Not Intended for Use in Canada

INSTRUCTIONS FOR USE HIV c

VITROS Immunodiagnostic Products 684 2779
HIV Combo Reagent Pack
VITROS Immunodiagnostic Products 684 2780
HIV Combo Calibrator

Intended Use
For in vitro diagnostic use only.

VITROS Immunodiagnostic Products HIV Combo Reagent Pack

For the simultaneous qualitative detection of antibodies to Human Immunodeficiency Virus types 1, including group M and
O, and/or 2 (anti-HIV-1 and anti-HIV-2) and HIV p24 antigen in human serum and plasma (heparin and EDTA) in adults,
pregnant women, adolescents and children, using the VITROS ECi/ECiQ/3600 Immunodiagnostic Systems and the
VITROS 5600/XT 7600 Integrated Systems.
The results of the VITROS HIV Combo test, in conjunction with other serological evidence and clinical information, may be
used as an aid in the diagnosis of infection with HIV-1 and/or HIV-2 in persons at high and low risk for HIV infection and as
a screening test for the detection of HIV-1 and/or HIV-2 in blood donors.

VITROS Immunodiagnostic Products HIV Combo Calibrator

For use in the calibration of the VITROS ECi/ECiQ/3600 Immunodiagnostic Systems and the VITROS 5600/XT 7600
Integrated Systems for the simultaneous qualitative detection of antibodies to Human Immunodeficiency Virus types 1,
including group M and O, and/or 2 (anti-HIV-1 and anti-HIV-2) and HIV p24 antigen in human serum and plasma (heparin
and EDTA) in adults, pregnant women, adolescents and children.

Summary and Explanation of the Test

Acquired Immunodeficiency Syndrome (AIDS) is caused by at least two types of Human Immunodeficiency Viruses
designated HIV–1 and HIV–2. The VITROS HIV Combo test uses 3 recombinant antigens derived from HIV–1 envelope
(env 13), HIV-1 group O envelope (env 70-3) and HIV–2 envelope (env 31). These antigens detect antibodies to HIV–1 and
antibodies to HIV–2 in the same test. The use of these recombinant antigens improves test specificity by avoiding non
specific reactions due to cross-reaction with human cell proteins which are present in cell lysates. The VITROS HIV Combo
test also uses antibodies to HIV p24 antigen to enable detection of HIV p24 antigen that may be present in early
seroconversion before the onset of antibody response enabling earlier diagnosis of HIV infection.

Principles of the Procedure

An immunometric technique is used; this involves a two stage reaction. In the first stage HIV antibody or antigen present in
the sample binds with biotinylated HIV recombinant antigen or biotinylated antibody coated on streptavidin wells. Unbound
sample is removed by washing. In the second stage horseradish peroxidase (HRP)-labeled recombinant HIV antigens and
antibodies are added in the conjugate reagent. The conjugate binds specifically to any human anti-HIV–1 or anti-HIV–2 (IgG
and IgM) or HIV antigen captured on the well in the first stage. Unbound conjugate is removed by washing.
The bound HRP conjugate is measured by a luminescent reaction. 1 A reagent containing luminogenic substrates (a luminol
derivative and a peracid salt) and an electron transfer agent is added to the wells. The HRP in the bound conjugate
catalyzes the oxidation of the luminol derivative, producing light. The electron transfer agent (a substituted acetanilide)
increases the level of light produced and prolongs its emission. The light signals are read by the system. The amount of
HRP conjugate bound is indicative of the amount of HIV antibody and/or p24 antigen present.

Reaction Sample
Test Type System * Incubation Time Time to first result Test Temperature Volume
ECi/ECiQ, 3600,
Immunometric 37 minutes 48 minutes 37 °C 80 μL
5600, XT 7600
* Not all products and systems are available in all countries.

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Warnings and Precautions

Reaction Scheme

Warnings and Precautions

WARNING: Potentially Infectious Material

Treat as if capable of transmitting infection.

Use caution when handling material of human origin. Consider all samples
potentially infectious. No test method can offer complete assurance that hepatitis B
virus, hepatitis C virus (HCV), human immunodeficiency virus (HIV 1+2) or other
infectious agents are absent. Handle, use, store and dispose of solid and liquid
waste from samples and test components, in accordance with procedures defined
by appropriate national biohazard safety guideline or regulation (e.g. CLSI
document M29). 2

The VITROS HIV Combo Calibrator contains:

HIV antibody negative plasma obtained from donors who were tested individually
and who were found to be negative for hepatitis B surface antigen, and for
antibodies to hepatitis C virus (HCV) and HIV, using approved methods (enzyme
immunoassays).

The HIV antibody positive plasma has been treated in order to reduce the titer of
potentially infectious virus. However, as no testing method can rule out the risk of
potential infection, handle as if capable of transmitting infection.

WARNING: Contains Mixture, 3(2H)-isothiazolone, 5-chloro-2-methyl- with 2-methyl-3(2H)-

isothiazolone (CAS 55965-84-9) 3

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Reagents

The VITROS HIV Combo Assay Reagent and VITROS HIV Combo Conjugate
Reagent contain 0.03% Mixture, 3(2H)-isothiazolone, 5-chloro-2-methyl- with 2-
methyl-3(2H)-isothiazolone respectively. H317: May cause an allergic skin reaction.
H319: Causes serious eye irritation. H411: Toxic to aquatic life with long lasting
effects. P261: Avoid breathing dust/fume/gas/mist/vapors/spray. P273: Avoid
release to the environment. P280: Wear protective gloves and eye protection. P333
+ P313: If skin irritation or rash occurs: Get medical advice/attention. P337 + P313:
If eye irritation persists: Get medical advice/attention. P391: Collect spillage.

WARNING WARNING

WARNING: Contains 2-Methyl-3-isothiazolone (CAS 2682-20-4) 3

The VITROS HIV Combo Calibrator contains 0.095% 2-Methyl-3-isothiazolone.

H317: May cause an allergic skin reaction. P261: Avoid breathing dust/fume/gas/
mist/vapors/spray. P280: Wear protective gloves. P333 + P313: If skin irritation or
rash occurs: Get medical advice/attention. P362 + P364: Take off contaminated
clothing and wash it before reuse. P501: Dispose of contents/container to an
approved waste disposal plant.

Refer to www.Orthoclinicaldiagnostics.com for the Safety Data Sheets and for

Ortho contact information.

WARNING

Reagents
Reagent Pack Contents
1 reagent pack containing:
• 100 coated wells (streptavidin, bacterial; binds ≥3 ng biotin/well) (biotin-mouse monoclonal anti-HIV p24, 0.3 μg/mL and
biotin-recombinant HIV antigens, 0.1025 μg/mL)
• 6.2 mL assay reagent (buffer with bovine gamma globulin, bovine serum albumin and antimicrobial agent)
• 16.2 mL conjugate reagent (HRP-recombinant HIV antigens, 0.021–0.266 μg/mL and HRP-mouse monoclonal anti-HIV
p24, 1.5 μg/mL) in buffer with goat serum, bovine serum albumin and antimicrobial agent

Reagent Pack Handling

• The reagent pack is supplied ready for use.
• The reagent pack contains homogeneous liquid reagents that do not require shaking or mixing prior to loading on the
system.
• As with all immunoassay protein-based solutions, inappropriate handling of the reagent pack can cause foam to occur on
the surface of the reagent. Avoid agitation, which may cause foaming or the formation of bubbles.
– If reagent packs are dropped or agitated, small levels of fine foam could be generated that may not be detected by
the system.
– Reagent packs containing fine foam that is not detected by the system, may show a negative bias.
– If you must use a dropped or agitated reagent pack before it has been allowed to settle, you should verify
performance by running high and low quality control samples in duplicate after loading the pack on the system.

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Specimen Collection, Preparation and Storage

Reagent Pack Storage and Preparation

Reagent Storage Condition Stability
Unopened Refrigerated 2–8 °C (36–46 °F) expiration date
Opened On system System turned on ≤12 weeks
Opened Refrigerated 2–8 °C (36–46 °F) ≤12 weeks
• The VITROS HIV Combo Reagent Pack is suitable for use until the expiration date on the carton when stored and
handled as specified. Do not use beyond the expiration date.
• Do not freeze unopened reagent packs.
• Load reagent packs directly from refrigerated storage to minimize condensation.
• Store opened refrigerated reagent packs in a sealed reagent pack storage box that contains dry desiccant.

Calibrator Contents
• 1 VITROS HIV Combo Calibrator (anti-HIV-1 positive human plasma in anti-HIV 1+2 negative human plasma, 2.0 mL)
with antimicrobial agent
• Lot calibration card
• Protocol card
• 8 calibrator bar code labels

Calibrator Handling
• Use only with reagent packs of the same lot number. Mix thoroughly by inversion and bring to 15–30 °C (59–86 °F)
before use. Each pack contains sufficient volume for a minimum of 6 determinations of each calibrator.
• Handle calibrators in stoppered containers to avoid contamination and evaporation. To avoid evaporation, limit the
amount of time calibrators are on the system. Refer to the operating instructions for your system.
• Return to 2–8 °C (36–46 °F) as soon as possible after use, or load only sufficient volume for a single determination.

Calibrator Storage and Preparation

Calibrator Storage Condition Stability
Unopened Refrigerated 2–8 °C (36–46 °F) expiration date
Opened Refrigerated 2–8 °C (36–46 °F) ≤13 weeks
Opened Frozen ≤-20 °C (≤-4 °F) ≤13 weeks
• VITROS HIV Combo Calibrator is supplied ready for use.
• The VITROS HIV Combo Calibrator is suitable for use until the expiration date on the carton when stored and handled as
specified. Do not use beyond the expiration date.
• The opened calibrator may be stored frozen (with no more than 1 freeze-thaw cycle).
• The VITROS HIV Combo test uses 80 μL of calibrator for each determination. The VITROS HIV Combo Calibrator may
be used directly on the VITROS Immunodiagnostic and VITROS Integrated Systems. Alternatively, transfer an aliquot of
each calibrator into a sample container (taking account of the minimum fill volume of the container), which may be bar
coded with the labels provided. For details on minimum fill volume of sample cups or containers, refer to the operating
instructions for your system.
• The VITROS HIV Combo Calibrator is automatically processed in duplicate.

Specimen Collection, Preparation and Storage

Patient Preparation
No special patient preparation is necessary.

Specimens Recommended
• Serum
• Serum Separator Tube (SST)
• Plasma Separator Tube (PST)
• Lithium heparin plasma
• Sodium heparin plasma
• Potassium EDTA plasma
• Sodium citrate plasma
• Acid Citrate Dextrose (ACD)
• Citrate Phosphate Dextrose (CPD)

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Testing Procedure

• Citrate Phosphate Dextrose Adenine (CPDA-1)

Note: The liquid citrate anticoagulants tested (sodium citrate plasma, ACD, CPD and
CPDA-1) have no clinically significant effect on negative samples. However HIV
reactive samples will show proportionally lower signal/cutoff values, due to dilution
by the liquid anticoagulant. High negative results (0.70–0.99) obtained on samples
collected with these anticoagulants should be interpreted accordingly.
Supplemental tests may be required.

Specimens Not Recommended

Do not use turbid specimens. Turbidity in specimens may affect test results.

Special Precautions

IMPORTANT: Certain collection devices have been reported to affect other analytes and tests. 4
Owing to the variety of specimen collection devices available, Ortho Clinical
Diagnostics is unable to provide a definitive statement on the performance of its
products with these devices. Confirm that your collection devices are compatible
with this test.

Specimen Collection and Preparation

• Collect specimens using standard procedures. 5, 6
• Samples should be thoroughly separated from all cellular material. Failure to do so may lead to an erroneous result.
• Thoroughly mix samples by inversion and bring to 15–30 °C (59–86 °F) before use.
• The VITROS HIV Combo test uses 80 μL of sample for each determination. This does not take account of the minimum
fill volume of the chosen sample container. For details on minimum fill volume of sample cups or containers, refer to the
operating instructions for your system.

Handling and Storage Conditions

• Handle samples in stoppered containers to avoid contamination and evaporation.
• The amount of time samples are on the system prior to analysis should be limited to avoid evaporation. Refer to the
operating instructions for your system.
• Return to 2–8 °C (36–46 °F) as soon as possible after use, or load sufficient volume for a single determination.
• Serum, lithium heparin plasma and potassium EDTA plasma samples may be stored for up to 24 hours at room
temperature (up to 30 °C [86°F]) or 7 days at 2–8 °C (36–46 °F).
• SST, PST, sodium heparin plasma, sodium citrate plasma, ACD, CPD and CPDA-1 samples may be stored for up to 24
hours at room temperature (up to 30 °C [86 °F]).
• Samples that will not be tested within the time frames outlined above should be stored at -20 °C [- 4°F] and may be
subjected to up to five freeze-thaw cycles.

IMPORTANT: Thoroughly mix thawed samples by multiple inversions or by vortex mixing and
bring to 15–30 °C (59–86 °F) before use.

Testing Procedure
Materials Provided
• VITROS Immunodiagnostic Products HIV Combo Reagent Pack
• VITROS Immunodiagnostic Products HIV Combo Calibrator

Materials Required but Not Provided

• VITROS Immunodiagnostic Products Signal Reagent
• VITROS Immunodiagnostic Products Universal Wash Reagent
• Quality control materials
• VITROS Immunodiagnostic Products Reagent Pack Storage Box (optional) with desiccant

Operating Instructions
Check the inventory regularly to aid the management of reagents and ensure that sufficient VITROS Signal Reagent,
VITROS Universal Wash Reagent and calibrated reagent lots are available for the work planned. When performing panels
of tests on a single sample, ensure that the sample volume is sufficient for the tests ordered.

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Calibration

For detailed information refer to the operating instructions for your system.

Note: Do not use visibly damaged product.

Default Test Name

The default test name which will appear on patient reports is HIV Combo. The default short name that will appear on the
test selection menus and laboratory reports is HIV c. These defaults may be reconfigured, if required. For detailed
information refer to the operating instructions for your system.

Calibration
Calibration Procedure
• Calibration is lot specific; reagent packs and calibrators are linked by lot number. Reagent packs from the same lot may
use the same calibration.
• A Master Calibration is established for each new reagent lot by performing multiple tests. This is the process by which a
lot-specific parameter [a] which links the signal at the cutoff (cutoff value) to the calibrator signal is determined.
Cutoff value = (a x Signal of Cal 1)
• Ensure that the Master Calibration for each new reagent lot is available on your system.
• Process calibrator in the same manner as samples. Load sufficient for the automatic duplicate determination. Calibration
need not be programmed if bar code labels are used; calibration will be initiated automatically.
• When the calibrator is processed, the validity of the calibration is assessed against quality parameters which compare
the actual signal of the calibrator with the expected signal. If the calibration is acceptable the cutoff value is calculated
and stored for use with any reagent pack of that lot.
• The quality of calibration cannot be completely described by a single parameter. The calibration report should be used in
conjunction with acceptable control values to determine the validity of the calibration.
• Recalibration is required after a pre-determined calibration interval, or when a different reagent lot is loaded.
• Calibration results are assessed against a range of quality parameters. Failure to meet any of the defined quality
parameter ranges will be coded in the calibration report. For actions to be taken following a failed calibration refer to the
operating instructions for your system.
Refer to the operating instructions for your system for detailed instructions on the calibration process.

When to Calibrate
• Calibrate when the reagent pack and calibrator lot changes.
• Calibrate every 28 days.
• After specified service procedures have been performed.
• If quality control results are consistently outside of your acceptable range.
For additional information on when to calibrate, refer to the operating instructions for your system.

Traceability of Calibration
The calibration of the VITROS HIV Combo test is traceable to an in-house reference calibrator which has been value
assigned to optimize the clinical sensitivity and specificity performance.

Calibration Model
Results are calculated as a normalized signal, relative to a cutoff value. During the calibration process a lot-specific
parameter is used to determine a valid stored cutoff value for the VITROS Immunodiagnostic and VITROS Integrated
Systems.

Quality Control
Quality Control Material Selection
External controls may be used in accordance with local, government regulations or accrediting organizations, as applicable.
Controls with suitable levels of anti-HIV-1, anti-HIV-2, anti-HIV-1 group O, and HIV p24 antigen are recommended for use
with the VITROS Immunodiagnostic and VITROS Integrated Systems
The performance of commercial control fluids should be evaluated for compatibility with this test before they are used for
quality control.
Control materials may show a difference when compared with other HIV Ag/Ab methods if they contain high concentrations
of preservatives, stabilizers, or other non-physiological additives, or otherwise depart from a true human sample matrix.
Appropriate quality control value ranges must be established for all quality control materials used with the VITROS HIV
Combo test.

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Results

Quality Control Procedure Recommendations

• Good laboratory practice requires that controls be processed to verify the performance of the test.
• Choose control levels that check the clinically relevant concentrations.
• To verify system performance, analyze control materials:
– After calibration
– According to local regulations or at least once each day that the test is being performed
– After specified service procedures are performed
If quality control procedures within your laboratory require more frequent use of controls, follow those procedures.
• Analyze quality control materials in the same manner as patient specimens.
• If control results fall outside your acceptable range, investigate the cause before deciding whether to report patient
results.
• Refer to published guidelines for general quality control recommendations. 7
For more detailed information, refer to the operating instructions for your system.

Quality Control Material Preparation and Storage

Refer to the manufacturer’s product literature for preparation, storage, and stability information.

Results
Results are automatically calculated by the VITROS Immunodiagnostic and VITROS Integrated Systems.

Result Calculation

Signal for test sample

Result =
Cutoff value

Interpretation of Results
Patient sample results will be displayed with a "Negative", "Retest?" or “Reactive” label.

Result (s/c) <0.90 ≥0.90 and <1.00 ≥1.00

Result Text Negative Retest? Reactive

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Limitations of the Procedure

A sample found “Retest?” or “Reactive” should be retested in duplicate to verify its status. If results on repeat testing are
<1.00 for both replicates, the sample should be considered negative. If either retest result is ≥1.00, the sample should be
considered Reactive and tested by supplemental tests to confirm the result. A repeatedly reactive sample, confirmed by
supplemental tests must be considered positive for anti-HIV-1 (including Group O), and/or anti HIV-2 antibody and/or p24
antigen. In the case of repeatedly “Retest?” results, analysis of follow-up samples is recommended.
The following table summarizes the interpretation of results obtained with the VITROS HIV Combo test on the VITROS
Immunodiagnostic and VITROS Integrated Systems.
VITROS HIV
Combo Test
Result (s/c) Action Interpretation
No further
<0.90
testing Specimen is negative for anti-HIV-1 (including Group O), anti-HIV-2 and p24 antigen.
Specimen is negative for anti-HIV-1 (including Group O), anti-HIV-2 and p24 antigen if
Retest in both duplicate results are <1.00 s/c. Specimen is reactive for anti-HIV-1 (including
≥0.90 and <1.00
duplicate Group O), and/or anti-HIV-2 and/or p24 antigen if 1 or both duplicate results are ≥1.00
s/c.
Retest in Specimen is reactive for anti-HIV-1(including Group O), and/or anti-HIV-2 and or p24
≥1.00
duplicate antigen if 1 or both duplicate results are ≥1.00 s/c.
• If a specimen is reactive the probability that HIV antibodies or antigen are present is high, especially in subjects at high
risk for HIV infection. After confirming the HIV Combo test result by repeating in duplicate, it is appropriate to investigate
reactive results by additional, more specific tests. Specimens found reactive by the VITROS HIV Combo test and
positive by additional, more specific tests are considered positive for antibodies to HIV-1 (including Group O), and/or
HIV-2 and/or p24 antigen. Clinical correlation is indicated with appropriate counselling, medical intervention and possibly
additional testing to decide whether a diagnosis of HIV infection is accurate.
• Interpretation of results from specimens found to be reactive by the VITROS HIV Combo test and negative by additional,
more specific tests is unclear. Further clarification may be obtained by testing another specimen obtained three to six
months later.
• The magnitude of a VITROS HIV Combo test result cannot be correlated to an endpoint titre.

Limitations of the Procedure

Known Interferences
The VITROS HIV Combo test was evaluated for interference consistent with CLSI document EP7. 8 Commonly encountered
substances were tested on two lots of reagents. Of the compounds tested, none was found to interfere with the clinical
interpretation of the test.
Refer to “Substances that do not Interfere” for a list of compounds tested that did not show interference.

Other Limitations
• The results from this or any other diagnostic test should be used and interpreted only in the context of the overall clinical
picture.
• Heterophilic antibodies in serum or plasma samples may cause interference in immunoassays. 9 These antibodies may
be present in blood samples from individuals regularly exposed to animals or who have been treated with animal serum
products. Results which are inconsistent with clinical observations indicate the need for additional testing.
• Certain drugs and clinical conditions are known to alter antibody concentrations in vivo. For additional information, refer
to one of the published summaries. 10 11 12
• Do not use quality control materials preserved with azide.

Performance Characteristics
Clinical Performance
Sensitivity
500 patient samples previously determined as positive were tested in the VITROS HIV Combo test.
Number Tested Number Reactive
HIV-1 400 400
HIV-2 100 100
Total 500 500

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Performance Characteristics

The sensitivity for this population of samples in the VITROS HIV Combo test was calculated as 100.00% (500/500) with an
exact 95% confidence interval of 99.26% to 100.00%.
In addition, 83 samples known to be infected with HIV-1 group M subtypes were reactive by the VITROS HIV Combo test.
Antibody Subtype Number Tested VITROS HIV Combo Number Reactive
A 5 5
A1 1 1
A2 2 2
B 2 2
C 7 7
D 4 4
F 2 2
F1 2 2
F2 5 5
G 7 7
H 3 3
H/A1 1 1
H/U 1 1
J 4 4
K 3 3
CRF01 2 2
CRF01/AE 5 5
CRF01/CRF15 1 1
CRF02/AG 11 11
CRF02/G 2 2
CRF06 1 1
CRF06/CPX 2 2
CRF09/K 1 1
CRF09/CPX 3 3
CRF11/CPX 2 2
CRF18/CPX 4 4
Total 83 83

Antigen Positive Sample Sensitivity

Fifty one HIV antigen positive samples were tested and shown to be reactive in the VITROS HIV Combo Test.
Detection of HIV-1 Antigen Genotypes
Three HIV-1 group M subtype specimens and 49 viral isolates were tested with the VITROS HIV Combo test. The reactivity
by HIV-1 antigen subtype and country of origin on the VITROS 3600 Immunodiagnostic System and a commercially
available CE marked 4th generation Ag/Ab combo test is presented in the table below.

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Performance Characteristics

Commercially Available 4th

VITROS HIV Combo Test Generation HIV Ag/Ab Test
Number of Number of Reactive Number of Reactive
Source (# of Specimens) Subtype Specimens Tested Interpretations (%) Interpretations (%)
Group M Specimens (3)
Uganda A 1 1 (100) 1 (100)
Uganda D 1 1 (100) 1 (100)
Romania F 1 1 (100) 1 (100)
Viral Isolates* (49)
Ghana (1), Kyrgyzstan(1), Uganda
(2) A 4 4 (100) 4 (100)
Brazil (1), Thailand (1), USA (6) B 8 8 (100) 7 (87.5)
Djibouti (1), Ethiopia (1), Senegal
(1), Somalia (1), Uganda (1), C 7 5 (71.4) 5 (71.4)
Zambia (1), Unknown (1)
Senegal (1), Uganda (2) D 3 3 (100) 3 (100)
Brazil (3), Romania (2) F 5 5 (100) 5 (100)
Kenya (1), Democratic Republic of
G 2 2 (100) 1 (50.0)
the Congo (1)
Democratic Republic of the Congo
H 1 1 (100) 1 (100)
(1)
Indonesia (2), Thailand (8) CRF01_AE 10 10 (100) 10 (100)
Djibouti (2), Liberia (1) CRF02_AG 3 3 (100) 3 (100)
Cameroon (2), USA (1), Spain (1) Group O 4 4 (100) 4 (100)
Unknown IIIB 1 1 (100) 0 (0.0)
Unknown HIV-2 1 1 (100) 1 (100)
Total 52 50 (96.2) 47 (90.4)
* Isolates tested after dilution to 200,000 RNA copies/mL in Defibrinated, Delipidized Plasma

Detection of HIV-1 p24 Antigen Standards

Sensitivity was determined by testing serial dilutions of the NIBSC HIV-1 p24 Antigen Standard (90/636) and the AFSSAPS
HIV-1 p24 Antigen Standard in three determinations using two reagent lots across all the VITROS Immunodiagnostic and
Integrated Systems. The overall sensitivity of the VITROS HIV Combo test for the NIBSC HIV-1 p24 Antigen Standard
(90/636) was ≤0.48 IU/mL. The overall sensitivity of the VITROS HIV Combo test for the AFSSAPS HIV-1 p24 Antigen
Standard was ≤13.1 pg/mL.

Seroconversion Panels
Thirty four commercially available seroconversion panels were tested on both the VITROS HIV Combo Test and a
commercially available CE-marked 4th generation Ag/Ab combo test. Results for the thirty four panels are summarized in
the following table. The table presents the number of reactive panel members, the days from first bleed to first reactive
result and the difference in days to first reactive between the two tests.

Days to Evidence of HIV Infection

Number of Reactive Panel Members Days to First Reactive Result
Commercially Available Commercially Available
VITROS HIV 4th Generation HIV VITROS HIV 4th Generation HIV Difference in Days to
Panel ID Combo Test Ag/Ab Test Combo Test Ag/Ab Test First Reactive Result *
PRB934 3 3 0 0 0
PRB950 3 2 18 21 3
PRB954 2 2 17 17 0
PRB966 3 3 44 44 0
6243 4 4 25 25 0
6244 2 2 27 27 0
6247 4 4 21 21 0
6248 2 2 18 18 0
9021 4 4 47 47 0
9079 17 17 40 40 0

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Performance Characteristics

Number of Reactive Panel Members Days to First Reactive Result

Commercially Available Commercially Available
VITROS HIV 4th Generation HIV VITROS HIV 4th Generation HIV Difference in Days to
Panel ID Combo Test Ag/Ab Test Combo Test Ag/Ab Test First Reactive Result *
HIV9012 4 3 14 16 2
HIV9014 6 6 0 0 0
HIV9077 16 16 42 42 0
HIV9020 3 3 89 89 0
HIV9018 4 3 25 28 3
HIV9015 2 2 30 30 0
PRB955 4 4 3 3 0
PRB930 4 4 0 0 0
PRB951 4 4 8 8 0
PRB963 2 2 17 17 0
HIV12007 6 6 117 117 0
HIV12008 6 5 23 28 5
HIV9013 1 1 25 25 0
HIV9028 2 2 53 53 0
HIV9032 8 7 22 24 2
HIV9075 3 3 22 22 0
HIV9089 3 3 16 16 0
PRB943 5 5 7 7 0
PRB956 2 2 47 47 0
PRB957 2 3 23 16 –7
PRB960 2 2 28 28 0
PRB961 2 2 27 27 0
PRB962 2 2 14 14 0
PRB964 1 1 22 22 0
Total 138 134 931 939 8
*Days to first reactive test result on the commercially available test minus the days to first reactive test result for the VITROS HIV
Combo test
The VITROS HIV Combo Test and the commercially available 4th generation Ag/Ab Combo Test were in agreement for 28
of the 34 panels. The VITROS HIV Combo Test became reactive one bleed earlier for five of the thirty four panels. The
commercially available 4th generation Ag/Ab Combo Test became reactive one bleed earlier for one panel.

Specificity
Samples from 5077 presumed healthy blood donors, and 608 clinical specimens were tested at two external sites in the
VITROS HIV Combo test and another commercially available CE marked 4th generation Ag/Ab Combo Test.
Samples Number of test samples Initially Reactive Repeatedly Reactive Confirmed Reactive
Donor 5077 16 8 0
Clinical 608 1 1 1*
*Sample confirmed as Reactive in a 3rd generation antibody immunoassay, a line immunoassay and a nucleic acid test (NAT). This
sample was excluded from the calculation of specificity.
The specificity of the VITROS HIV Combo test for the donor population was calculated as 99.84% (5069/5077) exact 95%
CI (99.69-99.93%). The specificity of the VITROS HIV Combo test for the clinical population was calculated as 100.00%
(607/607) exact 95% CI (99.39-100.00%).

Potentially Cross-Reacting Subgroups

The VITROS HIV Combo test was evaluated for potential cross-reactivity in HIV negative samples from medical conditions
unrelated to HIV infection. The results are summarized in the table below.

Summary of VITROS HIV Combo Test Results with Potentially Cross-Reacting Samples
Sample Category Number Tested Number Negative Number Reactive
HCV Antigen 6 6 0
HCV Antibody 10 10 0
HBsAg 6 6 0

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Performance Characteristics

Sample Category Number Tested Number Negative Number Reactive

HBc Antibody 10 10 0
HTLV I Antigen 6 6 0
HTLV II Antigen 6 6 0
HTLV I Antibody 6 6 0
HTLV II Antibody 6 6 0
EBV Antigen 6 6 0
EBV Antibody 10 10 0
HSV I Antigen 6 6 0
HSV II Antigen 6 6 0
HSV Antibody 12 12 0
Chlamydia 10 10 0
Gonorrhea 10 10 0
Syphilis 10 10 0
Multiparous Female 10 10 0
Pregnancy (1st Trimester) 8 8 0
Pregnancy ( 2nd Trimester) 8 8 0
Pregnancy (3rd Trimester) 8 8 0
Pre Flu Vaccine 5 5 0
Post Flu Vaccine 5 5 0
Influenza A Antigen 6 6 0
Influenza B Antigen 6 6 0
Rheumatoid Factor (RF) 8 8 0
Human Anti-Mouse Antibody 10 10 0
(HAMA)
Autoimmune Disease 10 10 0
Anti-Nuclear Antibodies 10 10 0
(ANA)
Hemophilia 11 11 0
Dialysis 10 10 0
Yeast Reactive: Candida 10 10 0
Super-oxide Dismutase 1 1 0
(SOD)
Cytomegalovirus Antigen 6 6 0
Cytomegalovirus Antibody 10 10 0
Toxoplasmosis infection 10 10 0
HAV Antigen 6 6 0
HAV Antibody 10 10 0
Rubella infection 10 10 0
Elevated IgG 11 11 0
Elevated IgM 7 7 0
Non-Viral Liver Disease 10 10 0
Pediatric (2-5 yrs) 4 4 0
Pediatric (6-10 yrs) 6 6 0
Pediatric (11-16 yrs) 6 6 0
Pediatric (17-21 yrs) 4 4 0
Elevated Cholesterol 10 10 0
Elevated Total Protein 9 9 0
Elevated Triglycerides 6 6 0

Precision
Precision was evaluated consistent with CLSI dcument EP5. 13 Two replicates each of 14 negative or diluted reactive
patient sample pools and 5 control samplesa were tested on 2 separate occasions per day on at least 20 different days. The
experiment was performed using 2 reagent lots on 2 different systems on the VITROS ECi/ECiQ Immunodiagnostic

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Performance Characteristics

System, VITROS 3600 Immunodiagnostic System and VITROS 5600 Integrated System. The data presented are a
representation of the product performance on each system.

VITROS 3600 Immunodiagnostic System

Within-run* Within-calibration** Within-lab*** No.
Panel Member Mean (S/C) SD CV (%) SD CV (%) SD CV (%) Observ. No. Days
Negative 0.06 0.010 N/A 0.013 N/A 0.013 N/A 88 22
Negative 0.07 0.008 N/A 0.011 N/A 0.012 N/A 88 22
Negative Control 0.07 0.009 N/A 0.014 N/A 0.014 N/A 88 22
Anti-HIV-1 0.56 0.024 4.4 0.051 9.3 0.053 9.3 88 22
Anti-HIV-1 0.98 0.031 3.2 0.076 7.8 0.076 7.6 88 22
Anti-HIV-1 2.17 0.057 2.7 0.121 5.7 0.119 5.4 88 22
Anti-HIV-1
Reactive Control 1.90 0.071 3.8 0.115 6.1 0.116 6.0 88 22
Anti-HIV-2 0.72 0.036 5.1 0.072 10.1 0.073 10.0 88 22
Anti-HIV-2 1.04 0.038 3.7 0.075 7.3 0.074 7.0 88 22
Anti-HIV-2 2.44 0.075 3.1 0.129 5.4 0.126 5.1 88 22
Anti-HIV-2
4.20 0.124 0.190 4.6 0.190 4.5 88 22
Reactive Control 3.0
Anti-HIV-1 Group
0.86 0.044 5.2 0.074 8.7 0.076 8.7 88 22
O
Anti-HIV-1 Group
1.10 0.046 4.2 0.080 7.3 0.081 7.3 88 22
O
Anti-HIV-1 Group
2.38 0.093 4.0 0.131 5.6 0.136 5.6 88 22
O
Anti-HIV-1 Group
O Reactive 3.30 0.106 3.3 0.172 5.3 0.167 88 22
Control 5.0
HIV p24 Ag 0.78 0.018 2.3 0.057 7.4 0.059 7.5 88 22
HIV p24 Ag 1.40 0.028 2.0 0.073 5.3 0.077 5.4 88 22
HIV p24 Ag 3.33 0.049 1.5 0.108 3.3 0.116 3.4 88 22
HIV p24 Ag
1.92 0.033 1.7 0.089 4.7 0.092 4.7 88 22
Reactive Control
* Within-run (repeatability). Between Duplicate precision averaged over all runs.
** Within-calibration. Total precision with weighted components of within-run, between–run and between-day variation.
***Within-lab. A measure of the effect of recalibration on total precision, calculated within reagent lot, using data from at least 4
calibrations.
a The Controls used were Bio-Rad VIROTROL Controls
N/A = Not applicable

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Performance Characteristics

VITROS 5600 Integrated System****

Within-run* Within-calibration** Within-lab*** No.
Panel Member Mean (S/C) SD CV (%) SD CV (%) SD CV (%) Observ. No. Days
Negativeb 0.05 0.007 N/A 0.009 N/A 0.009 N/A 84 21
Negativec 0.07 0.008 N/A 0.009 N/A 0.009 N/A 84 21
Negative Control 0.07 0.014 N/A 0.015 N/A 0.015 N/A 88 22
Anti-HIV-1 0.55 0.024 4.4 0.041 7.6 0.042 7.6 88 22
Anti-HIV-1 0.97 0.033 3.4 0.057 5.9 0.058 6.0 88 22
Anti-HIV-1 2.17 0.048 2.2 0.100 4.6 0.099 4.5 88 22
Anti-HIV-1
1.90 0.056 3.0 0.093 4.9 0.092 4.8 88 22
Reactive Control
Anti-HIV-2 0.70 0.038 5.5 0.061 8.8 0.060 8.6 88 22
Anti-HIV-2 1.03 0.043 4.2 0.061 6.0 0.061 5.9 88 22
Anti-HIV-2 2.45 0.069 2.8 0.124 5.1 0.125 5.1 88 22
Anti-HIV-2
4.26 0.123 2.9 0.156 3.7 0.156 3.6 88 22
Reactive Control
Anti-HIV-1 Group
0.84 0.032 3.9 0.061 7.3 0.061 7.3 88 22
O
Anti-HIV-1 Group
1.09 0.036 3.3 6.2 0.067 6.1 88 22
O 0.067
Anti-HIV-1 Group
2.37 0.095 4.0 0.133 5.6 0.143 6.0 22
O 88
Anti-HIV-1 Group
O Reactive 0.100 3.0 0.131 3.9 0.134 3.9 88 22
Control 3.38
HIV p24 Ag 0.80 0.021 2.7 0.046 5.8 0.046 5.8 88 22
HIV p24 Ag 1.47 0.035 2.4 0.066 4.5 0.068 4.6 88 22
HIV p24 Ag 3.52 0.057 1.6 0.099 2.8 0.115 3.2 88 22
HIV p24 Ag
2.03 0.037 1.8 0.072 3.6 0.076 3.7 88 22
Reactive Control
* Within-run (repeatability). Between Duplicate precision averaged over all runs.
** Within-calibration. Total precision with weighted components of within-run, between–run and between-day variation.
***Within-lab. A measure of the effect of recalibration on total precision, calculated within reagent lot, using data from at least 4
calibrations.
**** Performance characteristics for the VITROS 5600 System are applicable to the VITROS XT 7600 System.
a The Controls used were Bio-Rad VIROTROL Controls
b One test day removed from analysis due to a statistical outlier (1.42 S/C)
c One test day removed from analysis due to a statistical outlier (1.15 S/C)
N/A = Not applicable

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Performance Characteristics

VITROS ECi/ECiQ Immunodiagnostic System

Within-run* Within-calibration** Within-lab*** No.
Panel Member Mean (S/C) SD CV (%) SD CV (%) SD CV (%) Observ. No. Days
Negativeb 0.05 0.004 N/A 0.005 N/A 0.005 N/A 84 21
Negative 0.07 0.005 N/A 0.007 N/A 0.007 N/A 88 22
Negative Control 0.07 0.024 N/A 0.024 N/A 0.025 N/A 88 22
Anti-HIV-1 0.65 0.034 5.2 0.045 6.9 0.046 7.1 88 22
Anti-HIV-1 1.09 0.088 8.1 0.106 9.8 0.105 9.6 88 22
Anti-HIV-1 2.38 0.095 4.0 0.122 5.1 0.122 5.1 88 22
Anti-HIV-1
1.96 0.070 3.6 0.113 5.8 0.115 5.8 88 22
Reactive Control
Anti-HIV-2 0.72 0.030 4.2 0.045 6.3 0.048 6.7 88 22
Anti-HIV-2 1.06 0.045 4.3 0.070 6.7 0.073 6.9 88 22
Anti-HIV-2 2.43 0.076 3.1 0.130 5.4 0.142 5.8 88 22
Anti-HIV-2
4.11 0.175 4.3 0.196 4.8 0.204 5.0 88 22
Reactive Control
Anti-HIV-1 Group
0.96 0.056 0.073 7.7 0.070 7.3 88 22
O 5.9
Anti-HIV-1 Group
1.22 0.048 4.0 0.074 6.1 0.077 6.3 88
O 22
Anti-HIV-1 Group
2.61 0.168 6.5 0.190 7.3 0.190 7.3 88 22
O
Anti-HIV-1 Group
O Reactive 3.49 0.093 2.7 0.139 4.0 0.153 4.4 88 22
Control
HIV p24 Ag 0.93 0.026 2.8 0.045 4.9 0.049 5.3 88 22
HIV p24 Ag 1.65 0.041 2.5 0.069 4.2 0.070 4.2 88 22
HIV p24 Ag 3.87 0.057 1.5 0.120 3.1 0.137 3.5 88 22
HIV p24 Ag
2.29 0.038 1.7 0.066 2.9 0.079 3.4 88 22
Reactive Control
* Within-run (repeatability). Between Duplicate precision averaged over all runs.
** Within-calibration. Total precision with weighted components of within-run, between–run and between-day variation.
***Within-lab. A measure of the effect of recalibration on total precision, calculated within reagent lot, using data from at least 4
calibrations.
a The Controls used were Bio-Rad VIROTROL Controls
b One test day removed from analysis due to a statistical outlier (1.90 S/C)
N/A = Not applicable

Specificity
Substances that do not Interfere
The VITROS HIV Combo test was evaluated for interference consistent with CLSI document EP7. 8 Of the compounds
tested, none was found to interfere with the clinical interpretation of the test in negative and weakly reactive samples at the
concentrations indicated.
Test Substance Maximum Level Tested
Bilirubin (conjugated) 30 mg/dL 0.386 mmol/L
Bilirubin (unconjugated) 30 mg/dL 0.513 mmol/L
Biotin 20 ng/mL 82.0 nmol/L
Hemoglobin 500 mg/dL 0.078 mmol/L
Cholesterol 300 mg/dL 77.7 mmol/L
HAMA 263 ng/mL N/A
IgG 1680 mg/dL 16.80 g/L
RF 3020 IU/mL N/A
Total Protein 10.9 g/dL 109 g/L
Triglycerides 1250 mg/dL 14.13 mmol/L
N/A = Not applicable (alternate units are not provided)

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References

Patent Statements
HIV-1 and HIV-2 recombinant antigens used in the VITROS HIV Combo test are prepared under US license by Grifols
Diagnostic Solutions Inc. under a shared manufacturing agreement.

References
1. Summers M et al. Luminogenic Reagent Using 3-Chloro 4-Hydroxy Acetanilide to Enhance Peroxidase/Luminol
Chemiluminescence. Clin Chem. 41:S73; 1995.
2. CLSI. Protection of Laboratory Workers from Occupationally Acquired Infections; Approved Guideline - Fourth Edition.
CLSI document M29-A4. Wayne, PA: Clinical and Laboratory Standards Institute; 2014.
3. Regulation (EC) No 1272/2008 of the European Parliament and of the Council of 16 December 2008 on classification,
labelling and packaging of substances and mixtures, amending and repealing Directives 67/548/EEC and 1999/45/EC,
and amending Regulation (EC) No 1907/2006.
4. Calam RR. Specimen Processing Separator Gels: An Update. J Clin Immunoassay. 11:86–90; 1988.
5. CLSI. Collection of Diagnostic Venous Blood Specimens. 7th ed.CLSI standard GP41. Wayne, PA: Clinical and
Laboratory Standards Institute; 2017.
6. NCCLS. Procedures and Devices for the Collection of Diagnostic Capillary Blood Specimens; Approved Standard –
Fifth Edition. NCCLS document H4-A5 [ISBN 1-56238-538-0]. NCCLS, 940 West Valley Road, Suite 1400, Wayne, PA
19087-1898 USA, 2004.
7. CLSI. Statistical Quality Control for Quantitative Measurements: Principles and Definitions; Approved Guideline - Third
Edition. CLSI document C24-A3 [ISBN 1-56238-613-1]. CLSI, 940 West Valley Road, Suite 1400, Wayne, PA
19087-1898 USA, 2006.
8. CLSI. Interference Testing in Clinical Chemistry; Approved Guideline – Second Edition. CLSI document EP7-A2 (ISBN
1-56238-584-4). CLSI, 940 West Valley Road, Suite 1400, Wayne, Pennsylvania 19087-1898 USA, 2005.
9. Levinson SS. The Nature of Heterophilic Antibodies and Their Role in Immunoassay Interference , J Clin Immunoassay
15: 108–115 (1992).
10. Young DS. Effects of Drugs on Clinical Laboratory Tests ed. 4. Washington, D.C.: AACC Press; 1995.
11. Friedman RB, Young DS. Effects of Disease on Clinical Laboratory Tests. ed. 3. Washington, D.C.: AACC Press; 1997.
12. Tryding N, Tufvesson C, Sonntag O (eds). Drug Effects in Clinical Chemistry. ed. 7. Stockholm: The National
Corporation of Swedish Pharmacies, Pharmasoft AB, Swedish Society for Clinical Chemistry; 1996.
13. CLSI. Evaluation of Precision Performance of Quantitative Measurement Methods; Approved Guideline – Third Edition.
CLSI document EP5-A3 [ISBN 1-56238-968-8]. CLSI, 950 West Valley Road, Suite 2500, Wayne, PA 19087 USA,
2014.

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Glossary of Symbols

Glossary of Symbols

Revision History
Date of Revision Version Description of Technical Changes*
2022-10-03 9.0 • Warnings and Precautions:
– Updated Hazard and Precaution Statements to align with the new Safety
Data Sheets
– Added Globally Harmonized Symbol to comply with the Classification,
Labelling and Packaging (CLP) Regulations
• Intended Use:
– Polish translation corrected - "quantitative" changed to "qualitative"
– Korean translation corrected - Added "human"
* The change bars indicate the position of a technical amendment to the text with respect to the previous version of the document.

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Revision History

When this Instructions For Use is replaced, sign and date below and retain as specified by local regulations or laboratory
policies, as appropriate.

Signature Obsolete Date

Conditions of supply: all supplies are made subject to the standard terms and conditions of Ortho Clinical
Diagnostics or its distributors. Copies of these are available on request.

0459
Ortho-Clinical Diagnostics
1500 Boulevard Sébastien Brant
B.P. 30335
67411 Illkirch
CEDEX, France

Ortho-Clinical Diagnostics
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Pencoed
Bridgend
CF35 5PZ
United Kingdom
VITROS is a trademark of Ortho Clinical Diagnostics.
© Ortho Clinical Diagnostics, 2017–2022

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