Molecular QC and Your Lab

Areas of Discussion• Regulations and QC

• QC in the Molecular Testing Process

• Types of Controls

• Streamlining Efficiencies in Your Lab with Ready-to-Use Controls– Microbiologics Product Offering– Microbiologics Solutions for Your

Molecular Testing Needs

Regulations and QC

Why QC?Protects patients

Protects reputation

Ensures your testing procedures, equipment and materials are working properly

Ensures personnel competence

Improves daily workflow and produces consistent and reliable results

Reduces risk of misdiagnosis

A Good QC Program Test accuracy and reproducibility depends on:– Reagents– Media– Test method– Instruments– Personnel

A good QC program follows ten practices to ensure reliable test results

Ten QC Best Practices1. Use quality control materials with known values 2. Include QC in the procedure manual3. Monitor media, reagents, stains, antigens4. Monitor equipment5. Train and monitor personnel 6. Participate in inter-laboratory comparison

program7. Perform verification as needed8. Perform validation as needed9. Keep detailed records10.Evaluate results

Molecular QC• Many regulatory bodies have not specifically addressed

molecular testing

• Current source: Clinical & Laboratory Standards Institute (CLSI)– MM03 Molecular Diagnostic Methods for Infectious Diseases– MM17-A Verification and Validation of Multiplex Nucleic Acid

Assays – C24 Statistical Quality Control for Quantitative Measurement

Procedures - Principles and Definitions

MM03 Molecular Diagnostic Methods for Infectious Diseases• Addresses:

– Amplified and nonamplified nucleic acid methods

– Selection and qualification of nucleic acid sequences

– Establishment and evaluation of test performance characteristics, inhibitors and interfering substances

– Controlling false-positive reactions– Reporting and interpretation of results, QA,

regulatory issues and recommendations for manufacturers and clinical labs

MM17-A Verification and Validation of Multiplex Nucleic Acid Assays • Covers analytical verification and validation of qualitative and

semiquantitative multiplex assays– Not verification of quantitative assays or expression assays

• Addresses:– Sample preparation– Multiplex methods and technology– Reference and quality control materials– Analytic verification and validation– Data analysis– Reporting results

C24 Statistical Quality Control for Quantitative Measurement Procedures

• Explains the purpose of statistical QC for quantitative measurement procedures

• Outlines an approach for planning a QC strategy for a particular measurement procedure

• Describes the use of QC material and QC data

• Provides examples that demonstrate a practical QC planning process for medical labs

QC in the Molecular Testing Process

What’s the Difference?

Traditional Microbial Identification System

Phenotypic Technologies (identifies an organism based on how the organism reacts during a certain chemical reaction)

Molecular SystemsGenotypic Technologies (identifies an organism based on the organism’s genetic makeup)

Meeting Molecular Control Requirements

Extraction DetectionAmplification

• There are multiple stages involved with any diagnostic test• Each stage must be controlled; however, different formats may be required at each individual stage

Step 1: Extraction

• Nucleic acids (DNA/RNA) are extracted from specimens either manually or using an automated method

• Control Format Required: Whole bacteria or virus are best because they mimic the patient sample

Step 2: Amplification

• Production of multiple copies of a sequence of DNA for quantification or strain typing in a polymerase chain reaction (PCR)

• Control Format Required: Positive controls could be purified nucleic acid, such as a synthetic standard, or a whole genomic extract

Step 3: Detection

• Detecting the intended target• Control Format Required:

Positive controls could be purified nucleic acid, such as a synthetic standard, or a whole genomic extract

Types of Testing: Verification• Generally conducted upon

installation of a system or new use of a method requiring that testing be done a certain number of times to confirm the system or method is working properly– Labs often conduct the test 20 times

successivelyo However, the number of tests conducted

depends on the lab’s processes and the regulations it follows

Types of Testing: Validation• Performed if an instrument is

moved• Also performed on a semi-

routine basis to ensure an instrument or process is working properly (dependent on the lab and the regulations it follows)

Types of Testing: Ongoing QCFrequency of use is dependent on the specific regulations a lab must adhere to; however, ongoing QC products are generally used on a monthly basis or when new test supplies are received

Types of Controls

Internal Process Controls• Internal process controls are built into instruments to ensure

the instruments are running properly – They are simultaneously extracted and amplified (or only

amplified) in the same tube with the pathogen targetoNote: Internal process controls should always be combined with an

external positive control to prove the functionality of the reaction mix for amplification of the pathogen target This combination rules out inhibition, among other malfunctions, and confirms that a

negative result is truly negative

External Controls

An external control is an independent control that provides a true challenge to the system or process being testing to ensure this system or process is working properly

Quality Control Testing vs. Proficiency TestingQuality Control Testing• Verifies a test or equipment

is providing correct results• Should be conducted on a

routine basis• Cultures have been tested

and designed for quality control testing

• Quality Control material producers can be accredited – Microbiologics is ISO 13485, ISO 17025, and ISO 17034

Proficiency Testing• Verifies an individual is

performing a test properly and obtaining the correct results

• Should be conducted periodically

• PT samples are designed as unknown samples to be identified and then discarded

• Proficiency Providers

Microbiologics Product Offering

Inactivated(full process from

extraction to detection)

Genomic DNA(Amplification and

Detection)

Synthetic(Amplification and

Detection)

Helix Elite™ Molecular Standards

Microbiologics Molecular Product Overview

Microbiologics Molecular Product Offering

Instrument/Test Specific QC Sets and Panels

(ex. BioFire, BD MAX, Cepheid)

Inactivated Helix Elite pellet format (ex. BD MAX Enteric Parasite

Verification Panel)

Inactivated Helix Elite swab format (ex. Cepheid Xpert CT/NG Control

Panel)

KWIK-STIK format (ex. Bruker MALDI Biotyper CA System

Comprehensive Identification Panel)

Strain-Specific (ex. Influenza A, CT/NG)

Inactivated Helix Elite Molecular Standards

(ex. Influenza A)

Synthetic Helix Elite Molecular Standards

(ex. HPV)

KWIK-STIK/LYFO DISK

(ex. C. diff)

Inactivated Helix Elite™ Molecular Standards

• Format: lyophilized pellet containing an inactivated organism– Medium to high titer– Unassayed/qualitative

• Configuration: 5 pellets individually packaged to avoid cross contamination

Why a Full Process Control?• Full process controls, from extraction through detection, are

important because they are:– Used as a surrogate for real samples to test efficiency of the extraction

process– Used as “Spike-In Controls” to determine the effect of the matrix on the

overall recovery– Processed at the same time and in the same manner as the patient

sample to show test effectiveness

Why a Full Process Control?

• There are over 20 steps required in an extraction procedure. By including a positive control, it can:

oDetect human or system errors

oEvaluate new extraction kits using independent controls

How It Works

Applications• QC of commercial molecular testing systems• Verification and Validation of Laboratory Developed Tests

(LDTs)• Sensitivity• Specificity • Assay Optimization• Training• Proficiency • Lot-to-Lot Testing

Synthetic Helix Elite™ Molecular Standards

• Format: Synthetic RNA and DNA that contain consensus sequences of diagnostic regions from the target’s genome, representing the known genetic diversity of the microorganism

• Configuration: 1 vial of dried synthetic DNA or RNA (approximately 100 reactions) and 1 vial of molecular standard water for rehydration

Why Prepared DNA?• Saves time: extraction of whole organisms can be

very labor intensive and time consuming• Minimizes risk of

contamination: PCR is very sensitive. One DNA strand could produce a false-positive result. DNA can contaminate skin, gloves, clothing, and environment

Why Synthetic Helix Elite?

Manual Methods Microbiologics SolutionsTime consuming (from 3 hours to 1 day)

Ready-to-use upon arrival

Cross contamination risk DNA already extracted which helps to avoid cross contamination

Requires skilled technicians to perform extraction

Easy-to-use so minimal training is required

Live organism (health risk) Non-viable (safe)

How It Works

How It Works

Applications

• QC of commercial molecular testing systems

• Sensitivity• Specificity • Assay Optimization• Training• Proficiency • Lot-to-Lot Testing

QC Sets and Panels

• Each instrument-specific QC set or panel contains the manufacturer recommended quality control strains for a particular instrument or test in an easy-to-use format

• Each regulation-specific QC set or panel contains the regulation-specific (e.g. CLSI or EUCAST) recommended quality control strains for a particular test in an easy-to-use format

Packaging Configurations

• Live Culture QC Sets and Panels– KWIK-STIK™ 2 Pack (2 STIKs of

each microorganism)• Synthetic DNA/RNA QC Sets

and Panels• Inactivated Microorganism

QC Sets and Panels

Most Popular Molecular QC Sets & Panels

• BD MAX™

• BioFire FilmArray®

• Cepheid GeneXpert®

BD Control Products: KWIK-STIK

• Refrigerated storage• Configured for

verification purposesCat # Product Name Configuration BD Assay

8179 Enteric Bacterial Organism Set (Live Culture)

20 strains, one KWIK-STIK of each (20 KWIK-STIKs total) BD MAX™ Enteric Bacterial Panel

8174 MRSA Organism Set (Live Culture)

10 Staphylococcus strains, four KWIK-STIKs of each (40 KWIK-STIKs total) BD MAX™ MRSA Assays

8178 SA Organism Set (Live Culture) 14 Staphylococcus strains, one KWIK-STIK of each (14 KWIK-STIKs total)

BD MAX™ StaphSR and BD MAX™ MRSA XT Assays

8173 Strep B Organism Set (Live Culture)

10 Streptococcus strains, four KWIK-STIKs of each (40 KWIK-STIKs total) BD MAX™ GBS Assay

BD Control Products: Inactivated Pellets• Full process controls• Pre-pooled when

applicable to mimic processing of patient sample

• Ready-to-use• Room temperature storage• In vitro diagnostic (IVD)

How it Works?

• Transfer pellet to sample buffer tube

• Vortex, invert, vortex• Process according to BD

insert

BD Control Products: Inactivated Pellets

Cat # Product Name BD Assay8193 BD MAX™ CT/GC/TV 20-Day QC Panel BD MAX™ CT/GC/TV8202 BD MAX™ Enteric Parasite 20-Day QC Panel BD MAX™ Enteric Parasite Panel8204 BD MAX™ Enteric Parasite Control Panel BD MAX™ Enteric Parasite Panel

8191 Extended Enteric Bacterial Verification Panel (Inactivated Pellet) BD MAX™ Extended Enteric Bacterial Panel

8208 Vaginal Verification Panel BD MAX™ Vaginal Panel8209 Vaginal Control Panel BD MAX™ Vaginal Panel8210 Enteric Viral Verification Panel BD MAX™ Enteric Viral Panel8211 Enteric Viral Control Panel BD MAX™ Enteric Viral Panel8214 Human Papillomavirus (HPV) Control Panel BD Onclarity HPV Assay (BD Viper)8216 Human Papillomavirus (HPV) Verification Panel BD Onclarity HPV Assay (BD Viper)

BioFire Control Products

• Full process control• Pre-pooled pellets to mimic the

processing of a patient sample• Ready-to-use• Room temperature storage• In vitro diagnostic (IVD)

BioFire FilmArray Inactivated ControlsAttributes Verification Ongoing QC

Name Blood Culture Identification (BCID) Verification Panel

Blood Culture Identification (BCID) Control Panel

Cat # 8201 8215

Format Helix Elite (Inactivated Material) Helix Elite (Inactivated Pellet)

Components– 3 pools (1 vial of each pool)

• 27 targets (22 organisms in panel)– 3 vials of hydration buffer

6 pellets each pellet containing 22 organisms (27 targets identified)

Respiratory Panel 2 (RP2) Control Panel Coming Soon!

How Does the BCID Verification Panel Work?

1. Transfer 1.2 mL of water to each vial.2. Vortex to dissolve material.3. Start testing based on BioFire’s Verification Protocol.4. Store hydrated materials at 2°C-8°C for up to 72 hours for

use throughout the remainder of the verification testing.

How Does the BCID Control Panel Work?

• Hydrate pellet• Vortex• Process according to

BioFire’s insert

Cepheid GeneXpert Products

• Inactivated swab format that mimics the processing of a patient sample

• Positive and negative controls included in each kit

• Ready-to-use• Room temperature storage• In vitro diagnostic (IVD)

All Infectious Disease Assays Covered!

Catalog # QC Panel Description8200 Cepheid Xpert® C. difficile/epi Control Panel8187 Carbapenem-resistant enterobacteriaceae (CRE) Control Panel (Inactivated

Swab)8188 Cepheid Xpert® CT/NG Control Panel8190 Enterovirus (EV) Control Panel (Inactivated Swab)8194 Cepheid Xpert® GBS LB Control Panel8195 Cepheid Xpert® MRSA NxG Control Panel8196 Cepheid Xpert® SA Nasal Complete Control PanelComing in 2019 Cepheid Xpert® MTB/RIF Control Panel8205 Norovirus Control Panel (Inactivated Swab)8199 Cepheid Xpert® Respiratory Control Panel8189 Trichomonas vaginalis (TV) Control Panel (Inactivated Swab)8203 Vancomycin Resistant Enterococcus (VRE) Control Panel (Inactivated Swab)

How Does it Work?

Instructions for Sample Preparation Reagent

• Break Swab in Vial or Transport Medium Vial (depending on assay insert)

• Vortex• Transfer to cartridge

and test per assay instructions!

Instructions for Direct Inoculation

with Dry or Pre-Wet Swab

• Break swab in cartridge

• Test per assay instructions!

SummaryA good QC program is the foundation of your labMicrobiologics products help streamline your QC process to optimize efficiency and your budget– Our molecular controls feature:

oRoom temperature storageoPre-pooled when applicable for ease-of-use and time savingso IVD

Questions