tests or check on Centrifuges
Measure operating speeds periodically with a tachometer. Document the readings.
Maintain daily record of temperatures for refrigerated centrifuges. Verify performance of centrifuge timers by comparing to a known standard every 6 months.
Trained laboratory staff or certified contractors should perform tests or check on Autoclaves
Verify content processing using heat sensitive tape with each autoclave batch.
Verify effective sterilization with an appropriate biological indicator weekly.
Perform autoclave maintenance annually or as per manufacturer, including a pressure check and calibration of temperature device.
Check autoclave mechanical timing device periodically.
Maintain records of autoclave operation and maintenance in the equipment log.
Trained laboratory staff or certified contractors should perform tests or check on Analytical balances
Check accuracy with standard weights and class at a predetermined frequency. Document the results with an evaluation of their capacity.
Service and calibrate periodically using qualified personnel.
Maintain records of service and calibrations.
Place the analytical balance in a stable location so that vibration does not affect the readings.
Trained laboratory staff or certified contractors should perform tests or check on Biosafety cabinets/laminar air flow hoods
Verify air intake grills are not obstructed.
Certify cabinets/hoods annually by a trained service technician, certified maintenance department or company.
Check daily for air flow as instructed by manufacturer and document the results to verify the effectiveness of the hood’s personnel and environmental protective
functions.
Clean the ultraviolet lamp, if used, weekly with 70% alcohol.
Document daily and weekly cleaning.
Trained laboratory staff or certified contractors should perform tests or check on Generators
Maintain and keep readily accessible records to verify that the backup generator system is in place and
operational.
Follow SOPs that detail generator maintenance. Procedure should list the frequency and procedural steps of maintenance and testing.
Supporting logs should document monthly checks on critical generator components, including fluid levels (oil, coolant, and fuel), belts, battery, testing startup, and operation.
Trained laboratory staff or certified contractors should perform tests or check on Out of service equipment
Any equipment that is out of service for any reason should be clearly identified and removed from the laboratory
