BD Viper™ System with XTR™ Technology
CLSI Laboratory Procedure*
I. INTENDED USE
The BD ProbeTec™ Herpes Simplex Viruses (HSV 1 & 2) Qx Amplified DNA Assays, when tested
with the BD Viper™ System in Extracted Mode, use Strand Displacement Amplification technology for
the direct, qualitative detection and differentiation of Herpes Simplex virus type 1 (HSV1) and Herpes
Simplex virus type 2 (HSV2) DNA in clinician-collected external anogenital lesion specimens. The
assays are indicated for use with symptomatic individuals to aid in the diagnosis of anogenital HSV1
and HSV2 infections.
Warning: The BD ProbeTec Herpes Simplex Viruses (HSV 1 & 2) Qx Amplified DNA
Assays (HSV Qx DNA Assays) are not indicated for use with cerebrospinal fluid (CSF). The
assays are not intended to be used for prenatal screening or for individuals under the age of
17 years.
II. SUMMARY AND EXPLANATION
Herpes Simplex virus type 1 (HSV1) and type 2 (HSV2) are ubiquitous double-stranded neurotropic
DNA viruses of the Herpesviridae family that cause incurable lifelong infections and which result from
inoculation of the virus through abraded skin or mucous membranes. Both viruses may remain latent for
long periods and cause recurrent episodes of symptomatic disease.
The primary modes of transmission for HSV1 are via oral secretions and non-genital contact resulting in
a predominance of infections of the oropharynx, face, eyes and central nervous system. In recent years
the frequency of genital HSV1 infections has increased due to a lower rate of oral infection during
prepubescent childhood that renders individuals susceptible to genital infection in later life and a rise in
frequency of oro-genital contact.1,2 Nevertheless, seroprevalence studies show that up to 80% of children
infected with HSV1 by adulthood, with the highest rates among those in poor socioeconomic groups.3
In contrast with HSV1, infection with HSV2 is usually the result of sexual transmission. In the United
States, 20-25% of the population has antibodies to HSV2 by the age of 40 and overall there are at least
50 million people with genital herpes.3 In the majority of cases, symptoms are mild or unrecognized and
the infection remains undiagnosed, although intermittent shedding of infectious virus into the genital
tract still occurs. As a result, most transmission of genital herpes occurs through sexual contact by
persons who are asymptomatic or are unaware that they are infected. Infection with HSV2 is more
common in women than men and is linked to an increased risk of sexually transmitted Human
Immunodeficiency Virus (HIV).4 While transmission of herpes to neonates in utero or intrapartum is
rare; the consequences of such infections are severe and frequently fatal.
Classical primary genital herpes infection is preceded by localized pain or tingling, frequently
accompanied by fever, malaise and inquinal lymphadenopathy. Within days, vesicles appear on the labia
minora, introitus and urethra meatus of women and on the shaft and glans of the penis in men. The
perineum and perianal areas may also be affected, as may the upper thighs and buttocks. Cervical lesions
also occur frequently in women.
This “Sample Procedure” is not indicated as a substitute for your facility procedure manual, instrument manual, or reagent
labeling/package insert. This “Sample Procedure” is intended as a model for use by your facility to be customized to meet the needs of your
laboratory.
*
For use with Package Insert: BD ProbeTec Herpes Simplex Viruses (HSV 1 & 2) Qx Amplified DNA Assays [8086121(01) 2012-10]
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CLSI Laboratory Procedure*
Primary infection with HSV1 cannot be distinguished from that caused by HSV2 on clinical grounds
and the lesions may also be confused with those caused by other sexually transmitted diseases. As a
consequence, laboratory testing is required for definitive diagnosis to reduce symptoms and hasten the
healing of lesions. In addition, because the recurrence of HSV1 infections and subclinical shedding are
less frequent than for HSV2, determination of the etiology of infection and typing of the virus is useful
in the assessment of prognosis and counseling.1,2,6 The preferred method of diagnosis of herpes
infection has historically been viral isolation in tissue culture followed by type-specific
immunofluorescent detection; however, the enhanced sensitivity, robustness and rapid time-to-results of
amplified methods for the detection of viral DNA are leading to their increasingly widespread
adoption.1,3,6
When used with the BD Viper System in extracted mode, the BD ProbeTec HSV Qx Amplified DNA
Assays involve automated non-specific extraction of DNA from clinical specimens using chemical lysis
of cells, followed by binding of DNA to magnetic particles, washing of the bound nucleic acid and
elution in an amplification-compatible buffer. When present, HSV1 and/or HSV2 is detected by Strand
Displacement Amplification (SDA) of type-specific target sequences in the presence of a fluorescentlylabeled detector probe.7,8
III. PRINCIPLES OF PROCEDURE
The BD ProbeTec HSV Qx Amplified DNA Assays are designed for use with the BD Qx Swab
Diluent and BD Universal Viral Transport (UVT) [or identical Copan manufactured media
formulations-see COLLECTION KITS PROVIDED SEPARATELY section] specimen collection
and transport devices, applicable reagents, the BD Viper System and BD FOX Extraction. Swab
specimens are collected and transported in the prescribed transport devices which preserve the integrity
of HSV DNA over the specified ranges of temperature and time.
To maintain a consistent workflow with other specimen types that are processed on the BD Viper
System, the HSV specimens undergo a pre-warm step in the BD Viper Lysing Heater. After cooling, the
specimens are loaded onto the BD Viper System which then performs all the steps involved in
extraction and amplification of the target DNA, without further user intervention. The specimen is
transferred to an Extraction Tube that contains ferric oxide particles in a dissolvable film and dried
Extraction Control. A high pH is used to lyse the viruses to liberate their DNA into solution. Acid is
then added to lower the pH and induce a positive charge on the ferric oxide, which in turn binds the
negatively charged DNA. The particles and bound DNA are then pulled to the sides of the Extraction
Tube by magnets and the treated specimen is aspirated to waste. The particles are washed and a high pH
Elution Buffer is added to recover the purified DNA. Finally, a Neutralization Buffer is used to bring the
pH of the extracted solution to the optimum for amplification of the target.
The BD ProbeTec HSV Qx Amplified DNA Assays are based on the simultaneous amplification and
detection of target DNA using amplification primers and a fluorescently labeled detector probe.7,8 The
reagents for SDA are dried down in four separate disposable microwells: the Priming Microwells
contain the assay specific amplification primers, fluorescently labeled detector probe, nucleotides and
other reagents necessary for amplification, while the assay specific Amplification Microwells each
contain the two enzymes (a DNA polymerase and a restriction endonuclease) that are required for SDA.
The BD Viper System pipettes portions of the purified DNA solution from each Extraction Tube into
two separate Priming Microwells to rehydrate their contents, one well corresponding to HSV1 and the
other to HSV2. After a brief incubation, the reaction mixtures are transferred to corresponding, pre-
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CLSI Laboratory Procedure*
warmed Amplification Microwells which are sealed to prevent contamination and then incubated in one
of the two thermally controlled fluorescent readers. The presence or absence of HSV DNA is determined
by calculating the peak fluorescence (Maximum Relative Fluorescence Units [MaxRFU]) over the
course of the amplification process and by comparing this measurement to a predetermined threshold
value.
In addition to the fluorescent probe used to detect amplified HSV target DNA, a second labeled
oligonucleotide is incorporated in each reaction. The Extraction Control (EC) oligonucleotide is labeled
with a different dye than that used for detection of the HSV specific target and is used to confirm the
validity of the extraction process. The EC is dried in the Extraction Tubes and is rehydrated upon
addition of the specimen and extraction reagents. At the end of the extraction process, the EC
fluorescence is monitored by the BD Viper instrument and an automated algorithm is applied to both the
EC and HSV specific signals to report specimen results as positive, negative, or EC failure.
IV. REAGENTS PROVIDED
Each BD ProbeTec HSV Qx Reagent Pack contains:

HSV1 Qx Amplified DNA Assay Priming Microwells, 3 pouches of 32 microwells each: each
Priming Microwell contains approximately 108 pmol oligonucleotides, 27 pmol fluorescentlylabeled detector probe, 150 nmol dNTPs, with stabilizers and buffer components.

HSV1 Qx Amplified DNA Assay Amplification Microwells, 3 pouches of 32 microwells each:
each Amplification Microwell contains approximately 100 units of DNA polymerase and 500
units restriction enzyme, with stabilizers and buffer components.

HSV2 Qx Amplified DNA Assay Priming Microwells, 3 pouches of 32 microwells each: each
Priming Microwell contains approximately 105 pmol oligonucleotides, 36 pmol fluorescentlylabeled detector probe, 120 nmol dNTPs, with stabilizers and buffer components.

HSV Qx Amplified DNA Assay Amplification Microwells, 3 pouches of 32 microwells each:
each Amplification Microwell contains approximately 35 units of DNA polymerase and 500
units restriction enzyme, with stabilizers and buffer components.
NOTE: Each microwell pouch contains one desiccant bag.
MATERIALS PROVIDED SEPARATELY
Control Set for the BD ProbeTec Herpes Simplex Viruses (HSV 1 & 2) Qx Amplified DNA Assays: 24
HSV Qx Positive Control Tubes containing approximately 15,000 copies of pHSV1 and 16,500 copies of
pHSV2 linearized plasmids in carrier nucleic acid, and 24 HSV Qx Negative Control Tubes containing
carrier nucleic acid alone. The concentrations of the pHSV1 and pHSV2 plasmids are determined by UV
spectrophotometry.
Swab Diluent for the BD ProbeTec Qx Amplified DNA Assays: 48 tubes, each containing
approximately 2 mL of potassium phosphate/potassium hydroxide buffer with DMSO and preservative.
BD FOX Extraction Tubes for the BD ProbeTec Qx Amplified DNA Assays: 48 strips of 8 tubes,
each containing approximately 10 mg of iron oxide in a dissolvable film and approximately 240 pmol
fluorescently-labeled Extraction Control oligonucleotide.
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Extraction Reagent and Lysis Trough for the BD ProbeTec Qx Amplified DNA Assays: 12 Reagent and
12 Lysis troughs, each 4-cavity Extraction Reagent trough contains approximately 16.5 mL Binding
Acid, 117 mL Wash Buffer, 35 mL Elution Buffer, and 29 mL Neutralization Buffer with preservative;
each Lysis Trough contains approximately 11.5 mL Lysis Reagent.
V. EQUIPMENT/SUPPLIES PROVIDED SEPARATELY
BD Viper Instrument, BD Viper Instrument Plates, BD Viper Pipette Tips, BD Viper Tip Waste Boxes,
BD Viper Amplification Plate Sealers (Black), BD Viper Lysing Heater, BD Viper Lysing Rack, BD
Viper Neutralization Pouches, and Pierceable Caps for use on the BD Viper System (Extracted Mode),
BD Viper System Accessories.
COLLECTION KITS PROVIDED SEPARATELY
BD ProbeTec Qx Collection Kit for Endocervical and Lesion Specimens for use on the BD Viper
System in Extracted Mode, BD Universal Viral Transport Medium (UVT) (3 mL) with polyester-fibertip swab collection kit (Cat. No. 220221 [kit], 220220 [UVT medium] / 220239 [collection swab]) or
identical Copan Universal Transport Medium* (UTM-RT) and polyester-fiber-tip swab collection kit
(Cat. No. 302C.LC, 302C, 330C, 340C or 321C).
*The collection device and collection medium in the BD UVT kit are identical to the Copan UTM-RT
medium and collection device in the catalog numbers listed.
MATERIALS REQUIRED BUT NOT PROVIDED
Nitrile gloves, 1% (v/v) sodium hypochlorite*, pipettes capable of transferring 0.5 mL, phosphate
buffered saline (PBS).
*Mix 200 mL of bleach with 800 mL of water. Prepare fresh daily.
Storage and Handling Requirements: Reagents may be stored at 2 – 33ºC. Unopened Reagent Packs
are stable until the expiration date. Once a pouch is opened, the microwells are stable for 8 weeks if
properly sealed or until the expiration date, whichever comes first. Do not freeze.
VI. SPECIMEN COLLECTION AND TRANSPORT
The BD ProbeTec HSV Qx Assays on the BD Viper System in Extracted Mode are designed to detect
the presence of Herpes Simplex virus type 1 or Herpes Simplex virus type 2 from external anogenital
lesion specimens.
The devices that can be used to collect lesion specimens for testing on the BD Viper System in
Extracted Mode are:

BD ProbeTec Qx Collection Kit for Endocervical or Lesion Specimens for use with the BD
Viper System in Extracted Mode.

BD Universal Viral Transport Medium (UVT)-3 mL fill volume and regular sized polyesterfiber-tip swab with plastic shaft (Cat. No. 220221).
o Identical Copan Universal Transport Medium (UTM-RT) and polyester-fiber-tip swab
collection kit (Cat. No. 302C.LC, 302C, 330C, 340C or 321C) may also be used.
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For U.S. and international shipments, specimens should be labeled in compliance with applicable state,
federal, and international regulations covering the transport of clinical specimens and etiologic
agents/infectious substances. Time and temperature conditions for storage must be maintained during
transport. Once pre-warmed, Qx swab Diluent and diluted UVT specimens may be stored for up to 120
days at -20ºC prior to testing on the BD Viper System.
VII. LESION SWAB SPECIMEN COLLECTION, STORAGE AND TRANSPORT
The external anogenital lesion specimens for the BD ProbeTec HSV Qx Assays are collected with either
the BD ProbeTec Qx Swab Collection Kit for Endocervical or Lesion Specimens or the UVT collection
device (polyester-tipped swab with plastic shaft used with 3 mL fill volume). A 0.5 mL volume of the
UVT specimen must be added to a Qx Swab Diluent Tube during processing and thus has additional
stability claims. Stability information is provided for each specimen type (Tables 1, 2A, 2B and 3).
External Anogenital Swab Specimen Collection using BD ProbeTec Collection Kit for
Endocervical or Lesion Specimens for use with the BD ProbeTec HSV Qx Amplified DNA Assays
NOTE: All specimens should be obtained from the patient by appropriately trained individuals.
1. Open the inner Qx Swab packaging and dispose of the swab with the white shaft.
2. Remove the pink collection swab from the packaging.
3. Swab the base of the exposed lesion firmly to absorb exudates and cellular material with the
pink swab.
4. Uncap the Qx Swab Diluent tube.
5. Fully insert the pink collection swab into the Qx Swab Diluent tube.
6. Break the shaft of the swab at the score mark. Use care to avoid splashing of contents.
7. Tightly recap the tube.
8. Label the tube with patient information and date/time collected.
9. Transport to the laboratory.
BD ProbeTec Qx Swab Lesion Specimen Storage and Transport
The Qx Swab must be stored and transported to the laboratory and/or test site at either 2-30C for testing
within 14 days of collection, or -20C within 120 days of collection.
Table 1: Stability of Anogenital Lesion Specimens Collected with the BD Qx Swab Collection Kit
BD ProbeTec Qx Collection Kit for Endocervical or Lesion Specimens
Temperature Condition for
Transport to Test Site and
Storage
2 - 30C
-20C
Process Specimen According
to Instructions
 14 days of collection
 120 days of collection
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External Anogenital Swab Specimen Collection using Universal Viral Transport (UVT) Collection
Kit (or equivalent Copan collection kit) for use with the BD ProbeTec HSV Qx Amplified DNA
Assays
NOTE: All specimens should be obtained from the patient by appropriately trained individuals.
1. Remove the cap from the transport medium provided in the transport kit.
2. Expose the base of the lesion.
3. Open the swab packaging and dispose of the second swab if present (either the second swab with
the plastic shaft or the metal shaft minitip swab).
4. Using only one plastic shaft, polyester-fiber-tip swab provided in the UVT transport kit, swab the
lesion firmly to absorb exudates and cellular material at the base of the lesion.
5. Immediately place the swab used to collect the lesion specimen into the transport medium.
6. Break swab shaft by bending it against the vial wall evenly at the pre-scored line.
7. Replace the cap on the vial and close tightly.
8. Label the vial with patient information and date/time collected.
9. Transport to the laboratory.
UVT Lesion Specimen Storage and Transport
If the UVT specimen is collected, stored, and transported prior to transfer to the Qx Swab Diluent, the
UVT specimen may be stored under the conditions in Table 2A.
Table 2A: Storage and Transport of Anogenital Lesion Specimens Collected with UVT Collection
Kit (3 mL fill vial with polyester-fiber-tip swab with plastic shaft)
UVT Specimen
Temperature
Condition for
Transport to Test
Site and Storage
Process Specimen
According to
Instructions
20 - 35C
2 - 8C
-70C
Aliquot and prewarm
within 48 h of
collection
Aliquot and prewarm
within 14 days of
collection
Aliquot and prewarm
within 120 days of
collection
UVT specimen matrix can be stored and transported in the UVT vial within the following conditions:

Up to 48 h at 20 - 25C, or

Up to 14 days at 2 - 8C, or

Up to 120 days at -70C.
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Once the UVT specimen is aliquoted into Qx Swab Diluent, the specimen must be prewarmed.
NOTE: Wear clean gloves when handling the anogenital lesion specimen. If gloves come in contact
with specimen, immediately change gloves to prevent contamination of other specimens.
1. All UVT specimens should be at room temperature prior to processing.
2. Verify that the UVT contains a swab with a white shaft.
3. Remove a pre-filled Qx Swab Diluent Tube from the packaging.
4. Label the pre-filled Qx Swab Diluent Tube with the patient identification and date/time collected
on the UVT specimen.
5. Vortex the UVT specimen for 5 – 10 s.
6. Remove the black pierceable cap from the Qx Swab Diluent Tube.
7. Uncap the UVT specimen transport vial. Avoid touching the swab. Using a pipette, aseptically
transfer 0.5 mL of UVT specimen into the Qx Swab Diluent Tube.
8. Discard the pipette. NOTE: The pipette is intended for use with a single specimen.
9. Tightly recap the Qx Swab Diluent Tube containing 0.5 mL of UVT with a black pierceable cap
and invert 3 -4 times to ensure that specimen and diluent are well mixed.
10. The Qx Swab Diluent Tube containing 0.5 mL of UVT specimen is ready to be pre-warmed.
11. Re-cap the UVT vial using the capture cap containing the swab and store at 2 - 8C or - 70C.
12. Change gloves before proceeding to avoid contamination.
13. Repeat steps 1 – 10 for additional UVT external anogenital lesion specimens.
UVT Specimen Storage and Transport in Qx Swab Diluent
If the UVT specimen is collected and immediately transferred into Qx Swab Diluent, specimens may be
stored under the conditions in Table 2B prior to the pre-warm procedure.
Table 2B: Storage and Transport of UVT Anogenital Lesion Specimens in Qx Swab Diluent
UVT Specimen Transferred into Qx Swab Diluent
Temperature
Condition for
Transport to Test
Site and Storage
Process Specimen
According to
Instructions
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2 - 8C
-20C
Pre-warm within 24 h
after aliquotting
Pre-warm within 14
days of collection
Pre-warm within 120
days of collection
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BD Viper™ System with XTR™ Technology
CLSI Laboratory Procedure*
Specimen Processing Procedure for the BD ProbeTec Herpes Simplex Viruses (HSV 1 & 2) Qx
Amplified DNA Assays on the BD Viper System in Extracted Mode
NOTE: Allow all specimens to completely thaw at room temperature and mix by inversion prior to
proceeding.
1. Verify that the Qx Swab Diluent either contains a swab with a pink shaft or contains UVT
specimen (diluent is pinkish/purple in color). Also confirm that each tube has a black pierceable
cap.
2. Using the tube layout report, place the Qx Swab Diluent Tube in order in the BD Viper Lysing
Rack and lock into place.
3. Repeat steps 1 and 2 for additional external anogenital lesion swab specimens.
4. Specimens are ready to be pre-warmed.
5. Change gloves before proceeding to prevent contamination.
VIII. QUALITY CONTROL
Quality Control must be performed in accordance with applicable local, state and/or federal regulations
or accreditation requirements and you laboratory’s standard Quality Control procedures. It is
recommended that the user refer to pertinent CLSI guidance and CLIA regulations for appropriate
Quality Control practices.
The Control Set for the BD ProbeTec HSV Qx Amplified DNA Assays is provided separately. One
Positive and one Negative Control must be included on each plate and for each new reagent kit lot
number. Controls must be positioned according to the BD Viper Instrument User’s Manual. The HSV
Qx Positive Control will monitor for substantial reagent failure only. The HSV Qx Negative Control
monitors for reagent and/or environmental contamination.
The HSV Qx Positive Control contains cloned HSV1 and HSV2 target regions. These controls may be
used for internal quality control or users may develop their own internal quality control.13 Additional
controls may be tested according to guidelines or requirements of local, state, and/or federal regulations
or accrediting organizations. Refer to CLSI C24-A3 for additional guidance on appropriate internal
control testing practices. 13 The Positive Control contains approximately 15,000 copies per mL of
pHSV1 and 16,500 copies per mL of pHSV2 linearized plasmids.
The Extraction Control (EC) oligonucleotide is used to confirm the validity of the extraction process.
The EC is dried in the Extraction Tubes and is re-hydrated by the BD Viper System upon addition of the
specimen and extraction reagents. At the end of the extraction process, the EC fluorescence is monitored
by the instrument and an automated algorithm is applied to both the EC and the HSV specific signals to
report specimen results as positive, negative, or EC failure.
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QUALITY CONTROL PREPARATION
NOTE: The HSV 1 & 2 Qx Positive and Negative Controls (i.e. assay controls) do not require addition
of fluid by the user prior to loading in the BD Viper Lysing Rack. Do not re-hydrate the controls prior
to loading in the BD Viper Lysing Rack.
1. Using the tube layout report, place HSV Qx Positive Controls in the appropriate positions in the
BD Viper Lysing Rack.
2. Using the tube layout report, place HSV Qx Negative Controls in the appropriate positions in the
BD Viper Lysing Rack.
3. Controls and specimens are ready to be pre-warmed.
Specimen Processing Controls
Specimen processing controls may be tested in accordance with the requirements of appropriate
accrediting organizations. A positive specimen processing control should test the entire assay system.
For this purpose, known positive specimens can serve as controls by being processed and tested in
conjunction with unknown specimens.
Specimens used only as processing controls must be stored, processed, and tested according to the
package insert (see Specimen Collection and Transport). Specimen processing controls may also be
prepared in the laboratory using either commercially available AcroMetrix OptiQual HSV-1 (Cat. No.
95-1301) and AcroMetrix OptiQual HSV-2 (Cat. No. 95-1302) or Herpes Simplex virus 1 (ATCC
VR-539) and Herpes Simplex virus 2 (ATCC VR-734) following the procedure below. Specimens that
will be used as specimen processing controls will each need to be logged in as a specimen.
AcroMetrix Processing Control Procedure
NOTE: It is recommended to verify that controls give appropriate results before use as specimen
processing controls.
1. Thaw a vial of AcroMetrix OptiQual HSV-1 (Cat. No. 95-1301) and AcroMetrix OptiQual
HSV-2 (Cat. No. 95-1302).
2. Add 100 L of OptiQual HSV-1 and/or 100 L of OptiQual HSV-2 Control to a BD ProbeTec
Qx Swab Diluent Tube and tightly recap using a black pierceable cap.
3. Mix the solution by vortexing or with inversion.
4. Using the tube layout report, place the processing control sample in order in the BD Viper
Lysing Rack, log in as a specimen, and lock into place.
5. Process the controls according to the Pre-warming Procedure and then follow the Test Procedure.
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ATCC Processing Control Procedure
If a known positive specimen is not available, another approach is to assay a stock culture of Herpes
Simplex virus 1 ATCC #VR-539 and Herpes Simplex virus 2 ATCC #VR-734 prepared as described
below:
1. Thaw a vial of each Herpes Simplex virus 1 and Herpes Simplex virus 2 received from ATCC.
2. Prepare separate 10-fold serial dilutions for each virus to a 10-3-fold dilution (at least 4 mL final
volume) in phosphate buffered saline (PBS). It is recommended that the 10-3-fold dilution is
tested to confirm that the appropriate HSV1 and HSV2 results are obtained before use as
specimen processing controls.
3. Place 100 L of 10-3-fold dilution of HSV1 and 100 L of 10-3-fold dilution of HSV2 in a BD
ProbeTec Qx Swab Diluent Tube and tightly recap using a black pierceable cap.
4. Mix the solution by vortexing or with inversion.
5. Using the tube layout report, place the processing control sample in order in the BD Viper
Lysing Rack, log in as a specimen, and lock into place.
6. Process the controls according to the Pre-warming Procedure and then follow the Test Procedure.
General QC Information on the BD Viper System
The location of the microwells is shown in a color-coded plate layout screen on the LCD Monitor. The
plus symbol (+) within the microwell indicates the positive QC sample. The minus (-) symbol within the
microwell indicates the negative QC sample.
A QC pair must be logged in for each plate to be tested and for each reagent kit lot number. For each
plate, if you have not logged in QC samples, a message box appears that prevents saving the rack and
proceeding with the run until a QC pair is added.
A maximum of two QC pairs per rack is permitted per assay.
Running one plate on a BD Viper System
The first two positions (A1 and B1) are reserved for the positive (A1) and the negative (B1) controls
respectively. The first available position for a patient specimen is C1.
Running two plates on a BD Viper System
For plate one, the first two positions (A1 and B1) are reserved for the positive (A1) and the negative (B1)
controls respectively. The first available position for a patient specimen is C1.
For plate two (full plate) the last two positions (G12 and H12) are reserved for the positive (G12) and
negative (H12) controls, respectively.
For plate two (partial plate) the last two positions after the last patient specimen are assigned as the
positive and negative controls, respectively.
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Refer to BD Viper Instrument User’s Manual for more information.
NOTES:

Do not attempt to hydrate the controls prior to loading them into the BD Viper Lysing Rack. The
BD Viper System will rehydrate the controls during the assay run.

Other control materials may be added, provided they are logged in as specimens. Sample rack
position A1 must contain an HSV 1&2 Qx Positive Control, and sample rack position B1 must
contain an HSV 1&2 Qx Negative Control only. Refer to the BD Viper Instrument User’s
Manual for more information on placement of controls in mixed-batch mode.
IX. PRE-WARM PROCEDURE
NOTE: The pre-warm procedure must be applied once to all specimens to ensure that the specimen
matrix is homogenous prior to loading on the BD Viper System. Failure to pre-warm specimens may
have an adverse impact on the performance of the BD ProbeTec HSV Qx Assays and/or BD Viper
System. Specimens and processing controls (see specimen processing controls) must be pre-warmed;
however pre-warming of the assay controls is optional.
NOTE: Refrigerated or frozen specimens must be brought to room temperature prior to pre-warming.
1. Insert the BD Viper Lysing Rack into the BD Viper Lysing Heater.
2. Pre-warm the specimens for 15 min at 114C  2C.
3. Remove the Lysing Rack from the Lysing Heater and let specimens cool at room temperature for
a minimum of 15 min before loading into the BD Viper instrument.
4. Refer to the Test Procedure for testing specimens and controls.
5. After pre-warming, specimens may be stored frozen for 120 days at -20C or stored up to 7 days
at 2 - 30C without additional pre-warming prior to testing on BD Viper System.
NOTE: Frozen specimens must be thawed to room temperature prior to pre-warming.
Table 3: Post Pre-warm Stability
Post Pre-warm Specimen Stability
Temperature Condition for Storage
After Pre-warming step on BD
Viper According to Instructions
2 - 30C
-20C
Within 7 days of prewarming
Within 120 days of prewarming
X. TEST PROCEDURE
Refer to the BD Viper System Instrument User’s Manual for specific instructions for operating and
maintaining the components of the system. The optimum environmental conditions for the HSV Qx
Assays were found to be 18 - 27C and 20 – 85% Relative Humidity.
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XI. INTEPRETATION OF TEST RESULTS
The BD ProbeTec HSV Qx Amplified DNA Assays use fluorescent energy transfer as the detection
method to test for the presence of Herpes Simplex Type 1 and 2 in clinical specimens. All calculations
are performed automatically by the BD Viper software. The presence or absence of HSV1 or HSV2
DNA determined by calculating the peak fluorescence (MaxRFU) over the course of the amplification
process and by comparing this measurement to a predetermined threshold value. The magnitude of the
MaxRFU score is not indicative of the level of organism in the specimen. If the HSV1 or HSV2 specific
signal is greater than or equal to the threshold of 125 MaxRFU, the EC fluorescence is ignored by the
algorithm. If the HSV1 or HSV2 specific signal is less than a threshold of 125 MaxRFU, the EC
fluorescence is utilized by the algorithm in the interpretation of the result. If assay control results are not
as expected, patient results are not reported. See the Quality Control section for expected control values.
Reported results are determined as follows in Table 5, Interpretation of Test Results for the HSV1 and
HSV2 Qx Assays.
Interpretation of Quality Control Results
The HSV1 and HSV2 Positive Control and the HSV1 and HSV2 Negative Control must test as positive
and negative, respectively, in order to obtain patient results. If Positive and Negative Controls do not
perform as expected, the run is considered invalid and patient results will not be reported by the
instrument. If either of the controls does not provide the expected results, repeat the entire run using a
new set of controls, new extraction tubes, new extraction reagent trough, new lysis trough and new
microwells. If the repeat QC does not provide the expected results, contact BD Technical Services.
See Table 4 for Interpretation of Quality Control Results. For more information, see the BD Viper
Instrument User’s Manual.
Table 4: Interpretation of Quality Control Results
Control Type
Tube Result Report
Symbol
Max RFU
QC Disposition
HSV Positive Control
125
QC Pass
HSV Positive Control
125
QC Failure
Any value
QC Failure
HSV Negative Control
125
QC Pass
HSV Negative Control
125
QC Failure
Any value
QC Failure
HSV Positive Control
HSV Negative Control
or
or
or
or
= Fail,
= Extraction Transfer Failure,
= Error
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= Liquid Level Failure,
= Extraction Control Failure,
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BD Viper™ System with XTR™ Technology
CLSI Laboratory Procedure*
Table 5: Interpretation of Test Results for HSV1 and HSV2 Qx Assays
HSV Qx Assays Interpretation of Results
Tube
Report
Result
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HSV Qx
MaxRFU
Report
Interpretation
Result
Positive for HSV
DNA
Positive
125
HSV DNA detected by SDA
125
HSV DNA not detected by SDA
Negative for
HSV DNA
Negative
125
Extraction control failure. Repeat
test from initial specimen tube or
obtain another specimen for
testing.
Non-reportable
result
Extraction
Control Failure
Any value
Extraction Transfer Failure.
Repeat test from initial specimen
tube or obtain another specimen
for testing.
Non-reportable
result
Extraction
Transfer Failure
Any value
Liquid Level Failure. Repeat test
from initial specimen tube or
obtain another specimen for
testing.
Non-reportable
result
Liquid Level
Failure
Any value
Error. Repeat test from initial
specimen tube or obtain another
specimen for testing.
Non-reportable
result
Error
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BD Viper™ System with XTR™ Technology
CLSI Laboratory Procedure*
XII. MONITORING FOR THE PRESENCE OF DNA CONTAMINATION
At least monthly, the following test procedure should be performed to monitor the work area and
equipment for the presence of DNA contamination. Environmental monitoring is essential to detect
contamination prior to the development of a problem.
1. For each are to be tested, use a clean pink collection swab from the BD ProbeTec Qx Collection
Kit for Endocervical or Lesion Specimens.
2. Dip the pink collection swab into the BD ProbeTec Qx Swab Diluent Tube and wipe the first
area* using a broad sweeping motion.
3. Fully insert the pink collection swab into the BD ProbeTec Qx Swab Diluent Tube.
4. Break the shaft of the pink swab at the score mark. Use care to avoid splashing of contents.
5. Tightly recap the tube using the black pierceable cap.
6. Repeat for each desired area.
7. After all swabs have been collected and processed according to the Pre-warming Procedure,
follow the Test Procedure.
* Recommended areas to test include: Instrument deck: Pipette Tip Station Covers (2); Tube
Processing Station; Tube Alignment Block and Fixed Metal Base; Deck Waste Area, Priming and
Warming Heaters/Stage; Extraction Block; Plate Sealing Tool; Tip Exchange Stations (2); Instrument
Exterior: Upper Door Handle; Lower Door Handle; Waste Liquid Quick Release Valve; LCD Monitor
(Touchscreen); Keyboard/Scanner; Staging Area; Locking Plate and Fixed Metal Base; Accessories:
Tube Lockdown Cover, BD Viper Lysing Rack/Table Base; BD Viper Lysing Heater; Metal Microwell
Plates; Timer, Laboratory Bench Surfaces.
If an area gives a positive result or if contamination is suspected, clean the area with fresh 1% (v/v)
sodium hypochlorite. Make sure the entire area is wetted with the solution and allowed to remain on the
surface for at least 2 min or until dry. If necessary, remove excess cleaning solution with a clean towel.
Wipe the area with a clean towel saturated with water and allow the surface to dry. Retest the area.
Repeat cleaning process until negative results are obtained. If the contamination does not resolve,
contact BD Technical Services for additional information.
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BD Viper™ System with XTR™ Technology
CLSI Laboratory Procedure*
XIII. LIMITATIONS OF THE PROCEDURE
1. The device is not indicated for use with cerebrospinal fluid (CSF), endocervical specimens, or
any lesions other than clinician-collected external anogenital lesion specimens.
2. Optimal performance of the test requires adequate specimen collection, transport and handling.
Refer to the “Specimen Collection and Transport” sections of this insert.
3. A negative test does not exclude the possibility of infection because test results may be affected
by improper specimen collection/transport/handling (inadequate specimen collection), presence
of inhibitor(s), technical error, specimen mix-up, concurrent antiviral therapy, or the presence of
insufficient DNA for detection.
4. HSV viability and/or infectivity cannot be inferred from a positive test result since target DNA
may persist in the absence of infectious virus.
5. As with many diagnostic tests, results from the BD ProbeTec HSV Qx Amplified DNA Assays
should be interpreted in conjunction with other laboratory and clinical data available to the
physician.
6. The BD ProbeTec HSV Qx Amplified DNA Assays provide qualitative results. No correlation
can be drawn between the magnitude of the positive assay signal (MaxRFU) and the quantity of
virus in an infected specimen.
7. The predictive value of an assay depends on the prevalence of the disease in any particular
population. [Package Insert] See Table 7 for hypothetical predictive values when testing varied
populations.
8. Because the Positive Control for the BD ProbeTec HSV Qx Amplified DNA Assays is used in
testing for both HSV1 and HSV2, correct positioning of the microwell strips is important for
final results reporting.
9. Use of the BD ProbeTec HSV Qx Amplified DNA Assays is limited to personnel who have been
trained in the assay procedure and the BD Viper System.
10. The reproducibility of the BD ProbeTec HSV Qx Amplified DNA Assays was established using
seeded Qx Swab Diluent and UVT in Qx Swab Diluent to simulate both specimen types
recommended for these assays. [Package Insert] See Tables 16A-16D for reproducibility results.
11. The performance of the BD ProbeTec HSV Qx Amplified DNA Assays has not been evaluated
with specimens from patients less than 17 years of age.
12. This test detects and differentiates between HSV1 and HSV2 only. It does not detect or
differentiate any other Herpes virus types.
13. There is a risk of false negative results due to the presence of sequence variants in the targets of
the assay, procedural errors, amplification inhibitors in specimens, or numbers of organisms
below the claimed limits of the assays.
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BD Viper™ System with XTR™ Technology
CLSI Laboratory Procedure*
XIV. REFERENCES
1. Ryan, C., G. Kinghorn. 2006 Clinical assessment of assays for diagnosis of herpes simplex
infection. Expert Rev Mol Diagn 6: 767-775.
2. Gupta, R., T. Warren, A. Wald. 2007 Genital herpes. Lancet 370: 2127-2137.
3. Fatahzadeh, M., R.A. Schwartz. 2007 Human herpes simplex virus infections: epidemiology,
pathogenesis, symptomology, diagnosis, and management. J Am Acad Dermatol 57: 737-763.
4. Freeman, E., H.A. Weiss, J.R. Glynn, P.L. Cross, J.A. Whitworth and R.J. Hayes. 2006 Herpes
simplex virus 2 infection increases HIV acquisition in men and women: systematic review and
meta-analysis of longitudinal studies. AIDS 20: 73-83.
5. Whitley, R.J., B. Roizman. 2001. Herpes simplex virus infections. Lancet 357: 1513-1518.
6. Centers for Disease Control and Prevention. 2006. Sexually Transmitted Diseases Treatment
Guidelines, 2006. MMWR 55: RR-11.
7. Little, MC, J Andrews, R Moore, et al. 1999. Strand displacement amplification and
homogeneous real-time detection incorporated in a second-generation DNA probe system, BD
ProbeTec ET. Clin Chem 45: 777-784.
8. Hellyer, T.J., J.G. Nadeau. 2004. Strand displacement amplification: a versatile tool for
molecular diagnostics. Expert Rev Mol Diagn 4: 251-261.
9. Clinical and Laboratory Standards Institute. 2005. Approved Guidelines M29-A3. Protection of
laboratory workers from occupationally acquired infections, 3rd ed., CLIS. Wayne, PA.
10. Garner, J.S. 1996. Hospital Infection Control Practices Advisory Committee, U.S. Department of
Health and Human Services, Centers for Disease Control and Prevention. Guideline for isolation
precautions in hospitals. Infect. Control Hospital Epidemiol. 17:53-80.
11. U.S. Department of Health and Human Services. 2007. Biosafety in microbiological and
biomedical laboratories, HHS Publication (CDC), 5th ed. U.S. Government Printing Office,
Washington, D.C.
12. Directive 2000/54/EC of the European Parliament and of the Council 18 September 2000 on the
protection of workers from risks related to exposure to biological agents at work (seventh
individual directive within the meaning of Article 16(1) of Directive 89/391/EEC). Office
Journal L262, 17/10/2000, p. 0021-0045.
13. Clinical and Laboratory Standards Institute. 2006. Approved Guideline C24-A3. Statistical
quality control for quantitative measurement procedures: principles and definitions, 3rd ed. CLSI,
Wayne, PA.
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CLSI Laboratory Procedure*
XV. APPROVALS
Supervisor:________________________________ Date:_________________
Manager:__________________________________ Date:_________________
Director:__________________________________ Date:_________________
Effective Date:___________________
Reviewed by:___________________________________
Package Insert Reference:
BD ProbeTec Herpes Simplex Viruses (HSV 1 & 2) Qx Amplified DNA Assays
[8086121(01) 2012-10]
CLSI Revision: 2013/02
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