ELECSYS TESTOSTERONE II IMMUNOASSAY

K093421 · Roche Diagnostics Corp. · CDZ · Apr 23, 2010 · Clinical Chemistry

Device Facts

• Record ID: K093421
• Device Name: ELECSYS TESTOSTERONE II IMMUNOASSAY
• Applicant: Roche Diagnostics Corp.
• Product Code: CDZ · Clinical Chemistry
• Decision Date: Apr 23, 2010
• Decision: SESE
• Submission Type: Traditional
• Regulation: 21 CFR 862.1680
• Device Class: Class 1
• Attributes: Pediatric

Indications for Use

Immunoassay for the in vitro quantitative determination of testosterone in human serum and plasma. The electrochemiluminescence immunoassay “ECLIA” is intended for use on Elecsys and cobas e immunoassay analyzers. Measurements of testosterone are used in the diagnosis and treatment of disorders involving the male sex hormones (androgens), including primary and secondary hypogonadism, delayed or precocious puberty, impotence in males and, in females hirsutism (excessive hair) and virilization (masculinization) due to tumors, polycystic ovaries, and androgenital syndromes.

Device Story

The Elecsys Testosterone II Immunoassay is an in vitro diagnostic test for measuring testosterone levels in human serum and plasma. It utilizes a competitive immunoassay principle with streptavidin-coated microparticles and electrochemiluminescence (ECLIA) detection. The assay is performed on automated Elecsys and cobas e immunoassay analyzers. The system uses a 2-point calibration and a master curve provided via reagent bar code to determine results. Healthcare providers use the quantitative output to assess androgen status in patients presenting with symptoms of hypogonadism, puberty disorders, or virilization. Results are intended to be interpreted in conjunction with clinical history and other diagnostic findings. The device benefits patients by providing standardized, accurate hormone quantification to guide clinical management of endocrine disorders.

Clinical Evidence

No clinical trials performed. Analytical performance validated via bench testing: precision (CLSI EP5-A2), linearity (CLSI EP6-A), LoD/LoQ (CLSI EP17-A), and interference studies. Method comparison study (n=387) against predicate showed Deming regression Y=0.989X-2.87 (R=0.992). Pediatric method comparison (n=123) showed Y=1.06X-2.79 (R=0.93). Reference ranges established for healthy adults and pediatric populations.

Technological Characteristics

Competitive immunoassay using streptavidin-coated microparticles and electrochemiluminescence (ECLIA). Reagents include R1, R2, and M (microparticles). Standardized to ID-GC/MS. Compatible with Elecsys and cobas e series analyzers. Measuring range 2.5–1500 ng/dL. Connectivity via automated analyzer platform.

Indications for Use

Indicated for the quantitative determination of testosterone in human serum and plasma to aid in the diagnosis and treatment of androgen-related disorders, including primary/secondary hypogonadism, delayed/precocious puberty, and male impotence; and in females, hirsutism and virilization associated with tumors, polycystic ovaries, and adrenogenital syndromes.

Regulatory Classification

Identification

A testosterone test system is a device intended to measure testosterone (a male sex hormone) in serum, plasma, and urine. Measurement of testosterone are used in the diagnosis and treatment of disorders involving the male sex hormones (androgens), including primary and secondary hypogonadism, delayed or precocious puberty, impotence in males and, in females hirsutism (excessive hair) and virilization (masculinization) due to tumors, polycystic ovaries, and adrenogenital syndromes.

Predicate Devices

• Elecsys Testosterone Assay (k964889)

Submission Summary (Full Text)

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

A. 510(k) Number:
k093421

B. Purpose for Submission:
New device

C. Measurand:
Testosterone

D. Type of Test:
Quantitative, electro-chemiluminescence (ECLIA) assay

E. Applicant:
Roche Diagnostics

F. Proprietary and Established Names:
Elecsys® Testosterone II Immunoassay

G. Regulatory Information:

| Product Code | Classification | Regulation Section | Panel |
| --- | --- | --- | --- |
| Radioimmunoassay, testosterone and dihydrotestosterone (CDZ) | Class I, reserved | 21 CFR 862.1680
Testosterone test system | Clinical Chemistry (75) |

H. Intended Use:

1. Intended use(s):
Refer to Indications for use below.

2. Indication(s) for use:
Immunoassay for the in vitro quantitative determination of testosterone in human serum and plasma. The electrochemiluminescence immunoassay “ECLIA” is intended for use on Elecsys and cobas e immunoassay analyzers.

Measurements of testosterone are used in the diagnosis and treatment of disorders involving the male sex hormones (androgens), including primary and secondary hypogonadism, delayed or precocious puberty, impotence in males and, in females hirsutism (excessive hair) and virilization (masculinization) due to tumors, polycystic ovaries, and androgenital syndromes.

3. Special conditions for use statement(s):
For Prescription use only

The sponsor has the following limitations in their labeling:

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"In patients receiving therapy with high biotin doses (i.e. > 5 mg/day), no sample should be taken until at least 8 hours after the last biotin administration."

"In isolated cases, elevated testosterone levels can be seen in samples from female patients with end stage renal disease (ESRD)."

"In rare cases, interference due to extremely high titers of antibodies to analyte-specific antibodies, streptavidin or ruthenium can occur. These effects have been minimized by suitable test design."

"Implausible elevated testosterone values in women should be verified by an extraction method or a validated LC-MS/MS tandem method."

4. Special instrument requirements:
Elecsys 2010 analyzer

I. Device Description:

The device is supplied as ready-to-use, three-reagent kit. The Elecsys Testosterone II reagent kit consists of a Reagent Pack (contains R1, R2, and M).

Reagent 1 contains Anti-testosterone-Ab-biotin, Biotinylated monoclonal anti-testosterone antibody, releasing reagent 2-bromoestradiol, MES buffer, and preservative. Reagent 2 contains Testosterone-peptide, Testosterone derivative, MES buffer, and preservative. M contains Streptavidin-coated microparticles.

All human source materials were tested by FDA approved methods and found to be negative for HIV ½, HBsAg, and HCV.

J. Substantial Equivalence Information:

1. Predicate device name(s):
Elecsys Testosterone Assay

2. Predicate 510(k) number(s):
k964889

3. Comparison with predicate:

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| Feature | Elecsys Testosterone II Assay (Candidate device) | Elecsys Testosterone Assay (K964889) Predicate Device |
| --- | --- | --- |
| Intended Use | Immunoassay for the in vitro quantitative determination of testosterone in human serum and plasma. The electrochemiluminescence immunoassay “ECLIA” is intended for use on Elecsys and cobas e immunoassay analyzers. | Immunoassay for the in vitro quantitative determination of testosterone in human serum and plasma. The electrochemiluminescence immunoassay “ECLIA” is intended for use on the Boehringer Mannheim Elecsys 1010 and 2010 immunoassay analyzers. |
| Indications for Use | Measurements of testosterone are used in the diagnosis and treatment of disorders involving the male sex hormones (androgens), including primary and secondary hypogonadism, delayed or precocious puberty, impotence in males and, in females hirsutism (excessive hair) and virilization (masculinization) due to tumors, polycystic ovaries, and adrenogenital syndromes. | Same |
| Assay Protocol | Competition principle | Same |
| Detection Protocol | Electrochemiluminescence immunoassay (ECLIA) | Same |
| Traceability/Standardization | ID-GC/MS (Isotope Dilution Gas Chromatography/Mass Spectrometry) | Same |
| Sample Type | Human serum and plasma | Same |
| Measuring Range | 2.5 – 1500 ng/dL (0.087 – 52.0 nmol/L) | 2.0 – 1500 ng/dL (0.069 – 52.0 nmol/L) |
| Calibrator | Testosterone II CalSet II (Calibrators 1 and 2) | Same |
| Calibration Interval | Once per reagent lot and after 1 month (28 days) when using the same reagent lot. After 7 days (when using the same reagent kit on the analyzer). As required: e.g. quality control findings outside the specified limits | Same |

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| Controls | PreciControl Universal 1 and 2 | Same |
| --- | --- | --- |
| Reagent Stability | Unopened at 2-8°C – up to the expiration date. After opening at 2-8°C – 12 weeks. Onboard the analyzer – 8 weeks | Unopened at 2-8°C – up to the expiration date. After opening at 2-8°C – 8 weeks. Onboard the analyzer – 8 weeks |

## K. Standard/Guidance Document Referenced (if applicable):

- CLSI EP5-A2: Evaluation of Precision Performance of Clinical Chemistry Devices; Approved Guideline Vol 19, No 2.
- CLSI EP6-A: Evaluation of the Linearity of Quantitative Analytical Measurement Procedure: A Statistical Approach; Approved Guideline, Vol 23 No 16.
- CLSI EP17-A: Protocol for Determination of Limits of Detection and Limits of Quantitation; Approved Guideline. Vol 24, No 34.

## L. Test Principle:

The Elecsys Testosterone II immunoassay is based on a competitive test principle with streptavidin-coated microparticles and electrochemiluminescence detection. Results are determined using a calibration curve that is generated specifically on each instrument by a 2-point calibration and a master curve provided with the reagent bar code.

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

1. Analytical performance:

a. Precision/Reproducibility:

Following CLSI EP5-A, the sponsor evaluated the precision using seven human serum (HS) specimens and two controls, at the concentrations listed in the table below. Precision was evaluated in duplicate, 2 runs per day for 21 days (n=84) using Elecsys 2010 analyzer. The results are tabulated below.

| Sample | Mean | Within run precision | | Between run precision | |
| --- | --- | --- | --- | --- | --- |
| | | SD | CV (%) | SD | CV (%) |
| | ng/dL | ng/dL | | ng/dL | |
| HS 1 | 4.5 | 0.5 | 10.2 | 0.8 | 18.5 |
| HS 2 | 9.5 | 0.4 | 4.7 | 0.8 | 8.4 |
| HS 3 | 69.1 | 1.4 | 2.1 | 2.2 | 3.2 |
| HS 4 | 216 | 4.2 | 1.9 | 6.0 | 2.8 |
| HS 5 | 867 | 22.9 | 2.6 | 24.3 | 2.8 |

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| HS 6 | 1300 | 15.8 | 1.2 | 44.0 | 3.4 |
| --- | --- | --- | --- | --- | --- |
| HS 7 | 1450 | 22.0 | 1.5 | 35.0 | 2.4 |
| PCU1 | 630 | 8.8 | 1.4 | 18.2 | 2.9 |
| PCU2 | 250 | 4.7 | 1.8 | 9.7 | 3.7 |

# b. Linearity/assay reportable range:

The sponsor performed linearity studies in accordance with the CLSI EP6-A guideline. Seventeen levels of samples were prepared by diluting a high spiked serum pool sample with a low serum pool sample. The linearity samples were tested in triplicate on the Elecsys 2010 analyzer over the range of 0.5 to $1535.8\mathrm{ng / dL}$ . The observed values were plotted against the expected values and an appropriate line fitted by standard linear regression resulting in: $\mathrm{y} = 1.0122\mathrm{x} - 0.0299$ ; $\mathrm{R} = 0.9996$ .

The data support the sponsor's claim that the measuring range of this device is 2.5 to $1500\mathrm{ng / dL}$ .

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

Calibrators used with this device have been previously cleared in k003411. Control materials have been previously cleared in k090541. The sponsor provided the stability study protocols for reagent stability. Based on the real-time stability studies, the reagent kit is stable up to 12 weeks. The on-board stability for open vials is 56 days.

# d. Limit of Detection:

The Limit of Blank (LoB), Limit of Detection (LoD), and Limit of Quantitation (LoQ) were determined in accordance with CLSI EP17-A document. LoB were conducted using a blank sample tested five times on 2 Elecsys 2010 analyzers over 3 days using 2 runs, and 2 reagent lots $(N = 60)$ . For LoD, five serum samples were selected and diluted to obtain lower concentrations. The samples were run over 3 days with 2 runs per day using 2 reagent lots $(N = 60)$ . For LoQ, 7 serum samples were tested once per run, 2 runs per day for over 5 days $(N = 70)$ . LoB was determined to be $1.2\mathrm{ng / dL}$ and LoD was determined to be $2.5\mathrm{ng / dL}$ . The LoQ was calculated as the lowest concentration for which the CV is less than $20\%$ . The results demonstrated that the LoQ is $12.0\mathrm{ng / dL}$

The claimed measuring range of the device is 2.5 to $1500\mathrm{ng / dL}$

# e. Analytical specificity:

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Interference studies were performed using four different levels of testosterone samples spiked with 5 endogenous substances, using the same protocol as follows: One aliquot of each serum sample was spiked with the interfering substance; another aliquot was spiked with the same volume of the solvent (control sample). The interfering pool was then diluted into the dilution pool in 10% increments, generating a total of 11 different concentrations of interference substances. Results of the spiked samples were compared with the control samples and % recovery was calculated. Non-significant interference was defined as recovery of ±10% (for concentration range of >100 ng/dL), ±15% (for concentration range of >50 to 100 ng/dL), and ±7.5 ng/dL (for concentration range of 15 to 50 ng/dL). Based on the data, the sponsor claims no significant interference for the substances and concentrations listed below:

Icterus (bilirubin up to 30 mg/dL),

Hemolysis (Hemoglobin up to 600 mg/dL),

Lipemia (Intralipid up to 1000 mg/dL),

Biotin: up to 30 ng/mL,

Rheumatoid Factor: up to 1000 IU/mL.

In addition, common pharmaceutical compounds were spiked into native human serum samples and tested with Testosterone II assay. Two serum samples pools containing approximately 60 ng/dL and 500 ng/dL testosterone were spiked with potential interferents (multiples of maximal daily doses). The reference sample (control) without interferent was spiked with the respective amount of solvent. Based on the sponsor's definition of non-significant interference (greater than ±10% of control value), the sponsor claims no interference for the compounds and concentrations listed in the table below:

| Compound | Concentration |
| --- | --- |
| Acetylcystein | 150 mg/L |
| Ampicillin | 1000 mg/L |
| Ascorbic acid | 300 mg/L |
| Ca-Dobesilate | 200 mg/L |
| Cyclosporine | 5 mg/L |
| Cefoxitin | 2500 mg/L |
| Heparin | 5000 U |
| Levodopa | 20 mg/L |
| Methyldopa | 20 mg/L |
| Metronidazole | 200 mg/L |
| Phenylbutazone | 400 mg/L |
| Doxycyclin | 50 mg/L |
| Acetylsalicylic Acid | 1000 mg/L |

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| Rifampicin | 60 mg/L |
| --- | --- |
| Acetaminophen | 200 mg/L |
| Ibuprofen | 50 mg/L |
| Theophylline | 100 mg/L |
| Heparin Clexane | 5000 U |
| Dexamethasone | 20 mg/L |

Nandrolone was tested and was considered as interference substance; therefore, the sponsor has the following limitations in the labeling:

"Do not use samples from patients under Nandrolone treatment"

Cross-reactivity study was performed using native human serum samples spiked with potential cross-reactant compounds. The spiked and non-spiked samples were tested in duplicates on the Elecsys 2010 analyzer. Results are summarized below:

| Substances | Concentration tested (ng/mL) | Cross-reactivity (%) |
| --- | --- | --- |
| Androstendione | 100 | ≤ 2.5 |
| Cortisol | 1000 | ≤ 0.01 |
| Cortisone | 2000 | ≤ 0.001 |
| Danazol | 1000 | ≤ 0.5 |
| Dexamethasone | 2000 | ≤ 0.001 |
| DHEA | 1000 | ≤ 0.016 |
| DHEA-S | 50000 | ≤ 0.003 |
| D-5-Androstene-3β,17β-diol | 1000 | ≤ 0.29 |
| Estradiol | 1000 | ≤ 0.16 |
| Estrone | 1000 | ≤ 0.004 |
| Ethisterone | 1000 | ≤ 2.40 |
| Norgestrel | 1000 | ≤ 0.91 |
| Testosterone propionate | 100 | ≤ 2.46 |
| 5-α-Androstane-3β,17β-diol | 1000 | ≤ 2.11 |
| 5-α-Dihydro-testosterone | 500 | ≤ 0.86 |
| 11-β-Hydroxy-testosterone | 100 | ≤ 18.0 |
| 11-Keto-testosterone | 1000 | ≤ 3.22 |
| Prednisone | 1000 | ≤ 0.001 |
| Prednisolone | 1000 | ≤ 0.002 |
| Progesterone | 1000 | ≤ 0.001 |

f. Assay cut-off: Not Applicable

2. Comparison studies:

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a. Method comparison with predicate device:

Method comparisons between the candidate device and the predicate device were performed according to the CLSI EP9-A guideline. A total of 387 serum samples (239 males and 148 females) ranging from $2.5 - 1400\mathrm{ng / dL}$ were used in the study. The candidate device results on the Elecsys 2010 (Y) were compared to the corresponding predicate results on the Elecsys 2010 (X). Deming regression analysis resulted in: $\mathrm{Y} = 0.989\mathrm{X} - 2.87$ . $\mathrm{R} = 0.992$

An additional method comparison study using 123 pediatric samples (69 males age 7-18 years old and 54 females' age 8-18 years old) was performed. Samples ranging from 2.74 to $1048.2\mathrm{ng / dL}$ . Deming regression analysis resulted in: $\mathrm{Y} = 1.06\mathrm{X} - 2.79$ ; $\mathrm{R} = 0.93$ .

b. Matrix comparison:

The sponsor performed a matrix comparison study using 45 unaltered paired serum, K2-EDTA plasma, K3-EDTA plasma and Lithium Heparin plasma samples. Samples ranging from 10 to $1460\mathrm{ng / dL}$ were analyzed on the Elecsys 2010 analyzer. The plasma results (Y) were compared to the corresponding serum results (X) and the Passing/Bablok regression analysis results are summarized in the table below:

| Anticoagulant | Regression Analysis |
| --- | --- |
| Lithium Heparin | Y=1.0008 - 0.0001; R = 0.997 |
| K2-EDTA | Y=0.998 + 0.0005; R = 0.998 |
| K3-EDTA | Y=0.993 + 0.0022; R = 0.998 |

The sponsor claimed that K2-EDTA, K3-EDTA and Lithium heparin are acceptable anticoagulants.

3. Clinical studies:

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

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# 5. Expected values/Reference range:

A reference range study was performed using healthy adult (214) males and (160) females without intake of contraceptives and prescription drugs. Results are summarized below:

| | | Percentiles | | |
| --- | --- | --- | --- | --- |
| Test subjects | N | Median | 5-95th | 2.5-97.5th |
| | | ng/dL | | |
| Males | 136 | 536 | 249-836 | 218-906 |
| 20-49 years | | | | |
| Males | 78 | 476 | 193-740 | 132-892 |
| ≥ 50 years | | | | |
| Females | 89 | 27.1 | 8.4-48.1 | 5.0-52.2 |
| 20-49 years | | | | |
| Females | 71 | 16.2 | 2.9-40.8 | < 2.5-46.1 |
| ≥ 50 years | | | | |

In addition, a reference range study using pediatric population (95 males and 100 females) under 18 years old, who were in good endocrinological health was performed. Subjects were clinically characterized according to their age as well as Tanner Stage. Tanner Stage was characterized according to the method of Marshall and Tanner. $^{(a,b)}$ Results are summarized below:

Reference values for males under the age of 18 characterized by age

| Age | N | Range, ng/dL | |
| --- | --- | --- | --- |
| | | Min | Max |
| 7 | 3 | < 2.5 | < 2.5 |
| 8 | 6 | < 2.5 | < 2.5 |
| 9 | 8 | < 2.5 | < 2.5 |
| 10 | 2 | < 2.5 | < 2.5 |
| 11 | 13 | < 2.5 | 236.6 |
| 12 | 6 | 29.40 | 278.4 |
| 13 | 8 | < 2.5 | 432 |
| 14 | 9 | 40.1 | 778.4 |
| 15 | 23 | 78.7 | 762.6 |
| 16 | 14 | 237.5 | 1048.2 |
| 17 | 1 | 505.5 | 505.5 |
| 18 | 2 | 557.1 | 685.1 |

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Reference values for females under the age of 18 characterized by age

| Age | N | Range, ng/dL | |
| --- | --- | --- | --- |
| | | Min | Max |
| 7 | 0 | — | — |
| 8 | 11 | < 2.5 | 6.14 |
| 9 | 16 | < 2.5 | 7.49 |
| 10 | 7 | < 2.5 | 5.45 |
| 11 | 13 | < 2.5 | 17.06 |
| 12 | 10 | < 2.5 | 26.23 |
| 13 | 7 | < 2.5 | 23.66 |
| 14 | 4 | 11.21 | 28.82 |
| 15 | 21 | 7.64 | 39.77 |
| 16 | 8 | < 2.5 | 29.4 |
| 17 | 0 | — | — |
| 18 | 3 | 15.33 | 31.13 |

Reference values for males under the age of 18

| | | | Percentiles | |
| --- | --- | --- | --- | --- |
| Tanner Stage | N | Median | 5-95th | 2.5-97.5th |
| | | | ng/dL | |
| 1 | 26 | < 2.5 | < 2.5 | < 2.5-4.2 |
| 2 | 18 | 59.7 | < 2.5-432 | < 2.5-432 |
| 3 | 15 | 245 | 64.9-778 | 64.9-778 |
| 4 | 16 | 344 | 180-763 | 180-763 |
| 5 | 20 | 446 | 188-882 | 138-1050 |

Reference values for females under the age of 18

| | | | Percentiles | |
| --- | --- | --- | --- | --- |
| Tanner Stage | N | Median | 5-95th | 2.5-97.5th |
| | | | ng/dL | |
| 1 | 37 | < 2.5 | < 2.5-6.12 | < 2.5-7.5 |
| 2 | 12 | < 2.5 | < 2.5-10.4 | < 2.5-10.4 |
| 3 | 12 | 7.9 | < 2.5-23.7 | < 2.5-23.7 |
| 4 | 12 | 12.2 | < 2.5-26.8 | < 2.5-26.8 |
| 5 | 27 | 19.7 | 4.6-38.3 | 4.5-39.8 |

Sponsor state that each laboratory should investigate the transferability of the expected values to its own patient population and if necessary determine its own reference ranges.

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References:
a. Marshall, W.A., Tanner, J.M. Variations in the pattern of pubertal changes in boys. Arch. Dis. Childh. 1970;45: 13-23.
b. Marshall, W.A., Tanner, J.M. Variations in the pattern of pubertal changes in girls. Arch. Dis. Childh. 1969;44: 291-303.

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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