HUMAN BETA-2 MICROGLOBULIN KIT

K083289 · The Binding Site, Ltd. · JZG · May 6, 2009 · Immunology

Device Facts

• Record ID: K083289
• Device Name: HUMAN BETA-2 MICROGLOBULIN KIT
• Applicant: The Binding Site, Ltd.
• Product Code: JZG · Immunology
• Decision Date: May 6, 2009
• Decision: SESE
• Submission Type: Traditional
• Regulation: 21 CFR 866.5630
• Device Class: Class 2

Intended Use

The kit is intended for the quantitative in vitro determination of beta-2 microglobulin (β2M) in human serum and urine, using SPAPLUS™ analyser to aid the diagnosis of active rheumatoid arthritis and kidney disease. The test result is to be used in conjunction with other clinical and laboratory findings.

Device Story

The device is a quantitative, latex-enhanced turbidimetric immunoassay for measuring beta-2 microglobulin (β2M) in human serum and urine. It utilizes monospecific sheep anti-β2M antibodies coated onto polystyrene latex particles. When mixed with patient samples, the antibodies form insoluble immune complexes with β2M; the latex particles enhance light scattering, allowing turbidimetric detection. The SPAPLUS™ analyser measures transmitted light, which is inversely proportional to the protein concentration. The system automatically calculates concentrations using a stored calibration curve. It is intended for use in clinical laboratory settings by trained personnel. Results assist clinicians in diagnosing active rheumatoid arthritis and kidney disease when interpreted alongside other clinical and laboratory findings. The device includes antigen excess protection to prevent false-negative results at high concentrations.

Clinical Evidence

No clinical studies were performed. Evidence consists of analytical bench testing, including precision (intra-assay CV 0.6-2.0%, inter-assay CV 1.9-5.8%), linearity (r² > 0.999), and interference studies. A method comparison study was conducted on 103 serum samples and 49 urine samples comparing the SPAPLUS™ assay to the predicate BNII system, yielding high correlation (Serum r=0.996; Urine r=0.993).

Technological Characteristics

Latex-enhanced turbidimetric immunoassay. Reagents: monospecific sheep anti-β2M antibodies on polystyrene latex, reaction buffer, calibrators, and controls. Preservatives: Proclin™, sodium azide, EACA, benzamidine. Detection: photodiode-based optical lens system measuring light transmission. Connectivity: integrated with SPAPLUS™ analyser. Standard: CLSI EP-5A for precision.

Indications for Use

Indicated for quantitative in vitro determination of beta-2 microglobulin in human serum and urine to aid diagnosis of active rheumatoid arthritis and kidney disease in adult populations.

Regulatory Classification

Identification

A beta-2-microglobulin immunological test system is a device that consists of the reagents used to measure by immunochemical techniques beta-2-microglobulin (a protein molecule) in serum, urine, and other body fluids. Measurement of beta-2-microglobulin aids in the diagnosis of active rheumatoid arthritis and kidney disease.

Special Controls

*Classification.* Class II (special controls). The device is exempt from the premarket notification procedures in subpart E of part 807 of this chapter subject to the limitations in § 866.9.

Predicate Devices

• The Binding Site β2M on the BNII (k946069)

Submission Summary (Full Text)

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# 510(k) SUBSTANTIAL EQUIVALENCE DETERMINATION DECISION SUMMARY

A. 510(k) Number:
k083289

B. Purpose for Submission:
New device

C. Measurand:
Beta-2 Microglobulin

D. Type of Test:
Quantitative, latex-enhanced turbidimetric assay

E. Applicant:
The Binding Site, Ltd.

F. Proprietary and Established Names:
Human Beta-2 Microglobulin Kit for use on $\mathrm{SPAPLUS}^{\mathrm{TM}}$

G. Regulatory Information:
1. Regulation section:
21 CFR § 866.5630 Beta-2 microglobulin immunological test system
2. Classification:
Class II
3. Product codes:
JZG, Beta-2 Microglobulin, Immunological Test System
4. Panel:
Immunology 82

H. Intended Use:
1. Intended use(s):
The kit is intended for the quantitative *in vitro* determination of beta-2 microglobulin (β2M) in human serum and urine, using $\mathrm{SPAPLUS}^{\mathrm{TM}}$ analyser to aid the diagnosis of active rheumatoid arthritis and kidney disease. The test result is to be used in conjunction with other clinical and laboratory findings.
2. Indication(s) for use:
Same as intended use.
3. Special conditions for use statement(s):
For prescription use only.
4. Special instrument requirements:
$\mathrm{SPAPLUS}^{\mathrm{TM}}$ analyser

I. Device Description:
The device consists of the following: monospecific sheep anti-β2M antibody coated onto polystyrene latex in the presence of preservatives. Calibrators 1-6; low, high and elevated controls in liquid form; and β2M reaction buffer. The reagents contain 0.05% Proclin™, 0.099% sodium azide, 0.1% E-amino-n-caproic acid (EACA) and 0.01% benzamidine as preservative.

J. Substantial Equivalence Information:
1. Predicate device name(s):
The Binding Site β2M on the BNII
2. Predicate K number(s):

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k946069

3. Comparison with predicate:

| Similarities | | |
| --- | --- | --- |
| Item | Device | Predicate |
| Intended Use | Quantitative determination of β2M in serum or urine to aid in the diagnosis of active rheumatoid arthritis and kidney disease | Same |
| Detection Method | Turbidimetric immunoassay | Same |
| Traceability | Standardized against the 1stInternational Standard for Beta-2 Microglobulin (NIBSC) | Same |
| Sample Matrix | Human serum and urine | Same |
| Antibodies | Monospecific Sheep | Same |
| Differences | | |
| --- | --- | --- |
| Item | Device | Predicate |
| Controls | Low, High and Elevated levels liquid ready to use | Low, High and Elevated levels lyophilized |
| Instruments | SPAPLUS™ analyser | BNII analyser |
| Measuring range | Serum: 0.3 - 40 mg/L Urine: 0.03 - 20 mg/L | Serum: 0.7 – 22.5 mg/L Urine: 0.04 mg/L |
| Reference Range | Serum: 0.80 – 2.34 mg/L Urine: 0.030 – 0.202 mg/L | Serum: 1.22 – 2.46 mg/L Urine: <0.03– 0.23 mg/L |

K. Standard/Guidance Document Referenced (if applicable):
CLSI (NCCLS) EP-5A: Evaluation of Precision Performance of Clinical Chemistry.

L. Test Principle:
The determination of soluble antigen concentration by turbidimetric methods involves the reaction with specific antiserum to form insoluble complexes. When light is passed through the suspension formed a portion of the light is transmitted and focused onto a photodiode by an optical lens system. The amount of transmitted light is indirectly proportional to the specific protein concentration in the test sample. Concentrations are automatically calculated by reference to a calibration curve stored within the instrument.

Latex-enhanced antibodies. Some antigen-antibody reactions do not form sufficiently large immune complexes to be detected turbidimetrically. If the antibody is coated

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onto latex particles of a suitable size, the light scattering ability of the immune complexes formed with antigen is enhanced sufficiently to enable turbidimetric detection.

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

### 1. Analytical performance:

#### a. Precision/Reproducibility:

The intra-assay precision was determined by testing six serum samples twenty one times. The inter-assay precision was determined by testing three serum samples three times with two runs per day for 21 days. Results are summarized below.

Intra-assay:

| n=21 | β2M | | |
| --- | --- | --- | --- |
| | SD | Concentration (mg/L) | % CV |
| Sample 1 | 0.02 | 0.97 | 1.8 |
| Sample 2 | 0.03 | 4.94 | 0.6 |
| Sample 3 | 0.09 | 16.43 | 0.6 |
| Sample 4 | 0.035 | 20.74 | 1.7 |
| Sample 5 | 0.065 | 31.81 | 2.0 |
| Sample 5 | 0.62 | 36.53 | 1.7 |

Inter-assay:

| n=21 | β2M | | |
| --- | --- | --- | --- |
| | SD | Concentration (mg/L) | % CV |
| Sample 1 | 0.06 | 0.97 | 5.8 |
| Sample 2 | 0.10 | 4.94 | 2.1 |
| Sample 3 | 0.31 | 16.43 | 1.9 |

#### b. Linearity/assay reportable range:

Serum linearity across the assay range (0.3- 40.0 mg/L) was confirmed by testing three sera with concentrations from 9.17- 36.46 mg/L. The samples were serially diluted 10 times with buffer (1:10) down to the lower measuring range (0.3 mg/L). All testing were performed twice. The regression plot equations where y is the measured level of serum β2M concentration and x the theoretical concentration were:

$$
y = 0.9896x + 0.150 \, (\mathrm{mg/L}), \, r^2 = 0.9996 \text{ for serum } \beta 2 \mathrm{M}
$$

Urine linearity across the assay range (0.6-20.0 mg/L) was confirmed by testing two samples with concentrations of 0.853 mg/L and 18.300 mg/L. The samples were serially diluted off-line 10 times with buffer (1:20) down to the lower measuring range (0.04 mg/L). All testing were performed three times. The regression plot equations where y is the measured level of urine β2M concentration and x the theoretical concentration were:

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$$
y = 0.9929x - 0.069 \text{ (mg/L)}, r^2 = 0.9992 \text{ for urine } \beta_2M
$$

c. Traceability, Stability, Expected values (controls, calibrators, or methods):
An internal reference standard (IR8494) was assigned by comparison to the $1^{\text{st}}$ International Standard for Beta-2 Microglobulin (NIBSC Code: B2M).

Stability: The expiration date claims are 6 months for the $\beta_2M$ unopened Kit and 30 days for the opened $\beta_2M$ kit and on-board $\beta_2M$ kit.

d. Detection limit:
The detection limit was determined by testing a blank sample, the lowest calibrator, and a sample with value close to the blank sample (0.006 mg/L) 60 times each. The very low level sample (diluted low level urine) gave a mean value of 0.00672 absorbance unit giving an estimated value of 0.12 mg/L which is considered to be the limit of detection. The limit of quantitation for this assay is defined as the lowest point of the calibration curve: 0.03 mg/L based upon a neat sample dilution (minimum used for urine).

e. Analytical specificity:
Serum Interference by endogenous and other substances: A known $\beta_2M$ serum sample was tested with the following interferents: 5 g/L hemoglobin, 200 mg/L bilirubin, 10722 FTU of chyle and 600 IU/mL RF. No significant interference by these substances was observed.

Urine Interference by endogenous and other substances: A known $\beta_2M$ urine sample was tested in with the following interferents: 238 mg/L hemoglobin, 200 mg/L bilirubin, 200 mg/L ascorbic acid, and 100 mg/dL therapeutic IgG. No significant interference by these substances was observed.

The package insert states that "turbidimetric assays are not suitable for measurement of highly lipemic or hemolyzed samples, or samples containing high levels of circulating immune complexes due to the unpredictable degree of non-specific scatter these sample types might generate. Unexpected results should be confirmed using alternative assay method".

Antigen excess effect:
The possibility of antigen excess occurring when using the device on The Binding Site $\mathrm{SPA}_{\mathrm{PLUS}}^{\mathrm{TM}}$ was evaluated with samples with $\beta_2M$ concentration above the assay range (40 mg/L). No antigen excess effect up to 76 mg/L of $\beta_2M$ was observed. The package insert states: "The antigen excess protection (P flag) on the $\mathrm{SPA}_{\mathrm{PLUS}}$ has been tested up to a level which is equivalent to 760 mg/L.

f. Assay cut-off:
Not provided

2. Comparison studies:

a. Method comparison with predicate device:
Testing was performed on 103 sera (31 normal and 72 clinical) and 49 urine

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samples (24 normal and 25 known renal impaired samples).

The table below shows the comparison of 103 sera (ranging from 0.51 – 19.44 mg/L) and 49 urine samples (ranging from 0.04- 20.2 mg/L) that were tested with the Binding Site $\mathrm{SPAPLUS}^{\mathrm{TM}}$ $\beta 2\mathrm{M}$ assay and the predicate device Behring BNII System. Regression analysis of these samples is summarized below:

| The Binding Site SPAPLUS™ vs. Behring BNII analyzer | n | Regression Equation | r |
| --- | --- | --- | --- |
| Serum | 103 | y = 1.02x - 0.11 (mg/L) | 0.996 |
| Urine: | 49 | y = 0.97x + 0.00 (mg/L) | 0.993 |

b. Matrix comparison: Not applicable.

3. Clinical studies:

a. Clinical Sensitivity and specificity: None provided.
b. Other clinical supportive data (when a. is not applicable): Not applicable.

4. Clinical cut-off: Not provided

5. Expected values/Reference range: Adult normal serum range was assessed on a total 150 normal adult sera samples from healthy US blood donors (age 18-64 years old). The urine range was assessed on a total of 116 normal urines from UK healthy adult donors (age 18-65 years old). The assays were performed on the Binding Site $\mathrm{SPA}_{\mathrm{plus}}^{\mathrm{TM}}$ analyser. A non-parametric distribution of $\beta 2\mathrm{M}$ results were seen that gave a 95 percentile reference interval of $0.8 - 2.34\mathrm{mg / L}$ with a mean of $1.33\mathrm{mg / L}$ and a median of $1.26\mathrm{mg / L}$ for serum; and 95 percentile reference interval of $0.03 - 0.202\mathrm{mg / L}$ with a mean of $0.08\mathrm{mg / L}$ and a median of $0.07\mathrm{mg / L}$ for urine.

# 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.