HERPESELECT 1 AND 2 PLEXUS IGG, MODEL MP0900G

K062543 · Focus Diagnostics, Inc. · MXJ · Feb 1, 2007 · Microbiology

Device Facts

• Record ID: K062543
• Device Name: HERPESELECT 1 AND 2 PLEXUS IGG, MODEL MP0900G
• Applicant: Focus Diagnostics, Inc.
• Product Code: MXJ · Microbiology
• Decision Date: Feb 1, 2007
• Decision: SESE
• Submission Type: Traditional
• Regulation: 21 CFR 866.3305
• Device Class: Class 2

Indications for Use

Focus Diagnostics’ Plexus™ HerpeSelect®1 and 2 IgG is intended for qualitatively detecting the presence or absence of human IgG class antibodies to HSV-1 and HSV-2 in human sera. The test is indicated for testing sexually active adults or expectant mothers for aiding in the presumptive diagnosis of HSV-1 and HSV-2 infection. The predictive value of a positive or negative result depends on the population's prevalence and the pretest likelihood of HSV-1 and HSV-2 infection. The performance of this assay has not been established for use in a pediatric population, for neonatal screening, for testing of immunocompromised patients, for use by a point of care facility or for use with automated equipment.

Device Story

Plexus HerpeSelect 1 and 2 IgG is a chemiluminescent-based fluorescent bead assay for qualitative detection of human IgG antibodies to HSV-1 and HSV-2 in serum. The device utilizes an antigen bead suspension containing distinct gG1 and gG2 antigen bead types. Procedure: 1) Patient serum incubation with antigen beads; 2) Addition of phycoerythrin-conjugated goat anti-human IgG to form a sandwich; 3) Measurement of fluorescence from each bead type compared against a cut-off calibrator. Requires Luminex xMAP or Bio-rad BioPlex 2200 instrumentation. Used in clinical laboratories to aid in HSV infection diagnosis. Output provides qualitative antibody status, assisting clinicians in patient management and diagnostic decision-making.

Clinical Evidence

Clinical performance evaluated in 300 expectant mothers and 300 sexually active adults. Compared against HerpeSelect 1/2 Immunoblot IgG. Expectant mothers: HSV-1 agreement 96.5% (pos)/92.2% (neg); HSV-2 agreement 94.3% (pos)/95.5% (neg). Sexually active adults: HSV-1 agreement 91.0% (pos)/96.5% (neg); HSV-2 agreement 96.3% (pos)/97.4% (neg). CDC panel (n=100) showed 100% agreement. Reproducibility studies showed %CV ≤ 10%. Cross-reactivity with CMV, EBV, VZV was 0-5%.

Technological Characteristics

Multiplexed immunoassay using antigen bead suspension. Beads fluoresce at distinct wavelengths/intensities. Three-step procedure: incubation with patient sera, addition of phycoerythrin-conjugated goat anti-human IgG, and fluorescence measurement. Standalone laboratory assay; non-automated.

Indications for Use

Indicated for sexually active adults or expectant mothers to aid in the presumptive diagnosis of HSV-1 and HSV-2 infection. Not for pediatric, neonatal, or immunocompromised populations.

Regulatory Classification

Identification

Herpes simplex virus serological assays are devices that consist of antigens and antisera used in various serological tests to identify antibodies to herpes simplex virus in serum. Additionally, some of the assays consist of herpes simplex virus antisera conjugated with a fluorescent dye (immunofluorescent assays) used to identify herpes simplex virus directly from clinical specimens or tissue culture isolates derived from clinical specimens. The identification aids in the diagnosis of diseases caused by herpes simplex viruses and provides epidemiological information on these diseases. Herpes simplex viral infections range from common and mild lesions of the skin and mucous membranes to a severe form of encephalitis (inflammation of the brain). Neonatal herpes virus infections range from a mild infection to a severe generalized disease with a fatal outcome.

Special Controls

*Classification.* Class II (special controls). The device is classified as class II (special controls). The special control for the device is FDA's revised guidance document entitled “Class II Special Controls Guidance Document: Herpes Simplex Virus Types 1 and 2 Serological Assays.” For availability of the guidance revised document, see § 866.1(e).

Predicate Devices

• HerpeSelect 1 and 2 Immunoblot IgG
• HerpeSelect-1 ELISA IgG
• HerpeSelect-2 ELISA IgG

Submission Summary (Full Text)

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# 510(k) SUBSTANTIAL EQUIVALENCE DETERMINATION DECISION SUMMARY ASSAY ONLY TEMPLATE

A. 510(k) Number: K062543
B. Purpose for Submission: New device clearance
C. Measurand: Herpes Simplex Virus
D. Type of Test: Chemiluminescent technology for antibody technology
E. Applicant: Focus Diagnostics
F. Proprietary and Established Names: Plexus™ HerpeSelect®1 and 2 IgG

## G. Regulatory Information:

1. Regulation section: 866.3305
2. Classification: II
3. Product code: MXJ/MYF
4. Panel: Microbiology

## H. Intended Use:

Focus Diagnostics’ Plexus™ HerpeSelect®1 and 2 IgG is intended for qualitatively detecting the presence or absence of human IgG class antibodies to HSV-1 and HSV-2 in human sera. The test is indicated for testing sexually active adults or expectant mothers for aiding in the presumptive diagnosis of HSV-1 and HSV-2 infection. The predictive value of a positive or negative result depends on the population's prevalence and the pretest likelihood of HSV-1 and HSV-2 infection. The performance of this assay has not been established for use in a pediatric population, for neonatal screening, for testing of immunocompromised patients, for use by a point of care facility or for use with automated equipment.

2. Indication(s) for use: NA
3. Special conditions for use statement(s): NA
4. Special instrument requirements: Luminex xMAP® or Bio-rad BioPlex® 2200

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# I. Device Description:

The Focus Diagnostics Plexus HerpeSelect®1 and 2 IgG uses an Antigen Bead suspension that contains two distinct HSV antigen bead types that fluoresce at different wavelengths and/or intensities: gG1 beads and gG2 beads.

The Focus Diagnostics Plexus HerpeSelect®1 and 2 IgG is a three step procedure.

1. Patient sera are diluted, and the diluted sera are incubated with Antigen Beads. If HSV antibodies are present, then the antibodies bind to the corresponding antigen beads.
2. Phycoerythrin—conjugated goat anti-human IgG, (Conjugate) is added, and the Conjugate binds to the bound HSV antibody (if present), and forms a Conjugate-HSV antibody-antigen bead sandwich.
3. Fluorescence from each distinct HSV antigen bead type is measured and compared against a Cut-off Calibrator.

# J. Substantial Equivalence Information:

1. Predicate device name(s): HerpeSelect 1 and 2 Immunoblot IgG, HerpeSelect 1 ELISA IgG, HerpeSelect 2 ELISA IgG
2. Predicate 510(k) number(s): K000238, K02129, K021486
3. Comparison with predicate:

| Item | Device | Predicate |
| --- | --- | --- |
| | Focus Plexus HerpeSelect 1 and 2 IgG | Focus HerpeSelect 1 and 2 Immunoblot IgG |
| Similarities | | |
| Same intended use | Qualitatively detecting the presence or absence of human IgG class antibodies to HSV-1 and HSV-2 in human sera. | Qualitatively detecting the presence or absence of human IgG class antibodies to HSV-1 and HSV-2 in human sera. |
| Same indications for use | The test is indicated for testing sexually active adults or expectant mothers for aiding in the presumptive diagnosis of HSV-1 and HSV-2 infection. | The test is indicated for testing sexually active adults or expectant mothers for aiding in the presumptive diagnosis of HSV-1 and HSV-2 infection. |
| Same type specific HSV-1 antigen | Recombinant gG1 antigen | Recombinant gG1 antigen |
| Same type specific HSV-2 antigen | Recombinant gG2 antigen | Recombinant gG2 antigen |
| Same immunoglobulin type | IgG | IgG |
| Same sample matrix | Serum | Serum |
| Differences | | |
| Methodology | Luminex-based fluorescent bead assay | Immunoblot assay |
| CLIA complexity | High | Moderate |

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K. Standard/Guidance Document Referenced (if applicable): Class II Special Controls Guidance Document: Herpes Simplex Virus Types 1 and 2 Serological Assays (http://www.fda.gov/cdrh/oivd/guidance/1305.pdf)

L. Test Principle:

The Focus Diagnostics Plexus HerpeSelect®1 and 2 IgG uses an Antigen Bead suspension that contains two distinct HSV antigen bead types that fluoresce at different wavelengths and/or intensities: gG1 beads and gG2 beads.

The Focus Diagnostics Plexus HerpeSelect®1 and 2 IgG is a three step procedure.

1. Patient sera are diluted, and the diluted sera are incubated with Antigen Beads. If HSV antibodies are present, then the antibodies bind to the corresponding antigen beads.
2. Phycoerythrin—conjugated goat anti-human IgG, (Conjugate) is added, and the Conjugate binds to the bound HSV antibody (if present), and forms a Conjugate-HSV antibody-antigen bead sandwich.
4. Fluorescence from each distinct HSV antigen bead type is measured and compared against a Cut-off Calibrator.

M. Performance Characteristics (if/when applicable):

1. Analytical performance:

a. Precision/Reproducibility

Inter-laboratory Reproducibility and Inter/Intra-assay Reproducibility Focus, a clinical laboratory located in Southern California, and a university laboratory located in Northern California assessed the device's inter-laboratory reproducibility and inter/intra-assay reproducibility. Each of the three laboratories tested eleven samples in triplicate on five different days.

Inter-laboratory Reproducibility and Inter/Intra-assay Reproducibility

| Sample | HerpeSelect 1 Plexus IgG Results | | | | | HerpeSelect 2 Plexus IgG Results | | | | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | Intra- and Inter-assay | | | Inter-Lab | | Intra- and Inter-assay | | | Inter-Lab | |
| | Mean Index | Intra-assay % CV | Inter-assay % CV | Mean Index | % CV | Mean Index | Intra-assay % CV | Inter-assay % CV | Mean Index | % CV |
| 9 | 4.93 | 3.6 | 10.3 | 4.94 | 3.7 | 3.88 | 3.4 | 10 | 3.87 | 2.5 |
| 6 | 4.24 | 3.8 | 8.7 | 4.22 | 3.3 | 4.9 | 2.6 | 8.5 | 4.89 | 2.1 |
| 2 | 3.87 | 4.8 | 7.9 | 3.86 | 1.3 | 3.36 | 4.3 | 7.7 | 3.35 | 2 |
| 8 | 3.27 | 4.9 | 9.1 | 3.25 | 3 | 4.56 | 3.1 | 8.3 | 4.55 | 1.5 |
| 4 | 3.24 | 4.9 | 7.4 | 3.22 | 2.1 | 2.55 | 4.5 | 8.9 | 2.54 | 5.8 |

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| 1 | 3.04 | 4.3 | 8.9 | 3.02 | 2.3 | 2.71 | 3.8 | 9.3 | 2.7 | 2.1 |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| 12^{b} | 2.13 | 7.9 | 8.7 | 2.13 | 4.1 | 1.87 | 7.2 | 8.8 | 1.87 | 3.4 |
| 3 | 0.34 | 9.1 | 14.9 | 0.34 | 6.8 | 0.06 | 8.7 | 28.3 | 0.06 | 22.6 |
| 10^{c} | 0.19 | 9.9 | 213.1 | 0.19 | 59 | 0.12 | 11.4 | 334.2 | 0.4 | 103.8 |
| 10^{d} | 0.13 | 10 | 15.8 | 0.12 | 1.9 | 0.06 | 11.5 | 41.7 | 0.06 | 38.3 |
| 7 | 0.18 | 8.3 | 16.3 | 0.17 | 9.4 | 0.06 | 8.1 | 23.7 | 0.06 | 17.3 |
| 5 | 0.14 | 9 | 16 | 0.14 | 2.7 | 0.06 | 8.3 | 39.8 | 0.06 | 38.1 |

a. Excludes two runs at one site that were invalid because the Negative Control index was beyond the acceptable QC criteria (it appears that the Positive Control was run in those wells since the indices were about 1.9 for both gG1 and gG2)
b. Samples 12 (inter-lab reproducibility) and 14 (inter-lot reproducibility below) were separate samples, but they were made with the same sera Samples 11 did not have sufficient volume to be sent to investigators.
c. This line includes all data for Sample 10, including one run at Lab 2, where it appears that Sample I may have been run instead since the indices were about 2.7 for both gG1 and gG 2.
d. This line includes all data for Sample 10. except for one run at Lab 2, where it appears that Sample I may have been run instead since the indices were about 2.7 for both gGI and gG2

## Inter-Lot Reproducibility

Focus assessed the device's Inter-lot Reproducibility by testing eleven samples with three separate lots. The samples were run in triplicate. Each lot had a different set of gG-I and gG2 beads, a different lot of conjugate (made from 2 different stock conjugates), and a different lot of calibrator (made from 2 different combinations of positive and negative sera). The results of the studies are summarized in the tables below:

| Sample | HSV-1 | | HSV-2 | |
| --- | --- | --- | --- | --- |
| | Mean | Inter-Lot | Mean | Inter-Lot |
| | Index | %CV | Index | %CV |
| 9 | 5.2 | 7.4 | 3.9 | 12.8 |
| 6 | 4.36 | 8.5 | 4.76 | 9.2 |
| 2 | 3.6 | 7.3 | 3.19 | 9.3 |
| 4 | 3.29 | 7.9 | 2.54 | 6.4 |
| 8 | 3.23 | 11.3 | 4.45 | 9.4 |
| 1 | 3.14 | 5.8 | 2.73 | 4.8 |
| 12/14* | 2.22 | 10.5 | 1.86 | 7.8 |
| 3 | 0.31 | 17 | 0.11 | 50.9 |
| 7 | 0.15 | 31.3 | 0.08 | 21.8 |
| 5 | 0.1 | 45.6 | 0.06 | 24.9 |
| 10 | 0.09 | 50.6 | 0.06 | 26.9 |

*Samples 12 (inter-lab reproducibility above) and 14 (inter-lot reproducibility) were separate samples, but they were made with the same sera. Samples II and 13 did not have sufficient volume to be sent to investigators.

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b. Linearity/assay reportable range: not applicable
c. Traceability, Stability, Expected values (controls, calibrators, or methods): not applicable
d. Detection limit: not applicable
e. Analytical specificity

i. Cross-reactivity

**Cross-reactivity (n = 51)**

Focus assessed cross-reactivity with two groups of samples: a "HSV ELISA dual negative" group (n=37), and a "HSV ELISA mixed seroreactivity" group (n=14).

The HSV ELISA dual negative group (n=37) included samples that were sero-negative with both the HerpeSelect-1 ELISA IgG and HerpeSelect 2 ELISA IgG, and were sero-positive by at least one of a FDA cleared CMV ELISA IgG (n = 18), a home brew VZV ACIF (n=32), a FDA cleared EBV VCA IgG (n=31). The HerpeSelect 1 and 2 Plexus IgG was HSV-1 negative with all but one of the HSV ELISA dual negatives, and equivocal with one sample (the one sample was CMV+ VZV+ and EBV+). The HerpeSelect 1 and 2 Plexus IgG was HSV-2 negative with all but one of the HSV ELISA dual negatives, and equivocal with one sample (the one sample was CMV- VZV+ and EBV+).

The HSV ELISA mixed reactivity group (n=14) included samples that were sero-positive with either the HerpeSelect-1 ELISA IgG or HerpeSelect 2 ELISA IgG, and were sero-positive by at least one of a FDA cleared CMV ELISA IgG (HSV-1 neg n = 2, HSV-2 neg n = 9), a home brew VZV ACIF (HSV-1 neg n = 1, HSV-2 neg n = 2), a FDA cleared EBV VCA IgG (HSV-1 neg n = 1, HSV-2 neg n = 0). The HerpeSelect 1 and 2 Plexus IgG was HSV-1 negative with all of the HSV-1 ELISA negatives in the mixed reactivity group.

The HerpeSelect 1 and 2 Plexus IgG was HSV-2 negative with all of the HSV-2 ELISA negatives in the mixed reactivity group.

| Cross-reactant | HSV ELISAs | HerpeSelect-1 Plexus | | | | | HerpeSelect-2 Plexus | | | | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | | n | Neg | Eqv* | Pos | % POS | n | Neg | Eqv* | Pos | % POS |
| CMV IgG + | Dual Neg | 18 | 17 | 1 | 0 | 5.6% (1/18)
95%CI 0.1-27.3% | 18 | 18 | 0 | 0 | 0.0% (0/18) 95%CI
0.0-18.5% |
| | +/- or -/+ | 2 | 2 | 0 | 0 | 0.0% (0/2) 95%CI
0.0-84.2% | 9 | 9 | 0 | 0 | 0.0% (0/9) 95%CI
0.0-33.6% |
| | Total | 20 | 19 | 1 | 0 | 5.0% (1/20)
95%CI 0.1-24.9% | 27 | 27 | 0 | 0 | 0.0% (0/27) 95%CI
0.0-12.8% |

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iv. Interfering substances: not applicable

f. Assay cut-off

2. Comparison studies:

a. Method comparison with predicate device: Immunoblot IgG as the reference method, see clinical studies
b. Matrix comparison: not applicable

3. Clinical studies:

PERFORMANCE CHARACTERISTICS

Summary of Studies (see details below)

| Study | HerpeSelect 1 Plexus IgG Results | HerpeSelect 2 Plexus IgG Results |
| --- | --- | --- |
| Expectant Mothers (Indicated Population) | Agreement with positives | 96.5% |
| | Agreement with negatives | 92.2% |
| Sexually Active Adults (Indicated Population) | Agreement with positives | 91.0% |
| | Agreement with negatives | 96.5% |
| CDC HSV/CMV Panel | Agreement with positives | 100% |
| | Agreement with negatives | 100% |
| Low Prevalence Population | Agreement with negatives | 97.9% |
| Cross-reactivity with CMV, EBV and VZV. | Cross-reactivity | 0-5% |
| Reproducibility | %CV of positives | ≤10% |

Reactivity with Expectant Mothers (n = 300)

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Focus (n = 150) and an external investigator (n = 150) assessed the device's reactivity with sera from Expectant Mothers. The sera were sequentially submitted to the laboratory, archived, and masked. The external investigator was an University laboratory located in Northern California, and the sera were collected in the Pacific Northwestern United States. The HerpeSelect Plexus results were compared to the HerpeSelect 1 ELISA IgG and the HerpeSelect 2 ELISA IgG, using the Focus HerpeSelect 1 and 2 Immunoblot IgG as the reference method.

## HSV-1 Reactivity

The Focus HerpeSelect 1 Immunoblot IgG was: HSV-1 positive for 170 samples, HSV-1 negative with 128 samples, and HSV Common Antigen band positive for two samples. The Plexus HerpeSelect 1 agreed with: 96.5% (164/170) of Immunoblot positives, and 92.2% (118/128) of Immunoblot negatives. The two Immunoblot HSV Common Antigen band positives were both negative in the Plexus.

## HSV-2 Reactivity

The Focus HerpeSelect 2 Immunoblot IgG was: HSV-2 positive for 122 samples, HSV-2 negative with 176 samples, and HSV Common Antigen band positive for two samples. The Plexus HerpeSelect 2 agreed with: 94.3% (115/122) of Immunoblot positives, and 95.5% (168/176) of Immunoblot negatives. The two Immunoblot HSV Common Antigen band positives were both negative in the Plexus.

Plexus HerpeSelect 1 IgG Reactivity with Expectant Mothers (n = 300)

| Lab | Herpe - Select Immunoblot | Plexus HerpeSelect-1 | | | | | HerpeSelect-1 ELISA | | | | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | | n | Ne g | Eqv | Pos | % Agreement | n | Ne g | Eqv | Pos | % Agreement |
| Site 1 | Pos | 84 | 6 | 0 | 78 | 92.9% (78/84) 95%CI 85.1-97.3% | 84 | 5 | 1 | 78 | 92.9% (78/84) 95%CI 85.1-97.3% |
| Focus | Pos | 86 | 0 | 0 | 86 | 100% (86/86) 95%CI 95.8-100% | 86 | 0 | 0 | 86 | 100% (86/86) 95%CI 95.8-100% |
| Combined Labs | Pos | 170 | 6 | 0 | 164 | 96.5% (164/170) 95%CI 92.5-98.7% | 170 | 5 | 1 | 164 | 96.5% (164/170) 95%CI 92.5-98.7% |
| Site 1 | Neg | 66 | 61 | 1 | 4 | 92.4% (61/66) 95%CI 83.2-97.5% | 66 | 59 | 2 | 5 | 89.4% (59/66) 95%CI 79.4-95.6% |
| Focus | Neg | 62 | 57 | 2 | 3 | 91.9% (57/62) 95%CI 82.2-97.3% | 62 | 59 | 1 | 2 | 95.2% (59/62) 95%CI 86.5-99.0% |
| Combined Labs | Neg | 128 | 118 | 3 | 7 | 92.2% (118/128) 95%CI 86.1-96.2% | 128 | 118 | 3 | 7 | 92.2% (118/128) 95%CI 86.1-96.2% |
| Site 1 | Com | 0 | 0 | 0 | 0 | Na | 0 | 0 | 0 | 0 | Na |
| Focus | Com | 2 | 2 | 0 | 0 | Na | 2 | 2 | 0 | 0 | Na |

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| Combined Labs | Com | 2 | 2 | 0 | 0 | Na | 2 | 2 | 0 | 0 | Na |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |

Plexus HerpeSelect 2 IgG Reactivity with Expectant Mothers (n = 300)

| Lab | Herpe - Select Immunoblot | Plexus HerpeSelect-2 | | | | | HerpeSelect-2 ELISA | | | | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | | n | Ne g | Eqv | Pos | % Agreement | n | Ne g | Eqv | Pos | % Agreement |
| Site 1 | Pos | 60 | 3 | 1 | 56 | 93.3% (56/60)
95%CI 83.8-98.2% | 60 | 2 | 0 | 58 | 96.7% (58/60)
95%CI 88.5-99.6% |
| Focus | Pos | 62 | 2 | 1 | 59 | 95.2% (59/62)
95%CI 86.5-99.0% | 62 | 1 | 0 | 61 | 98.4% (61/62)
95%CI 91.3-100% |
| Combined Labs | Pos | 122 | 5 | 2 | 115 | 94.3% (115/122)
95%CI 88.5-97.7% | 122 | 3 | 0 | 119 | 97.5% (119/122)
95%CI 93.0-99.5% |
| Site 1 | Neg | 90 | 88 | 0 | 2 | 97.8% (88/90)
95%CI 92.2-99.7% | 90 | 86 | 0 | 4 | 95.6% (86/90)
95%CI 89.0-98.8% |
| Focus | Neg | 86 | 80 | 3 | 3 | 93.0% (80/86)
95%CI 85.4-97.4% | 86 | 80 | 1 | 5 | 93.0% (80/86)
95%CI 85.4-97.4% |
| Combined Labs | Neg | 176 | 168 | 3 | 5 | 95.5% (168/176)
95%CI 91.2-98.0% | 176 | 166 | 1 | 9 | 94.3% (166/176)
95%CI 89.8-97.2% |
| Site 1 | Com | 0 | 0 | 0 | 0 | na | 0 | 0 | 0 | 0 | na |
| Focus | Com | 2 | 2 | 0 | 0 | Na | 2 | 2 | 0 | 0 | Na |
| Combined Labs | Com | 2 | 2 | 0 | 0 | Na | 2 | 2 | 0 | 0 | Na |

## Reactivity with Sexually Active Adults (n = 300)

Focus (n = 150) and an external investigator (n = 150) assessed the device's reactivity with sera from sexually active adults. The sera were sequentially submitted to the laboratory, archived, and masked. The external investigator was a clinical laboratory located in Southern California, and the sera were collected in the Pacific Northwestern United States. The HerpeSelect Plexus results were compared to the HerpeSelect 1 ELISA IgG and the HerpeSelect 2 ELISA IgG, using the Focus HerpeSelect 1 and 2 Immunoblot IgG as the reference method.

## HSV-1 Reactivity

The Focus HerpeSelect 1 Immunoblot IgG was: HSV-1 positive for 157 samples, HSV-1 negative with 142 samples, and HSV Common Antigen band positive for one sample.

The Plexus HerpeSelect 1 agreed with: 91.0% (142/156) of Immunoblot positives (one sample was not run on the Plexus device), and

96.5% (137/142) of Immunoblot negatives.

## HSV-2 Reactivity

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The Focus HerpeSelect 2 Immunoblot IgG was: HSV-2 positive for 109 samples, HSV-2 negative with 190 samples, and HSV Common Antigen band positive for one sample. The Plexus HerpeSelect 2 agreed with: $96.3\%$ (105/109) of Immunoblot positives, and $97.4\%$ (184/189) of Immunoblot negatives (one sample was not run on the Plexus device).

Plexus HerpeSelect 1 IgG Reactivity with Sexually Active Adults (n = 300)

| Lab | Herpe - Select Immunoblo t | Plexus HerpeSelect-1 | | | | | HerpeSelect-1 ELISA | | | | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | | n | Ne g | Eqv | Pos | % Agreement | n | Ne g | Eqv | Pos | % Agreement |
| Site 2 | Pos | 71 | 4 | 2 | 65 | 91.5% (65/71) 95%CI 82.5-96.8% | 71 | 3 | 0 | 68 | 95.8% (68/71) 95%CI 88.1-99.1% |
| Focus | Pos | 85* | 5 | 3 | 77 | 90.6% (77/85) 95%CI 82.3-95.9% | 86 | 4 | 2 | 80 | 93.0% (80/86) 95%CI 85.4-97.4% |
| Combined Labs | Pos | 156 | 9 | 5 | 142 | 91.0% (142/156) 95%CI 85.4-95.0% | 157 | 7 | 2 | 147 | 93.6% (147/157) 95%CI 88.6-96.9% |
| Site 2 | Neg | 79 | 78 | 1 | 0 | 98.7% (78/79) 95%CI 93.1-100% | 79 | 77 | 1 | 1 | 97.5% (77/79) 95%CI 91.2-99.7% |
| Focus | Neg | 63 | 59 | 2 | 2 | 93.7% (59/63) 95%CI 84.5-98.2% | 63 | 60 | 0 | 3 | 95.2% (60/63) 95%CI 86.7-99.0% |
| Combined Labs | Neg | 142 | 137 | 3 | 2 | 96.5% (137/142) 95%CI 92.0-98.9% | 142 | 137 | 1 | 4 | 96.5% (137/142) 95%CI 92.0-98.9% |
| Site 2 | Com | 0 | 0 | 0 | 0 | na | 0 | 0 | 0 | 0 | na |
| Focus | Com | 1 | 1 | 0 | 0 | na | 1 | 1 | 0 | 0 | na |
| Combined Labs | Com | 1 | 1 | 0 | 0 | na | 1 | 1 | 0 | 0 | na |

*One of 300 samples was not run in the Plexus HerpeSelect, and that one sample was HSV-1 negative/HSV-2 positive in the Immunoblot.

Plexus HerpeSelect 2 IgG Reactivity with Sexually Active Adults (n = 300)

| Lab | Herpe - Select Immunoblo t | Plexus HerpeSelect-2 | | | | | HerpeSelect-2 ELISA | | | | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | | n | Ne g | Eqv | Pos | % Agreement | n | Ne g | Eqv | Pos | % Agreement |
| Site 2 | Pos | 47 | 3 | 1 | 43 | 91.5% (43/47) 95%CI 79.6-97.6% | 47 | 1 | 0 | 46 | 97.9% (46/47) 95%CI 88.7-99.9% |
| Focus | Pos | 62 | 0 | 0 | 62 | 100%(62/62) 95%CI 94.2-100% | 62 | 0 | 1 | 61 | 98.4%(61/62) 95%CI 91.3-100% |

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| Combined Labs | Pos | 109 | 3 | 1 | 105 | 96.3% (105/109) 95%CI 90.9-99.0% | 109 | 1 | 1 | 107 | 98.2% (107/109) 95%CI 93.5-99.8% |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| Site 2 | Neg | 103 | 100 | 1 | 2 | 97.1% (100/103) 95%CI 91.7-99.4% | 103 | 102 | 0 | 1 | 99.0% (102/103) 95%CI 94.7-100% |
| Focus | Neg | 86* | 84 | 0 | 2 | 97.7% (84/86) 95%CI 91.8-99.7% | 87 | 84 | 1 | 2 | 96.6% (84/87) 95%CI 90.3-99.3% |
| Combined Labs | Neg | 189 | 184 | 1 | 4 | 97.4% (184/189) 95%CI 93.9-99.1% | 190 | 186 | 1 | 3 | 97.9% (186/190) 95%CI 94.7-99.4% |
| Site 2 | Com | 0 | 0 | 0 | 0 | na | 0 | 0 | 0 | 0 | na |
| Focus | Com | 1 | 1 | 0 | 0 | na | 1 | 1 | 0 | 0 | na |
| Combined Labs | Com | 1 | 1 | 0 | 0 | na | 1 | 1 | 0 | 0 | na |

*One of 300 samples was not run in the Plexus HerpeSelect, and that one sample was HSV-1 negative/HSV-2 positive in the Immunoblot.

# Agreement with CDC Panel $(\mathbf{n} = 100)$

The following information is from a serum panel obtained from the CDC and tested by Focus Diagnostics. The results are presented as a means to convey further information on the performance of this assay with a masked, characterized serum panel. This does not imply an endorsement of the assay by the CDC.

The test panel consists of 100 samples. This panel contains duplicate samples of 50 test sera. The duplicates serve to test for reproducibility. There are 16 HSV-1 positive, 7 HSV-2 positive, 11 double-positive and 16 double-negative sera resulting in 54 HSV-1 positive and 36 HSV-2 positive specimens.

# Determination of positive and negative samples

Of the 54 HSV-1 positive samples, the HerpeSelect® Plexus IgG correctly identified $100\%$ (54/54).
Of the 36 HSV-2 positive samples, the HerpeSelect® Plexus IgG correctly identified $100\%$ (36/36).
Of the 22 double positive samples, the HerpeSelect® Plexus IgG correctly identified $100\%$ (22/22).
Of the 32 double negative samples, the HerpeSelect® Plexus IgG correctly identified $100\%$ (32/32).

Agreement with CDC Panel (n = 100)

| Sample Type | CDC Result | | n | HerpeSelect-1 Plexus Results | | | | HerpeSelect-2 Plexus Results | | | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | HSV1 | HSV2 | | Neg | Eqv | Pos | % Agreement | Neg | Eqv | Pos | % Agreement |
| HSV-1 Positive | Pos | Neg | 32 | 0 | 0 | 32 | 100% (32/32) 95%CI 89.1-100% | 32 | 0 | 0 | 100% (32/32) 95%CI 89.1-100% |
| HSV-2 Positive | Neg | Pos | 14 | 14 | 0 | 0 | 100% (14/14) 95%CI 76.8-100% | 0 | 0 | 14 | 100% (14/14) 95%CI 76.8-100% |
| Dual | Pos | Pos | 22 | 0 | 0 | 22 | 100% (22/22) | 0 | 0 | 22 | 100% (22/22) |

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| Positive | | | | | | | 95%CI 84.6-100% | | | | 95%CI 84.6-100% |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| Dual Negative | Neg | Neg | 32 | 32 | 0 | 0 | 100% (32/32)
95%CI 89.1-100% | 32 | 0 | 0 | 100% (32/32)
95%CI 89.1-100% |

## CDC Panel Reproducibility

All paired sera were correctly identified: The Focus Diagnostics HerpeSelect® 1 and 2 Plexus IgG identified 16 out of 16 paired HSV-1 positive and HSV-2 negative (100%), 7 out of 7 paired HSV-2 positive and HSV-1 negative (100%), 11 out of 11 paired double-positive (100%) and 16 out of 16 paired double-negative (100%) samples.

## Specificity with a Low Prevalence Population (n = 77)

Focus (n = 77) assessed the device's reactivity with sera from a low prevalence population. Focus selected sera from patients aged 18 and 19 years, and that had been submitted to a clinical laboratory in Southern California from states having a history of low sexually transmitted disease prevalence. Focus excluded sera that were submitted for sexually transmitted diseases, herpesvirus testing, and tests indicating the patient may be immunocompromised. The sera were sequentially selected, archived and masked. The HerpeSelect Plexus results were compared to the Focus HerpeSelect 1 and 2 Immunoblot IgG.

## HSV-1 Reactivity

The Focus HerpeSelect 1 Immunoblot IgG was: HSV-1 positive for 28 samples, HSV-1 negative with 47 samples, and HSV Common Antigen band positive for two samples.

The Plexus HerpeSelect 1 agreed with: 96.4% (27/28) of Immunoblot positives (one HSV-1 Immunoblot positive was equivocal in the Plexus device), and 97.9% (46/47) of Immunoblot negatives (one HSV-1 Immunoblot negative was positive in the Plexus device). One Immunoblot HSV Common Antigen band positive was equivocal in the Plexus, and the other sample was Plexus negative.

## HSV-2 Reactivity

The Focus HerpeSelect 2 Immunoblot IgG was: HSV-2 positive for four samples, HSV-2 negative with 71 samples, and HSV Common Antigen band positive for two samples. The Plexus HerpeSelect 2 agreed with: 75.0% (3/4) of Immunoblot positives (one HSV-1 Immunoblot positive was equivocal in the Plexus device), and 100% (71/71) of Immunoblot negatives (one HSV-1 Immunoblot negative was positive in the Plexus device). Both Immunoblot HSV Common Antigen band positives were negative in the Plexus.

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Specificity with Low Prevalence Population (n = 77)

| HerpeSelect Immunoblot | Plexus HerpeSelect-1 | | | | | Plexus HerpeSelect-2 | | | | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | n | Ne g | Eq v | Pos | % Agreement | n | Ne g | Eq v | Pos | % Agreement |
| Pos | 28 | 0 | 1 | 27 | 96.4% (27/28) 95%CI 81.6-99.9% | 4 | 0 | 1 | 3 | 75.0% (3/4) 95%CI 19.4-99.4% |
| Neg | 47 | 46 | 0 | 1 | 97.9% (46/47) 95%CI 88.7-99.9% | 71 | 71 | 0 | 0 | 100% (71/71) 95%CI 94.9-100% |
| Com | 2 | 1 | 1 | 0 | Na | 2 | 2 | 0 | 0 | na |

4. Clinical cut-off:

5. Expected values/Reference range:

EXPECTED VALUES

An outside investigator assessed the device with masked, archived and unselected sera from 1) sexually active adults (n = 300), and 2) from expectant mothers (n = 300). The reference methods were Focus Diagnostics HerpeSelect 1 ELISA IgG and HerpeSelect 2 ELISA IgG. The observed prevalences and the hypothetical predictive values for the two populations are shown below. The positive predictive value will decrease proportionally to the prevalence of HSV infection as reflected in the table below. The calculations are based on HerpeSelect 1 and 2 Plexus IgG having

1) HSV-1 sensitivity of 96.5% and a HSV-1 specificity of 92.2% (expectant mothers), and
2) HSV-2 sensitivity of 94.3% and a HSV-2 specificity of 95.5% (expectant mothers).
3) HSV-1 sensitivity of 91.0% and a HSV-1 specificity of 96.5% (Sexually Active Adults), and
4) HSV-2 sensitivity of 96.3% and a HSV-2 specificity of 97.4% (Sexually Active Adults).

Observed Prevalence with Sexually Active Adults

| Sero-status | Observed Prevalence | | |
| --- | --- | --- | --- |
| | HerpeSelect ELISA | HerpeSelect Immunoblot | HerpeSelect Plexus |
| HSV-1 neg | 48.3% (145/300) | 47.3% (142/300) | 49.2% (147/299) |
| HSV-1 + | 50.3% (151/300) | 52.0% (156/300) | 48.2% (144/299) |
| HSV-2 neg | 62.7% (188/300) | 63.0% (189/300) | 62.9% (188/299) |
| HSV-2 + | 36.7% (110/300) | 36.3% (109/300) | 36.4% 9109/299) |

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Observed Prevalence with Expectant Mothers

| Sero-status | Observed Prevalence | | |
| --- | --- | --- | --- |
| | HerpeSelect ELISA | HerpeSelect Immunoblot | HerpeSelect Plexus |
| HSV-1 neg | 41.7% (125/300) | 42.6% (128/300) | 42.0% (126/300) |
| HSV-1 + | 57.0% (171/300) | 56.7% (170/300) | 57.0% (171/300) |
| HSV-2 neg | 57.0% (171/300) | 58.7% (176/300) | 58.3% (175/300) |
| HSV-2 + | 42.7% (128/300) | 40.7% (122/300) | 40.0% (120/300) |

Expectant Mothers Prevalence vs. Hypothetical Predictive Values

| Prevalence | HSV-1 | | HSV-2 | |
| --- | --- | --- | --- | --- |
| | PPV | NPV | PPV | NPV |
| 50% | 92.5 | 92.2 | 95.4 | 95.5 |
| 40% | 89.2 | 94.7 | 93.3 | 97.0 |
| 30% | 84.1 | 96.5 | 90.0 | 98.0 |
| 25% | 80.5 | 97.3 | 87.5 | 98.5 |
| 20% | 75.6 | 97.9 | 84.0 | 98.8 |
| 15% | 68.6 | 98.5 | 78.7 | 99.2 |
| 10% | 57.9 | 99.1 | 70.0 | 99.5 |
| 5% | 39.4 | 99.6 | 52.4 | 99.8 |

Sexually Active Adults Prevalence vs. Hypothetical Predictive Values

| Prevalence | HSV-1 | | HSV-2 | |
| --- | --- | --- | --- | --- |
| | PPV | NPV | PPV | NPV |
| 50% | 96.3 | 96.5 | 97.4 | 97.4 |
| 40% | 94.5 | 97.6 | 96.1 | 98.3 |
| 30% | 91.8 | 98.5 | 94.1 | 98.9 |
| 25% | 89.7 | 98.8 | 92.5 | 99.1 |

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| 20% | 86.7 | 99.1 | 90.3 | 99.3 |
| --- | --- | --- | --- | --- |
| 15% | 82.1 | 99.4 | 86.7 | 99.5 |
| 10% | 74.3 | 99.6 | 80.5 | 99.7 |
| 5% | 57.8 | 99.8 | 66.1 | 99.9 |

Note: Sexually active adult and expectant mother populations in different geographic areas may produce different frequency distributions from the table above. Each laboratory should establish frequency distributions for their specific patient populations.

N. Proposed Labeling:

The labeling is sufficient and it satisfies the requirements of 21 CFR Part 809.10.

O. Conclusion:

The submitted information in this premarket notification is complete and supports a substantial equivalence decision.