S. AUREUS PNA FISH

K060099 · Advandx, Inc. · NXX · May 8, 2006 · Microbiology

Device Facts

• Record ID: K060099
• Device Name: S. AUREUS PNA FISH
• Applicant: Advandx, Inc.
• Product Code: NXX · Microbiology
• Decision Date: May 8, 2006
• Decision: SESE
• Submission Type: Traditional
• Regulation: 21 CFR 866.3700
• Device Class: Class 1

Indications for Use

S. aureus PNA FISH is a qualitative nucleic acid hybridization assay intended for presumptive identification of Staphylococcus aureus from blood cultures.

Device Story

S. aureus PNA FISH is a qualitative nucleic acid hybridization assay; utilizes peptide nucleic acid (PNA) probes to target specific ribosomal RNA sequences in Staphylococcus aureus. Input: positive blood culture samples. Process: hybridization of fluorescently labeled PNA probes to target RNA within fixed cells; visualization via fluorescence microscopy. Output: visual identification of S. aureus based on fluorescence pattern. Used in clinical microbiology laboratories by trained laboratory personnel. Provides rapid identification of S. aureus from blood cultures, enabling faster targeted antimicrobial therapy and improved patient management.

Clinical Evidence

Bench testing and multi-site clinical laboratory evaluations. Compared against standard culture identification procedures across 1,438 samples using BacT/Alert, ESP, and BACTEC systems. Overall sensitivity 99.2% (513/517) and specificity 99.5% (916/921). Reproducibility >95%. Analytical specificity 100% for S. aureus strains; potential for false positives with S. schleiferi due to single base mismatch.

Technological Characteristics

Nucleic acid hybridization assay using peptide nucleic acid (PNA) probes. Employs fluorescence in situ hybridization (FISH) technology. Requires fluorescence microscopy for visualization. Qualitative detection method.

Indications for Use

Indicated for presumptive identification of Staphylococcus aureus from positive blood cultures. For prescription use only.

Regulatory Classification

Identification

Staphylococcus aureus serological reagents are devices that consist of antigens and antisera used in serological tests to identify enterotoxin (toxin affecting the intestine) producing staphylococci from cultured isolates. The identification aids in the diagnosis of disease caused by this bacterium belonging to the genus Staphylococcus and provides epidemiological information on these diseases. Certain strains of Staphylococcus aureus produce an enterotoxin while growing in meat, dairy, or bakery products. After ingestion, this enterotoxin is absorbed in the gut and causes destruction of the intestinal lining (gastroenteritis).

Submission Summary (Full Text)

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

A. 510(k) Number:
k060099

B. Purpose for Submission:
New product *Staphylococcus aureus* PNA FISH™

C. Measurand:
*S. aureus* specific 16S ribosomal RNA

D. Type of Test:
Fluorescent In Situ Hybridization (FISH) using protein nucleic acid (PNA) probes

E. Applicant:
AdvanDx, Inc.

F. Proprietary and Established Names:
*S. aureus* PNA FISH™, *Staphylococcus aureus* Culture Identification Kit

G. Regulatory Information:
1. Regulation section:
21 CFR Part 866.3700 – Staphylococcus aureus serological reagents
2. Classification:
Class I
3. Product code:
NXX
4. Panel:
83 Microbiology

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H. Intended Use:

1. Intended use(s):

S. aureus PNA FISH is a qualitative nucleic acid hybridization assay intended for presumptive identification of Staphylococcus aureus from blood cultures.

2. Indication(s) for use:

This submission is for the new product S. aureus PNA FISH, which is a qualitative nucleic acid hybridization assay, intended for the presumptive identification of Staphylococcus aureus from blood cultures containing Gram-positive cocci in clusters (GPCC).

3. Special conditions for use statement(s):

Prescription use only.

4. Special instrument requirements:

AdvanDx Teflon-coated Microscope Slides. A fluorescent microscope equipped with an AdvanDx Dual Band Filter.

I. Device Description

The S. aureus PNA FISH Culture Identification Kit contains a 3 mL bottle of fixation solution, a 1.5 mL bottle of fluorescein-labeled PNA probe in hybridization solution, a 50 mL bottle of concentrated wash solution, which must be diluted prior to use, and a 3 mL bottle of mounting medium. The one-well, Teflon-coated microscope slides, glass coverslips and the external quality control organism slides are sold separately. User-prepared quality control organism slides are also acceptable. After processing, the slides must be examined within two hours by using a fluorescent microscope equipped with a dual band filter.

J. Substantial Equivalence Information:

1. Predicate device name(s):

a. AccuProbe S. aureus Culture Identification Test

2. Predicate K number(s):

a. K902213

3. Comparison with predicate(s):

| Similarities | | |
| --- | --- | --- |
| Item | Device | Predicate |
| Intended Use | Intended for the in vitro identification of S. aureus | Same |
| Organism | Blood cultures with GPCC | Same |

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| Differences | | |
| --- | --- | --- |
| Item | Device | Predicate |
| | PNA FISH | AccuProbe |
| Technology Method | Fluorescence in situ hybridization | Hybridization protection assay |
| Time to result | 2.5 hours from time of smear preparation | 18 – 24 hours following positive blood culture detection (e.g. 0 – 5 days) |
| Sample | Smear of blood cultures | Colonies grown on solid media culture, isolated from liquid blood culture media |
| Control organisms | S. aureus and S. epidermidis | S. aureus and S. epidermidis |
| Mechanism of identification | S. aureus specific 16S rRNA | S. aureus specific 16S rRNA |
| Interpretation of results | Qualitative fluorescent microscopy | Quantitative luminometer |

**K. Standard/Guidance Document Referenced (if applicable):**
Not applicable

**L. Test Principle:**
One drop of fluorescein-labeled, *S. aureus*-specific PNA probe is added to a methanol, heat, or flame fixed smear, prepared from liquid blood culture media with GPCC. Hybridization is performed during a 90 +/- 5 minute incubation at 55 +/- 1° C, in an incubator or on a slide warmer. The slide is examined by fluorescent microscopy within two hours of staining. *S. aureus* is presumptively identified as multiple bright green fluorescent clusters of cocci in multiple fields on a reddish background, whereas non-*S. aureus* cells will not fluoresce.

**M. Performance Characteristics (if/when applicable):**

**1. Analytical performance:**
The performance data were generated by multiple, geographically distinct, clinical laboratories. Each laboratory compared the device results to corresponding results obtained by standard culture identification procedures. These laboratories reported the sensitivity and specificity results in peer reviewed literature articles, and during presentations performed at professional meetings (posters).

**a. Precision/Reproducibility:**
Inter-laboratory and Intra-laboratory testing demonstrated >95% reproducibility. The ten isolate study described in the guidance document was used. (10 organisms tested 3 times on 3 days at 3 sites.)

**b. Linearity/assay reportable range:**
Not applicable

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c. Traceability, Stability, Expected values (controls, calibrators, or methods):

The recommended QC isolates, S. aureus ATCC #29213 and S. epidermidis ATCC #14990, provided by the manufacturer as External Control Slides, or as user-prepared slides from stock or liquid blood cultures, are recommended for testing in parallel for each batch of tests performed. At one testing (Spain) site the S. aureus PNA FISH user-prepared organism control slides were tested a sufficient number of times to demonstrate that the device can produce acceptable quality control results >95% of the time (285/285).

The stability of the S. aureus PNA FISH reagents were evaluated using real-time data. The sponsor periodically removed samples from five lots of S. aureus PNA FISH reagent from storage for testing. The analytical fluorescence performance and a functional performance of each lot were tested 5 times over 30 months after the manufacturing date. The fluorescent dye on the probe is light sensitive and is considered the least stable component of the kit. There was no change in performance for at least 24 months when stored at 2 - 8°C and protected from light, as indicated in the product labeling.

d. Detection limit:

The claimed detection limit for S. aureus in blood cultures was determined to be approximately 1 × 10⁵ colony forming units (CFU) per mL by serial dilutions of a S. aureus positive culture. This is consistent with the analytical sensitivity of slide-based staining techniques and is not limited by the test itself, but rather by the general requirement for 1 × 10⁵ CFU/mL for interpretation by standard light microscopy.

e. Analytical specificity:

Specificity of S. aureus PNA FISH probe was evaluated using cultures of 17 reference strains and 48 clinical isolates, methicillin-resistant and methicillin-susceptible S. aureus, coagulase-negative Staphylococcus species, and other clinically relevant and phylogenetically related bacterium, and yeast species. The analytical specificity for S. aureus strains positive by PNA FISH was 100% (17/17), and 96% (46/48) of the other strains were negative. Of these two false-positive strains, one was S. schleiferi and the other one was one of five Stomatococcus strains.

The target sequence is unique for S. aureus. False positive results with S. schleiferi may occur due to a single base mismatch. All other gram positive bacteria will likely have two or more mismatches and not be reactive. Other non-S. aureus species may potentially cause false-positive results.

f. Assay cut-off:

Not applicable

2. Comparison studies:

a. Method comparison of device to AccuProbe, as the reference method:

The S. aureus PNA FISH™, Staphylococcus aureus Culture Identification Kit was compared to the AccuProbe S. aureus Culture Identification Test (K902213) hybridization protection method. PNA tests were performed on 157 isolates on

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GPCC isolated from the ESP Blood Culture System. The results are presented in the table below. The data demonstrates substantial equivalence to the predicate for the presumptive identification of S. aureus isolated from blood cultures with GPCC.

| N=157 | Sensitivity | Specificity | Blood Culture System Evaluated |
| --- | --- | --- | --- |
| S. aureus PNA FISH | 98.6% (68/69) | 100.0% (88/88) | ESP |

AccuProbe = AccuProbe S. aureus Culture Identification Test (Gen-Probe, San Diego, CA)

ESP = ESP Blood culture system (Trek Diagnostics, Cleveland, OH)

The performance of S. aureus PNA FISH was evaluated using three different automated blood culture media systems at multiple sites. The data demonstrated that the S. aureus PNA FISH™ can be used with major blood culture systems, and results from testing are similar to results obtained by conventional methods. The results are displayed in the table below.

Automated blood culture system media evaluation comparing device results to results obtained by subculture and subsequent identification by standard methods, by site

| Blood Culture System Evaluated | S. aureus Prevalence Rate | Sensitivity | Specificity | Study |
| --- | --- | --- | --- | --- |
| BacT/Alert | 42.5% (37/87) | 97.3% (36/37) | 100% (50/50) | Site A |
| ESP | 32.8% (57/174) | 100% (57/57) | 99.2% (115/117) | Site B |
| BACTEC | 34.2% (68/199) | 98.5% (67/68) | 98.5% (129/131) | Site B |
| BacT/Alert | 39.0% (74/190) | 100% (74/74) | 99.1% (115/116) | Site B |
| BacT/Alert | 44.9% (128/285) | 100% (128/128) | 99.4 (156/157) | Site C |
| BacT/Alert | 30.4% (153/503) | 98.7% (151/153) | 100% (350/350) | Site D |
| Total | 36.0% (517/1438) | 99.2% (513/517) | 99.5% (916/921) | |

BacT/Alert = BacT/Alert Blood culture system (bioMérieux, Durham, NC)

BACTEC = BACTEC Blood culture system (Becton Dickenson, Sparks, MD)

ESP = ESP Blood culture system (Trek Diagnostics, Cleveland, OH)

b. Matrix comparison:

Not applicable

3. Clinical studies:

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a. Clinical Sensitivity:
Not applicable

b. Clinical specificity:
Not applicable

c. Other clinical supportive data (when a. and b. are not applicable):
Not applicable

4. Clinical cut-off:
Not applicable

5. Expected values/Reference range:
The expected S. aureus positive result rate from GPCC positive blood culture bottles is 30% - 43%, depending on institutional and patient population.

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.

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