1 HIV Drug Resistance Training Module 13: Equipment and Supplies.

1

HIV Drug Resistance Training

Module 13:

Equipment and Supplies

2

Topics

Overview of Responsibilities Equipment and Supplies Needed for

Genotyping Equipment Use and Care

3

Objectives

Identify specialized equipment and supply needs for a genotyping lab.

Describe how to ensure quality through appropriate care and use of equipment.

4

A Systems Approach to Laboratory Quality

Organization

Stock Management

SOP, Documents & Records

Process Improvemen

t

Personnel

Quality Control

Occurrence Management

Specimen Management

Equipment

Data Management

Assessment

Safety & Waste

Management

5

Functioning Equipment is Vital to Quality Service

Produces reliable

test results

Lowers repair costs

Prevents delays in

testing

Maintains productivity

QualityResults

FunctioningEquipment

6

overview of responsibilities

What are the responsibilities of lab management?What are the responsibilities of lab staff/technicians?

7

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

8

Staff/Technician Responsibilities

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

9

Discussion

How will you assign responsibilities for equipment among different members of management and staff?

10

equipment and supplies needed for genotypingWhat equipment and supplies do all labs need for HIVDR testing?What equipment and supply decisions depend on the testing method?

11

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)

12

Core Laboratory Supplies

Micropipettes Agarose gel electrophoresis, power supply Refrigerators and freezers Distilled, deionized water supply Sink Ice machine Timers

13

Equipment for Genotyping

DNA sequencer PCR machine Dead-air cabinet (PCR workstation) Computer attached to sequencer Specialized software dependent on Platform

used Sequencing kit

May be influenced by platform (ViroSeq,

TruGene, or In-house)

14

Laboratory Supplies

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)

15

Types of Pipettes

Single, multichannel, automated, repeater Use disposable, single-use, pipette tips Positive displacement and aerosol barrier

tips recommended for pre-PCR work

16

Equipment: Pre-Amplification Area, In-house Assay

Essential:– 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

17

PCR Workstations (Dead-air Cabinets)

18

Equipment: Post-Amplification Area,In-house Assay

Essential:– 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

19

ABI Sequencing Instruments

20

Alternative Sequencing Instruments

Beckman CEQ8000/8800

TruGene

21

Remember Workflow!

Consider lab layout (module 4) when planning equipment needs and placement

LAB 1

"Specimen Processing" Lab

LAB 2

"Post-amplification" Lab

-80°C

freezer

BSC: bio-safety cabinet

Ice machine

Ice machine

-80°C

freezer

-20°C

freezer

wor

kbe

nch

Sequencer Gel electrophoresis areaPCR

machines

Lab coat rack

Lab coat rack

NA extraction equipment

PCR machines

Dead air cabinet

De

ad a

ir

cab

ine

tC

en

trif

ug

e

sink Eye wash

-20°C freezer

BSC BSC

Eye wash

Ce

ntr

ifu

ge

LAB 1


LAB 2


-80°C

freezer-80°C

freezer

BSC: bio-safety cabinet

Ice machine

Ice machine

-80°C

freezer-80

°C

freezer-20°C

freezer-20°C

freezer

wor

kbe

nch


machines

Lab coat rack

Lab coat rack


PCR machines

Dead air cabinet

De

ad a

ir

cab

ine

tC

en

trif

ug

e

sink Eye wash

-20°C freezer-20°C freezer

BSCBSC BSCBSC

Eye wash

Ce

ntr

ifu

ge-20°C

freezer

wor

kben

ch

Ice machine

LAB 1

"No Template" Lab

LAB 2


LAB 4


LAB 3

"Nested PCR" Lab

-80°C

freezer

-20°C freezer

BSC

wor

kben

ch

Ice machine

Ice machine

-80°C

freezer-20°C

freezer

wor

kben

ch


machinesworkbench PCR

machines

-20°C freezer

Cen

trif

ug

e

Pass-through window

Lab coat rack

Lab coat rack

Lab coat rack

Lab coat rack


workbench

sink

sink

sink

sinkEye

wash

Eye wash

BSC

Cen

trif

ug

e


wor

kben

ch

Ice machine

LAB 1

"No Template" Lab

LAB 2


LAB 4


LAB 3

"Nested PCR" Lab

-80°C

freezer-80°C

freezer


BSCBSC

wor

kben

ch

Ice machine

Ice machine

-80°C

freezer-80°C

freezer

-20°C

freezer-20°C

freezer

wor

kben

ch


machinesworkbench PCR

machines


Cen

trif

ug

e

Pass-through window

Lab coat rack

Lab coat rack

Lab coat rack

Lab coat rack


workbench

sink

sink

sink

sinkEye

wash

Eye wash

BSCBSC

Cen

trif

ug

e

22

Reflection

What is our “shopping list?”

23

equipment use and care

What can we do to make sure our genotyping equipment is properly cared for?

24

Maintenance

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"

25

Introducing New Equipment

Calibrate or verify specified performance upon installation– 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

26

Maintenance: Function Checks

Verify that equipment is working properly Performed routinely

– Daily, weekly, monthly– After adjustment or repair

Example:– Monitoring refrigerator temperatures– Verifying pipette accuracy– Checking centrifuge speed

27

Refrigerator and Freezer: Use and Care

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

28

Refrigerator & Freezer:Temperature Checks

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.)

29

Refrigerator & Freezer:Temperature Log

30

Keep a Log for All Maintenance Activities

31

Exercise: Create a Maintenance Activity List

Purpose:– Use what you have learned in this module and

create a maintenance checklist specific to your test site

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

32

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?

33

Summary

Overview of Responsibilities Equipment and Supplies Needed for

Genotyping Equipment Use and Care

1 HIV Drug Resistance Training Module 13: Equipment and Supplies.

Documents

Transcript of 1 HIV Drug Resistance Training Module 13: Equipment and Supplies.