Abbot SARS-CoV-2 Instructions Manual

Abbott RealTime

en

SARS-CoV-2

09N77-095

51-608445/R4

09N77-095

51-608445/R4

NOTE: Changes highlighted

For use under an Emergency Use Authorization (EUA) Only

Instructions for Use

INTRODUCTION

This Emergency Use Authorization (EUA) package insert must be read carefully prior to use. EUA package insert instructions must be followed
accordingly. Reliability of EUA assay results cannot be guaranteed if there are any deviations from the instructions in this package insert.

NAME

Abbott RealTime SARS-CoV-2

INTENDED USE

The Abbott RealTime SARS-CoV-2 assay is a real-time (rt) reverse transcriptase (RT) polymerase chain reaction (PCR) test intended for the
qualitative detection of nucleic acid from SARS-CoV-2 in nasal swabs, self-collected at a health care location or collected by a healthcare worker
and nasopharyngeal (NP) and oropharyngeal (OP) swabs, and bronchoalveolar lavage fluid (BAL) collected by a healthcare worker, from individuals
suspected of COVID-19 by their healthcare provider.
Testing is limited to laboratories certified under the Clinical Laboratory Improvement Amendments of 1988 (CLIA), 42 U.S.C. §263a, that meet
requirements to perform high complexity tests.
Results are for the identification of SARS-CoV-2 RNA. The SARS-CoV-2 RNA is generally detectable in respiratory specimens during the acute phase
of infection. Positive results are indicative of the presence of SARS-CoV-2 RNA; clinical correlation with patient history and other diagnostic information
is necessary to determine patient infection status. Positive results do not rule out bacterial infection or co-infection with other viruses. Laboratories
within the United States and its territories are required to report all positive results to the appropriate public health authorities.

Negative results do not preclude SARS-CoV-2 infection and should not be used as the sole basis for patient management decisions. Negative results
must be combined with clinical observations, patient history, and epidemiological information.

The Abbott RealTime SARS-CoV-2 assay is intended for use by qualified and trained clinical laboratory personnel specifically instructed and trained in
the techniques of real-time PCR and in vitro diagnostic procedures. The Abbott RealTime SARS-CoV-2 assay is only for use under the Food and Drug
Administration’s Emergency Use Authorization.

SUMMARY AND EXPLANATION OF THE TEST

The Abbott RealTime SARS-CoV-2 assay is real-time reverse transcription polymerase chain reaction (rRT-PCR) test on the Abbott m2000 System. The
SARS-CoV-2 primer and probe sets are designed to detect RNA respiratory specimens collected from patients who are suspected of COVID-19 by their
health care provider.

BIOLOGICAL PRINCIPLES OF THE PROCEDURE

The Abbott RealTime SARS-CoV-2 assay consists of 2 reagent kits:

• Abbott RealTime SARS-CoV-2 Amplification Reagent Kit

• Abbott RealTime SARS-CoV-2 Control Kit

The Abbott RealTime SARS-CoV-2 assay is a dual target assay for the RdRp and N genes.
An RNA sequence that is unrelated to the SARS-CoV-2 target sequence is introduced into each specimen at the beginning of sample preparation. This

unrelated RNA sequence is simultaneously amplified by RT-PCR and serves as an internal control (IC) to demonstrate that the process has proceeded
correctly for each sample.

The Abbott RealTime SARS-CoV-2 assay detects the SARS-CoV-2 virus and IC target sequences through the use of target-specific fluorescent-labeled
oligonucleotide probes. The probes do not generate a signal unless they are specifically bound to the amplified product. The two SARS-CoV-2-specific
probes are labeled with the same fluorophore and the IC-specific probe is labeled with a different fluorophore, thus allowing for simultaneous detection
of both SARS-CoV-2 and IC amplified products in the same reaction well.

The Abbott RealTime SARS-CoV-2 assay is performed on the Abbott m2000 System consisting of a sample preparation unit, the Abbott m2000sp, and
an amplification and detection unit, the Abbott m2000rt. Application parameters specific to the Abbott RealTime SARS-CoV-2 assay are contained on an
assay-specific application specification file, distributed electronically, stored on portable media and loaded onto the Abbott m2000sp and Abbott m2000rt
instruments.

Sample Preparation

The Abbott m2000sp provides automated sample preparation using a magnetic microparticle-based protocol and reagents (Abbott mSample
Preparation System

During the sample preparation protocol, SARS-CoV-2 virions are disrupted by guanidine isothiocyanate, nucleic acids are captured on the magnetic
microparticles, and inhibitors and unbound sample components are removed by washing steps. The bound nucleic acids are eluted off the
microparticles with buffer and transferred to a 96 deep-well plate. The nucleic acids are then ready for amplification. The Internal Control (IC) is
introduced into each specimen at the beginning of the sample preparation process to demonstrate that the process was completed correctly for each
specimen and control.

A positive control and a negative control are processed from the start of sample preparation for each test order to evaluate run validity.
The purpose of sample preparation is to extract and concentrate the target nucleic acids to make the target accessible for amplification, and to

remove potential inhibitors of amplification from the extract.

) to process respiratory specimens.

DNA

1

The Abbott mSample Preparation System
sample components. The bound nucleic acids are eluted and transferred to a 96 deep-well plate. The nucleic acids are then ready for amplification.
The IC is taken through the entire sample preparation procedure along with the controls and specimens.

The Abbott m2000sp automated instrument system is used to prepare samples for the Abbott RealTime SARS-CoV-2 assay. The Abbott m2000sp
provides automated sample eluate transfer and reaction assembly in the Abbott 96-Well Optical Reaction Plate.

uses magnetic particle technology to capture nucleic acids and washes the particles to remove unbound

DNA

Reagent Preparation and Reaction Plate Assembly

The Abbott m2000sp combines the Abbott RealTime SARS-CoV-2 assay amplification reagent components (SARS-CoV-2 Oligonucleotide Reagent,
Thermostable rTth Polymerase Enzyme, and Activation Reagent). The Abbott m2000sp dispenses the resulting master mix to the Abbott 96-Well
Optical Reaction Plate along with aliquots of the nucleic acid samples prepared by the Abbott m2000sp. The plate is ready, after manual application of
the optical seal, for transfer to the Abbott m2000rt.

Amplification

During the amplification reaction on the Abbott m2000rt, the target RNA is converted to cDNA by the reverse transcriptase activity of the thermostable
rTth DNA polymerase. First, the SARS-CoV-2 and IC reverse primers anneal to their respective targets and are extended during a prolonged incubation
period. After a denaturation step, in which the temperature of the reaction is raised above the melting point of the double-stranded cDNA : RNA product,
a second primer anneals to the cDNA strand and is extended by the DNA polymerase activity of the rTth enzyme to create a double-stranded DNA
product.

During each round of thermal cycling, amplification products dissociate to single strands at high temperature allowing primer annealing and extension
as the temperature is lowered. Exponential amplification of the product is achieved through repeated cycling between high and low temperatures,
resulting in a billion-fold or greater amplification of target sequences. Amplification of the three targets (SARS-CoV-2 RdRp, SARS-CoV-2 N, and IC)
takes place simultaneously in the same reaction.

The target sequences for the Abbott RealTime SARS-CoV-2 assay are in the SARS-CoV-2 RdRp and N genes of the SARS-CoV-2 genome. The
selected target sequences are highly conserved and also specific to this strain of coronavirus.

The IC target sequence is derived from the hydroxypyruvate reductase gene from the pumpkin plant, Cucurbita pepo, and is delivered in an Armored
RNA® particle that has been diluted in negative human plasma.

Detection

During the read cycles of amplification on the Abbott m2000rt, the temperature is lowered further to allow fluorescent detection of amplification
products as the SARS-CoV-2 and IC probes anneal to their targets (real-time fluorescence detection). The SARS-CoV-2 probes have a fluorescent
moiety that is covalently linked to the 5′ end and has a quencher molecule at its 3′ end. In the absence of target sequences, the probes adopt a
conformation that brings the quencher close enough to the excited fluorophore to absorb its energy before it can be fluorescently emitted. When
the probe binds to its complementary sequence in the target, the fluorophore and the quencher are held apart, allowing fluorescent emission and
detection. The IC probe is a single-stranded DNA oligonucleotide with a fluorophore at the 5′ end and a quencher at the 3′ end. In the absence of
IC target sequences, probe fluorescence is quenched. In the presence of IC target sequences, probe hybridization to complementary sequences
separates the fluorophore and the quencher and allows fluorescent emission and detection. The SARS-CoV-2 and IC specific probes are each labeled
with a different fluorophore, thus allowing for simultaneous detection of both amplified products.

PREVENTION OF NUCLEIC ACID CONTAMINATION

The possibility of nucleic acid contamination is minimized because:

• Reverse transcription, PCR amplification, and oligonucleotide hybridization occur in a sealed Abbott 96-Well Optical Reaction Plate.

• Detection is carried out automatically without the need to open the Abbott 96-Well Optical Reaction Plate.

• Pipettes with aerosol barrier tips or disposable transfer pipettes are used for all pipetting. The disposable pipettes or pipette tips are discarded

after use.

• Separate, dedicated areas are used to perform the Abbott RealTime SARS-CoV-2 assay. Refer to the SPECIAL PRECAUTIONS section of this
package insert.

REAGENTS

Abbott RealTime SARS-CoV-2 Amplification Reagent Kit (List No. 09N77-095)

1. Abbott RealTime SARS-CoV-2 Internal Control

(4 vials, 1.2 mL per vial)

• < 0.01% noninfectious Armored RNA with internal control sequences in negative human plasma. Negative human plasma tested and found to be
non-reactive by appropriate FDA-licensed, approved, or cleared tests for antibody to HCV, antibody to HIV-1, antibody to HIV-2, HIV-1 Ag, HBsAg,
and Syphilis. The material is also tested and found to be negative by appropriate FDA-licensed, approved, or cleared PCR methods for HIV RNA,
HCV RNA, and HBV DNA. Preservatives: 0.1% ProClin® 300 and 0.15% ProClin 950.

2. Abbott RealTime SARS-CoV-2 Amplification Reagent Pack (List No. 9N77)
(4 packs, 24 tests/pack)

• 1 bottle (0.141 mL) Thermostable rTth Polymerase Enzyme (2.9 to 3.5 Units/µL) in buffered solution.

• 1 bottle (1.0 mL) SARS-CoV-2 Amplification Reagent containing synthetic oligonucleotides (6 primers and 3 probes), and dNTPs in a buffered

solution with a reference dye. Preservative: 0.10% ProClin 300 and 0.15% ProClin 950.

• 1 bottle (0.400 mL) Activation Reagent. 30 mM manganese chloride solution. Preservatives: 0.10% ProClin 300 and 0.15% ProClin 950.

Abbott RealTime SARS-CoV-2 Control Kit (List No. 09N77-085)

1. Abbott RealTime SARS-CoV-2 Negative Control
(8 vials, 1.3 mL per vial) Contains 1.0% ammonium sulfate and 7.9% detergent in a buffer solution.

2. Abbott RealTime SARS-CoV-2 Positive Control
(8 vials, 1.3 mL per vial) Contains non-infectious, recombinant Sindbis virus containing SARS-CoV-2 RNA sequences, 1.0% ammonium sulfate, and

7.9% detergent in a buffer solution.

WARNINGS AND PRECAUTIONS

For Use Under An Emergency Use Authorization Only.

This assay is only for in vitro diagnostic use under the FDA Emergency Use Authorization.
For Prescription Use Only.

• This test has not been FDA cleared or approved;

• This test has been authorized by FDA under an EUA for use by authorized laboratories;

• This test has been authorized only for the detection of nucleic acid from SARS-CoV-2, not for any other viruses or pathogens; and

• This test is only authorized for the duration of the declaration that circumstances exist justifying the authorization of emergency use of in vitro

diagnostic tests for detection and/or diagnosis of COVID-19 under Section 564(b)(1) of the Federal Food, Drug and Cosmetic Act, 21 U.S.C.

§ 360bbb-3(b)(1), unless the authorization is terminated or revoked sooner.

2

Safety Precautions

Refer to the Abbott m2000sp and Abbott m2000rt Operations Manuals, Hazard Section, for instructions on safety precautions.
Important information regarding the safe handling, transport and disposal of this product is contained in the Safety Data Sheet.

CAUTION: This preparation contains human sourced and/or potentially infectious components. Components sourced from human blood have
been tested and found to be nonreactive by appropriate FDA-licensed, approved, or cleared tests for antibody to HCV, antibody to HIV-1, antibody to
HIV-2, HIV-1 Ag, HBsAg, and Syphilis. The material is also tested and found to be negative by appropriate FDA-licensed, approved, or cleared PCR
methods for HIV RNA, HCV RNA, and HBV DNA. No known test method can offer complete assurance that products derived from human sources or
inactivated microorganisms will not transmit infection. These reagents and human specimens should be handled as if infectious using laboratory safety
procedures, such as those outlined in Biosafety in Microbiological and Biomedical Laboratories,1 OSHA Standards on Bloodborne Pathogens,2 CLSI
Document M29-A4,3 and other appropriate biosafety practices.4 Therefore all human sourced materials should be considered infectious.

These precautions include, but are not limited to, the following:

• Wear gloves when handling specimens or reagents.

• Do not pipette by mouth.

• Do not eat, drink, smoke, apply cosmetics, or handle contact lenses in areas where these materials are handled.

• Clean and disinfect spills of specimens by including the use of a tuberculocidal disinfectant such as 1.0% sodium hypochlorite or other suitable

disinfectant.

• Decontaminate and dispose of all potentially infectious materials in accordance with local, state, and federal regulations.

Components of the Abbott RealTime SARS-CoV-2 Internal Control, Oligonucleotide Reagent, and Activation Reagent contain the following components:
2-Methyl-4-isothiazol-3-one:

• Reaction mass of: 5-chloro-2-methyl-4-isothiazolin-3-one (EC no. 247-500-7) and 2-methyl-2H-isothiazol-3-one (EC no. 220-239-6)(3:1)

• Reaction mass of: 5-chloro-2-methyl-4-isothiazolin-3-one (EC no. 247-500-7) and 2-methyl-4-isothiazolin-3-one (EC no. 220-239-6)(3:1)

Potassium Hydroxide
The following warnings apply:

1

4

Warning

H 317 May cause an allergic skin reaction.

H319 Causes serious eye irritation.

H315 Causes skin irritation.

H402 Harmful to aquatic life.*

H412 Harmful to aquatic life with long lasting effects.

P261 Avoid breathing mist/vapors/spray.

P264 Wash hands thoroughly after handling.

P272 Contaminated work clothing should not be allowed out of the workplace.

P280 Wear protective gloves/protective clothing/eye protection.

P273 Avoid release to the environment,

P302+P352 IF ON SKIN: Wash with plenty of water.

P333+P313 If skin irritation or rash occurs: Get medical advice/attention.

P362+P364 Take off contaminated clothing and wash before reuse.

P305+P351+P338 IF IN EYES: Rinse cautiously with water for several minutes. Remove

contact lenses, if present and easy to do. Continue rinsing.

P337+P313 If eye irritation persists: Get medical advice/attention.

P501 Dispose of contents/container in accordance with local regulations.

* Not applicable where regulation EC 1272/2008 (CLP) has been implemented.

Important information regarding the safe handling, transport, and disposal of this product is contained in the Safety Data Sheet.
Safety Data Sheets are available from your Abbott Representative.

SPECIAL PRECAUTIONS

As with any test procedure, good laboratory practice is essential to the proper performance of this assay. Due to the high sensitivity of this test, care
should be taken to keep reagents and amplification mixtures free of contamination.

• For in vitro diagnostic use under Emergency Use Authorization only.

• Positive results are indicative of the presence of SARS-CoV-2 RNA.

• Laboratories within the United States and its territories are required to report all positive results to the appropriate public health authorities.

• All patient samples should be handled as if infectious, using good laboratory procedures as outlined in Biosafety in Microbiological and Biomedical

Laboratories1 and in the CLSI Document M29-A4.3 Only personnel proficient in handling infectious materials and the use of the Abbott RealTime
SARS-CoV-2 assay and the Abbott m2000 System should perform this procedure.

Handling Precautions for Specimens

• The Abbott RealTime SARS-CoV-2 assay is only for use with respiratory specimens that have been handled and stored as described in the

SPECIMEN COLLECTION, STORAGE, AND TRANSPORT TO THE TEST SITE section.

• Inadequate or inappropriate specimen collection, storage, and transport are likely to yield false test results. Training in specimen collection is highly

recommended due to the importance of specimen quality. Refer to CLSI MM13-A 5 as an appropriate resource.

• During preparation of samples, compliance with good laboratory practices is essential to minimize the risk of cross-contamination between samples

and the inadvertent introduction of ribonucleases (RNases) into samples during and after the extraction procedure.

• Proper aseptic technique should always be used when working with RNA.

• Amplification technologies such as PCR are sensitive to accidental introduction of product from previous amplification reactions. Incorrect results

could occur if either the clinical specimen or the reagents used become contaminated by accidental introduction of even a few molecules of
amplification product. Measures to reduce the risk of contamination in the laboratory include physically separating the activities involved in
performing PCR in compliance with good laboratory practices.

3

Work Areas

15°C

-25°C

15°C

The m2000sp and the m2000rt instruments may be operated in the same location. The use of 2 dedicated areas (Sample Preparation Area and
Amplification Area) within the laboratory is recommended when performing the Abbott RealTime SARS-CoV-2 assay.

The Sample Preparation Area is dedicated to processing samples (specimens and Abbott RealTime SARS-CoV-2 Controls) and to adding processed
samples and controls to the 96-Well Optical Reaction Plate. All reagents used in the Sample Preparation Area should remain in this dedicated area at
all times. Laboratory coats, pipettes, pipette tips, and vortexers used in the Sample Preparation Area must remain in this area and not be moved to the
Amplification Area. Do not bring amplification product into the Sample Preparation Area.

The Amplification Area is dedicated to the amplification and detection of amplified product. Laboratory coats and equipment used in the Amplification
Area must remain in this area and not be moved to the Sample Preparation Area.

• Components contained within a kit are intended to be used together. Do not mix components from different kit lots. For example, do not use the

negative control from control kit lot X with the positive controls from control kit lot Y.

• Do not use kits or reagents after the expiration dates shown on kit labels.

• Work area and instrument platforms must be considered potential sources of contamination. Change gloves after contact with potential

contaminants (specimens, eluates, and/or amplified product) before handling unopened reagents, negative control, positive controls, or specimens.
Refer to the Abbott m2000sp and Abbott m2000rt Operations Manuals for instrument cleaning procedures.

• If the Abbott m2000sp instrument run is aborted, dispose of all commodities and reagents according to the Abbott m2000sp Operations Manual.

• If the Abbott m2000sp master mix addition protocol is aborted, seal the Abbott 96-Well Optical Reaction Plate in a sealable plastic bag and

dispose according to the Abbott m2000sp Operations Manual, Hazards section, along with the gloves used to handle the plate.

• If the Abbott m2000rt instrument run is interrupted or aborted, seal the Abbott 96-Well Optical Reaction Plate in a sealable plastic bag and dispose

according to the Abbott m2000rt Operations Manual along with the gloves used to handle the plate.

• Decontaminate and dispose of all potentially biohazardous materials in accordance with local, state, and federal regulations.4 All materials should

be handled in a manner that minimizes the chance of potential contamination of the work area.

NOTE: Autoclaving the sealed Reaction Plate will not degrade the amplified product and may contribute to the release of the amplified product

by opening the sealed plate. The laboratory area can become contaminated with amplified product if the waste materials are not carefully

handled and contained.

Aerosol Containment

To reduce the risk of nucleic acid contamination due to aerosols formed during manual pipetting, aerosol barrier pipette tips must be used for all
manual pipetting. The pipette tips must be used only 1 time. Clean and disinfect spills of specimens and reagents as stated in the Abbott m2000sp
and Abbott m2000rt Operations Manuals.

Contamination and Inhibition

The following precautions should be observed to minimize the risks of RNase contamination, cross-contamination between samples, and inhibition:

• Wear appropriate personal protective equipment at all times.

• Use powder-free gloves.

• Change gloves after having contact with potential contaminants (such as specimens, eluates, and/or amplified product).

• To reduce the risk of nucleic acid contamination due to aerosols formed during pipetting, pipettes with aerosol barrier tips must be used for all

pipetting. The length of the tip should be sufficient to prevent contamination of the pipette barrel. While pipetting, care should be taken to avoid
touching the pipette barrel to the inside of the sample tube or container. The use of extended aerosol barrier pipette tips is recommended.

• Change aerosol barrier pipette tips between ALL manual liquid transfers.

• The Abbott mSample Preparation System

reagents for every new Abbott RealTime SARS-CoV-2 assay run. At the end of each run, discard all remaining reagents from the worktable as
stated in the Abbott m2000sp Operations Manual and the Abbott mSample Preparation System

reagents are single use only. Use new reagent troughs or vessels, reaction vessels, and newly opened

DNA

product information sheet.

DNA

STORAGE INSTRUCTIONS

Abbott RealTime SARS-CoV-2 Amplification Reagent Kit (List No. 09N77-095)

-

• Abbott RealTime SARS-CoV-2 Amplification Reagent Packs and Internal Control (IC) vials must be stored at – 25 to – 15°C when
not in use. Care must be taken to separate the Abbott RealTime SARS-CoV-2 Amplification Reagent Pack that is in use from direct
contact with samples and controls.

Abbott RealTime SARS-CoV-2 Control Kit (List No. 09N77-085)

-

-25°C

• The Abbott RealTime SARS-CoV-2 Negative and Positive Controls must be stored at – 25 to – 15°C.

SHIPPING CONDITIONS

• Abbott RealTime SARS-CoV-2 Amplification Reagent Kit: Ship on dry ice.

• Abbott RealTime SARS-CoV-2 Control Kit: Ship on dry ice.

If you receive reagents that are in a condition contrary to label recommendation, or that are damaged, contact your Abbott Representative.

INDICATION OF INSTABILITY OR DETERIORATION OF REAGENTS

When a positive or negative control value is out of the expected range, it may indicate deterioration of the reagents. Associated test results are invalid
and samples must be retested.

SPECIMEN COLLECTION, STORAGE, AND TRANSPORT TO THE TEST SITE

Human respiratory specimens may be used with the Abbott RealTime SARS-CoV-2 assay. Refer to the CDC Interim Guidelines for Collecting,
Handling, and Testing Clinical Specimens from Persons Under Investigation (PUIs) for Coronavirus Disease 2019 (COVID-19) 6 https://www.cdc.gov/
coronavirus/2019-nCoV/lab/guidelines-clinical-specimens.html or the FDA FAQs on Diagnostic Testing for
SARS-CoV-2 https://www.fda.gov/medical-devices/emergency-situations-medical-devices/faqs-diagnostic-testing-sars-cov-2

An Abbott multi-Collect Specimen Collection Kit (List No. 09K12-01 (CE), 09K12-02 (CE), 09K12-03 or 09K12-04) or Abbott Universal Collection Kit
(List No. 09N77-055) can be used for the transport of nasopharyngeal swab specimens or the collection and transport of nasal and oropharyngeal
swab specimens from the collection site to the testing laboratory. Neither the swab nor the transfer pipette are authorized for nasopharyngeal
specimen collection. The Transport Tube contains Specimen Transport Buffer which is used to stabilize nucleic acid until sample preparation.
Transport and store transport tube at 2 to 25°C for up to 48 hours. If delivery and processing exceed 48 hours, specimens should be transported in dry
ice and once in laboratory frozen at –70°C or colder.

4