HIV Drug Resistance Training
Module 13:
Equipment and Supplies
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Topics
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Overview of Responsibilities
Equipment and Supplies Needed for Genotyping
Equipment Use and Care
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Objectives
Identify specialized equipment and supply needs
for a genotyping lab.
 Describe how to ensure quality through
appropriate care and use of equipment.
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3
A Systems Approach to Laboratory Quality
Organization
Personnel
Equipment
Stock
Management
Quality Control
Data
Management
SOP,
Documents &
Records
Occurrence
Management
Assessment
Process
Improvement
Specimen
Management
Safety &
Waste
Management
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Functioning Equipment is Vital to
Quality Service
Produces
reliable
test results
Functioning
Equipment
Lowers
repair costs
Prevents
delays in
testing
Quality
Results
Maintains
productivity
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overview of responsibilities
What are the responsibilities of lab management?
What are the responsibilities of lab staff/technicians?
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Management Responsibilities
In the area of…
Lab mgt. must ensure…
Ordering pieces of equipment
Each piece meets needs and
specifications of lab
Receiving and installing
equipment
The lab is ready for installation
Accountability and supervision
Assigning oversight responsibility for
equipment
Standard operating procedures
Develop/delegate and approve
equipment SOPs
Operating equipment
Train staff
Troubleshooting
Train staff
Inventory control
Establish inventory record
Maintenance
Develop maintenance program
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Staff/Technician Responsibilities
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Attend training and learn all procedures
Properly use the instrument by following SOPs
Perform routine maintenance
– Routine function check
– Troubleshooting
– Maintenance log
Take corrective actions: notifying lab manager
when problems arise
 Keep records on maintenance and problems
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Discussion

How will you assign responsibilities for equipment
among different members of management and
staff?
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equipment and supplies needed
for genotyping
What equipment and supplies do all labs need for HIVDR testing?
What equipment and supply decisions depend on the testing
method?
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Core Laboratory Equipment
Microcentrifuge
Vortex
Biological safety cabinet
Agarose gel electrophoresis
Computer, printer, Internet
connection
 UV Imaging system
 Reliable power supply
(Uninterruptible power
supply, UPS, for short outages)
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Core Laboratory Supplies
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Micropipettes
Agarose gel electrophoresis, power supply
Refrigerators and freezers
Distilled, deionized water supply
Sink
Ice machine
Timers
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Equipment for Genotyping
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May be influenced by platform
(ViroSeq, TruGene, or Inhouse)
DNA sequencer
PCR machine
Dead-air cabinet (PCR workstation)
Computer attached to sequencer
Specialized software dependent on Platform used
Sequencing kit
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Laboratory Supplies
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Sequencer supplies: capillary array, polymer,
buffers
PCR supplies: tubes, plates, trays
RNA/DNA purification: columns, buffers
Electrophoresis: Agarose, ethidium bromide or
other stains, buffers, MW markers/standards
Sterile, RNAse-free water
Pipette tips (aerosol-resistant for pre-PCR)
Polaroid film or special printing paper
Disposable gloves
Lab coats (disposable or not)
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Types of Pipettes
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Single, multichannel, automated, repeater
Use disposable, single-use, pipette tips
Positive displacement and aerosol barrier tips
recommended for pre-PCR work
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Equipment: Pre-Amplification Area,
In-house Assay
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Essential:
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Bench with sink / tap water
Biohazard flow, class IIb
Dead air cabinet (master mix preparation)
-20°C freezer and 4°C refrigerator
Microcentrifuge 12,500-15,000 g
Vortex
Timers
Dedicated set of micropipettes for master mix and RNA
extraction
Optional:
– Ultracentrifuge 21,000-25,000 g
– Dead air cabinet (RNA extraction)
– -80°C freezer
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PCR Workstations (Dead-air Cabinets)
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Equipment: Post-Amplification Area,
In-house Assay
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Essential:
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Bench with sink / tap water
Dead air cabinet (nested reaction)
Thermal cyclers (e.g. ABI, Bio-Rad, Hybaid, etc)
Agarose gel apparatus
Photo documentation of agarose gel
DNA sequencer (e.g. ABI, Beckman)
Computer + programs (editing)
Microcentrifuge 450-550 g
Vortex
Dedicated set of micropipettes
Timers
-20°C freezer
4°C refrigerator
Optional:
– -80°C freezer
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ABI Sequencing Instruments
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Alternative Sequencing Instruments
Beckman CEQ8000/8800
TruGene
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Remember Workflow!
Ice
machine
LAB 4
"Postamplification" Lab
Sequencer
NA extraction
equipment
Gel electrophoresis area
PCR
machines
Eye
wash
workbench
workbench
workbench
Centrifuge
workbench
"Specimen
Processing" Lab
sink
Lab coat
rack
Lab coat
rack
PCR
machines
Eye
wash
Dead air
cabinet
Ice
machine
LAB 2
-20°C
freezer
PCR
machines
sink
-20°C
freezer
LAB 3
Eye
wash
BSC
LAB 1
Eye
wash
Lab coat
rack
-20°C
freezer
BSC
-80°C
freezer
Lab coat
rack
sink
Ice
machine
-80°C
freezer
NA extraction
equipment
-80°C
freezer
Lab coat
rack
workbench
-80°C
freezer
Ice
machine
"Specimen
Processing" Lab
Centrifuge
BSC
Dead air
cabinet
-20°C
freezer
BSC
LAB 2
Passthrough
window
"Nested PCR"
Lab
sink
Ice
machine
LAB 1
"No Template"
Lab
Lab coat
rack
-20°C
freezer
-20°C
freezer
Centrifuge
sink
Centrifuge
Consider lab layout (module 4) when planning
equipment needs and placement
workbench
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"Postamplification" Lab
Sequencer
Gel electrophoresis area
PCR
machines
BSC: bio-safety cabinet
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Reflection
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What is our “shopping list?”
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equipment use and care
What can we do to make sure our genotyping equipment is
properly cared for?
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Maintenance
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Properly maintained equipment is needed
for quality results
Maintenance: Routine and biannual or
yearly scheduled maintenance
– Reduces breakdowns
– Reduces costs in the long run
– Minimizes "down time"
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Introducing New Equipment
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Calibrate or verify specified performance upon
installation
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Pipettes
Centrifuge
Vortex
Etc.
Perform assay equivalence testing
– Key components of the genotyping process
(sequencer, PCR machine)
– 10-20 samples previously reported and verify results
within expected range of variability
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Maintenance: Function Checks
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Verify that equipment is working properly
Performed routinely
– Daily, weekly, monthly
– After adjustment or repair
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Example:
– Monitoring refrigerator temperatures
– Verifying pipette accuracy
– Checking centrifuge speed
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Refrigerator and Freezer:
Use and Care
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Keep organized
Periodically clean inside and outside
Ensure door is completed sealed when closing
CAUTION! – DO NOT store food items or
beverages in laboratory refrigerator or freezer
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Refrigerator & Freezer:
Temperature Checks
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Monitor daily
– Refrigerator: 4°C (2°C to
8°C)
– Freezer: -20°C (+/- 5°C)
or -80°C (+/- 10°C)
If possible install
electronic monitoring and
alarm system
 Avoid "frost-free" freezers
for storage of critical
reagents (enzymes,
specimens, etc.)
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Refrigerator & Freezer:
Temperature Log
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Keep a Log for All Maintenance Activities
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Exercise: Create a Maintenance Activity List
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Purpose:
– Use what you have learned in this module and create a
maintenance checklist specific to your test site
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Process:
– Work in groups of 3-4 (or by test site)
– Create a list of maintenance activities on a daily,
weekly, monthly, and yearly basis
– You can use the handout as a starting point.
– Activity time = 10 minutes
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Discussion
What equipment and supplies do all labs need for
HIVDR testing?
 What equipment and supply decisions depend on
the testing method?
 What challenges do you face in your setting
regarding equipment and inventory supply?
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32
Summary



Overview of Responsibilities
Equipment and Supplies Needed for Genotyping
Equipment Use and Care
33