(90 days)
The APTIMA Combo 2 Assay is a target amplification nucleic acid probe test that utilizes target capture for the in vitro qualitative detection and differentiation of ribosomal RNA (rRNA) from Chlamydia trachomatis (CT) and/or Neisseria gonorrhoeae (GC) to aid in the diagnosis of chlamydial and/or gonococcal urogenital disease using the PANTHER System as specified.
On the PANTHER System, the assay may be used to test the following specimens from symptomatic and asymptomatic individuals: clinician-collected endocervical, vaginal and male urethral swab specimens, clinician-collected gynecological specimens collected in the PreservCyt Solution, patient-collected vaginal swab specimens, and male urine specimens.
Patient-collected vaginal swab specimens are an option for screening women when a pelvic exam is not otherwise indicated. The vaginal swab specimen collection kit is not for home use.
The APTIMA Combo 2 Assay combines the technologies of target capture, transcriptionmediated amplification (TMA), and dual kinetic assay (DKA).
Specimens are collected and transferred into their respective specimen transport tubes. The transport solutions in these tubes release the rRNA targets and protect them from degradation during storage. When the APTIMA Combo 2 Assay is performed in the laboratory, the target rRNA molecules are isolated from specimens by use of capture oligomers via target capture that utilizes magnetic microparticles. The capture oligomers contain sequences complementary to specific regions of the target molecules as well as a string of deox yadenosine residues. A separate capture oligomer is used for each target. During the hybridization step, the sequence specific regions of the capture oligomers bind to specific regions of the target molecules. The capture oligomer:target complex is then captured out of solution by decreasing the temperature of the reaction to room temperature. This temperature reduction allows hybridization to occur between the deoxyadenosine region on the capture oligomer and the poly-deoxythymidine molecules that are covalently attached to the magnetic particles. The microparticles, including the captured target molecules bound to them, are pulled to the side of the reaction vessel using magnets and the supernatant is aspirated. The particles are washed to remove residual specimen matrix that may contain amplification reaction inhibitors. After the target capture steps are completed, the specimens are ready for amplification.
Target amplification assays are based on the ability of complementary oligonucleotide primers to specifically anneal and allow enzymatic amplification of the target nucleic acid strands. The APTIMA Combo 2 Assay replicates a specific region of the 23S rRNA from CT and a specific region of the 16S rRNA from GC via DNA intermediates. A unique set of primers is used for each target molecule. Detection of the rRNA amplification product sequences (amplicon) is achieved using nucleic acid hybridization. Single-stranded chemiluminescent DNA probes, which are complementary to a region of each target amplicon, are labeled with different acridinium ester molecules. The labeled DNA probes combine with amplicon to form stable RNA:DNA hybrids. The Selection Reagent differentiates hybridized from unhybridized probe, eliminating the generation of signal from unhybridized probe. During the detection step, light emitted from the labeled RNA:DNA hybrids is measured as photon signals in a luminometer, and are reported as Relative Light Units (RLU). In DKA, differences in the kinetic profiles of the CT and GC labeled probes allow for the differentiation of signal; kinetic profiles are derived from measurements of photon output during the detection read time. The chemiluminescent detection for CT signal has very rapid kinetics and has the "flasher" kinetic type. The chemiluminescent detection reaction for GC signal is relatively slower and has the "glower" kinetic type. Assay results are determined by a cut-off based on the total RLU and the kinetic curve type.
APTIMA Combo 2® Assay (on PANTHER® System) - Acceptance Criteria and Study Details
The APTIMA Combo 2® Assay is a nucleic acid amplification test for the qualitative detection and differentiation of ribosomal RNA from Chlamydia trachomatis (CT) and/or Neisseria gonorrhoeae (GC) to aid in the diagnosis of chlamydial and/or gonococcal urogenital disease. This 510(k) submission (K132251) specifically focuses on clearing the assay for use with male urine specimens on the PANTHER System.
1. Acceptance Criteria and Reported Device Performance
The provided document details the performance characteristics for male urine, alongside other specimen types, in terms of sensitivity and specificity based on two clinical studies. While explicit "acceptance criteria" in a numeric format (e.g., minimum sensitivity of X%) are not directly stated, the reported performance metrics demonstrate the device's efficacy across various specimen types and symptom statuses. The regulatory approval implies these performance levels met the FDA's requirements for substantial equivalence.
Reported Device Performance for Male Urine (from Table 4 for CT and Table 7 for GC):
| Specimen Type | Analyte | Prevalence (%) | Sensitivity % (95% CI) | Specificity % (95% CI) | PPV % (95% CI) | NPV % (95% CI) |
|---|---|---|---|---|---|---|
| Male Urine (MU) | CT | 11.5 | 95.2 (91.3-97.4) | 99.8 (99.4-99.9) | 98.5 (95.8-99.7) | 99.4 (98.9-99.7) |
| Male Urine (MU) | GC | 4.2 | 98.7 (92.9-99.8) | 99.7 (99.3-99.9) | 93.8 (86.7-97.8) | 99.9 (99.7-100) |
Additional detailed performance by symptom status is provided in Table 5 (CT) and Table 8 (GC) and by individual study in Table 6 (CT) and Table 9 (GC).
2. Sample Sizes and Data Provenance for Test Set
The clinical performance data was derived from two multi-center clinical studies conducted in the United States. The data is prospective, as specimens were collected from enrolled symptomatic and asymptomatic individuals for the purpose of the study.
Test Set Sample Sizes:
- Clinical Study 1: Included male urethral swab, vaginal swab, PreservCyt Solution liquid Pap, female endocervical swab, and male urine samples.
- Male subjects (urine): 580 enrolled. 580 male urine samples were tested.
- For CT performance analysis, 1799 male urine samples were included (data from Clinical Study 1 and 2 combined, see Table 4).
- For GC performance analysis, 1797 male urine samples were included (data from Clinical Study 1 and 2 combined, see Table 7).
- Clinical Study 2: Primarily focused on male urine specimens.
- Male subjects: 1492 enrolled, 1478 male urine samples from non-withdrawn subjects were tested.
- For CT performance analysis, 1799 male urine samples were included (data from Clinical Study 1 and 2 combined, see Table 4).
- For GC performance analysis, 1797 male urine samples were included (data from Clinical Study 1 and 2 combined, see Table 7).
A breakdown of male urine samples by study and symptom status for performance evaluation:
- CT (Table 6):
- Study 1 Asymptomatic: 323 samples
- Study 2 Asymptomatic: 979 samples
- Study 2 Symptomatic: 497 samples
- Total (Study 1 + Study 2) for male urine (Table 4): 1799 samples
- GC (Table 9):
- Study 1 Asymptomatic: 320 samples
- Study 2 Asymptomatic: 980 samples
- Study 2 Symptomatic: 497 samples
- Total (Study 1 + Study 2) for male urine (Table 7): 1797 samples
3. Number of Experts and Qualifications for Ground Truth
The document does not explicitly state the number of "experts" used to establish the ground truth or their specific qualifications (e.g., radiologist with X years of experience), as this is a diagnostic assay for infectious disease rather than image-based diagnosis.
Instead, the ground truth ("infected status") was established using cleared nucleic acid amplification tests (NAATs). The reference methods are described as:
- "cleared nucleic acid amplification tests (NAATs)" for Clinical Study 1 (male urethral swab, male and female urine, and PreservCyt Solution liquid Pap samples).
- "cleared NAATs" for Clinical Study 2 (male urethral swab and urine samples). Specifically, the infected status algorithm used "urethral swab and urine sample results from one reference CT and GC NAAT and urine sample results from two additional reference CT and GC NAATs to generate four reference results for each analyte."
The qualifications of the individuals performing these reference NAATs are not specified but would presumably be laboratory professionals trained in molecular diagnostics.
4. Adjudication Method for the Test Set
The adjudication method for establishing the "infected status" (ground truth) for the clinical test set was based on an algorithm using multiple reference NAATs.
- Clinical Study 1: "Subjects were categorized as infected if a positive result occurred in each of the two reference NAATs." (See Tables 10, 11, 13, and 14 for specific algorithms for different specimen types and analytes). For female subjects, if positive NAAT results occurred only in urine, they were considered infected for urine evaluation but non-infected for other non-urine specimens.
- Clinical Study 2 (Male Urine): "Subjects were categorized as infected if a positive result occurred in at least two of the reference NAATs." (See Tables 13 and 15).
This method is a form of "consensus" or "composite comparator" ground truth, where multiple established diagnostic tests are used to determine the true infection status in the absence of a single universally accepted gold standard.
5. Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study
No Multi-Reader Multi-Case (MRMC) comparative effectiveness study was mentioned or performed. This device is an in vitro diagnostic assay, not an imaging device requiring human reader interpretation or AI assistance for human readers. Therefore, the concept of "human readers improve with AI vs without AI assistance" is not applicable here. The assay provides a qualitative result directly.
6. Standalone (Algorithm Only) Performance
Yes, a standalone performance study was conducted. The performance data presented in the tables (Tables 4, 5, 6, 7, 8, 9) for sensitivity, specificity, PPV, and NPV represent the standalone performance of the APTIMA Combo 2 Assay on the PANTHER System, without human-in-the-loop interpretation being part of the result generation. The device itself performs the detection and differentiation of rRNA and provides a qualitative result.
7. Type of Ground Truth Used
The type of ground truth used was "composite comparator" or "reference NAAT consensus". For both clinical studies, the "infected status" was established by comparing results from two or more FDA-cleared nucleic acid amplification tests (NAATs) performed on the same or corresponding samples, rather than pathology (histology), clinical outcomes data, or a single expert's opinion.
8. Sample Size for the Training Set
The document does not explicitly state a separate "training set" sample size in the context of device development. For in vitro diagnostic devices, "training" often refers to internal analytical studies and optimization during development, rather than a distinct clinical "training set" in the way it's used for AI or machine learning models. The provided clinical studies (Clinical Study 1 and Clinical Study 2) represent the validation or test sets used to establish clinical performance.
Analytical sensitivity (Limit of Detection) studies were conducted using dilutions of CT organisms and GC organisms (7). These analytical studies involve controlled samples to determine the detection limits and would be part of the internal development and analytical verification processes, but are not typically referred to as a "training set" in this context.
9. How the Ground Truth for the Training Set Was Established
As noted above, a distinct "training set" with established ground truth in the clinical context is not explicitly described. For the analytical sensitivity studies, the "ground truth" (i.e., known concentration of organisms) was established by spiking known concentrations of CT and GC organisms into various matrices (e.g., Specimen Transport Medium, urine) and testing dilutions. This allows for the determination of the limit of detection (LOD) for the assay.
§ 866.3390
Neisseria spp. direct serological test reagents.(a)
Identification. Neisseria spp. direct serological test reagents are devices that consist of antigens and antisera used in serological tests to identifyNeisseria spp. from cultured isolates. Additionally, some of these reagents consist ofNeisseria spp. antisera conjugated with a fluorescent dye (immunofluorescent reagents) which may be used to detect the presence ofNeisseria spp. directly from clinical specimens. The identification aids in the diagnosis of disease caused by bacteria belonging to the genusNeisseria, such as epidemic cerebrospinal meningitis, meningococcal disease, and gonorrhea, and also provides epidemiological information on diseases caused by these microorganisms. The device does not include products for the detection of gonorrhea in humans by indirect methods, such as detection of antibodies or of oxidase produced by gonococcal organisms.(b)
Classification. Class II (performance standards).