The AquaLite® Ferritin Bioluminescent Immunoassay (BIA) Kit (or the AquaLite® Ferritin Assay) is an in vitro diagnostic product intended for use in clinical laboratories for the quantitative determination of human ferritin in serum and plasma. Ferritin measurements are used in the diagnosis of diseases affecting iron metabolism.

Device Description

The AquaLite® Ferritin Bioluminescent Immunoassay Kit uses a mouse monoclonal antiferritin antibody that is pre-coated onto polystyrene tubes (solid phase). Samples (serum or plasma), or appropriate calibrators or controls, are pipetted (25 uL) into the precoated tubes. A mouse monoclonal anti-ferritin antibody covalently linked to AquaLite® (150 uL) is then added to the tubes. Ferritin in the sample simultaneously combines with the antibody on the solid phase and conjugate antibody to form an immune complex or "sandwich" bound to the solid phase. Complex formation is complete after a 60-minute incubation period at room temperature on a standard orbital shaker. The tubes are then washed to remove unbound conjugate.

The washed tubes are placed in a luminometer that is capable of reading a triggered, flash-type reaction in 12 x 75 mm tubes. Injection of the calcium trigger buffer causes AquaLite® to oxide its self-contained luciferin molecule, producing a flash of light, which is measured by the luminometer. The intensity of the light emitted from antibody bound to the tubes is directly proportional to the concentration of the ferritin in the sample. To calculate results, the light intensity (in relative light units, RLU) of the ferritin calibrators is plotted against ferritin concentration (in ng per mL) to yield a calibration curve. This curve is used to relate the light intensity generated from the samples and controls to ferritin concentration in ng/mL.

Note: Samples that generate signals greater than the signal from the highest calibrator are off-scale. These samples must be diluted and re-assayed. Remember to multiply the results from diluted samples by the dilution factor used.

AI/ML Overview

The provided document describes the SeaLite Sciences, Inc. AquaLite® Ferritin Assay, a bioluminescent immunoassay for the quantitative determination of human ferritin in serum and plasma.

Here's an analysis of the acceptance criteria and study information:

1. Table of Acceptance Criteria and Reported Device Performance

Performance Characteristic	Acceptance Criteria (Implicit)	Reported Device Performance	Comments
Sensitivity	Not explicitly stated (e.g., lower limit of detection based on clinical need)	0.03 ng/mL	Determined by adding the mean signal of 20 replicates of the zero-level calibrator + 2 standard deviations.
Specificity (Cross-Reactivity)	Not explicitly stated (e.g., % cross-reactivity with structurally similar molecules within a certain range).	85% and 103% for spiked liver ferritin in two human serum samples.	Measures "intact ferritin." The values indicate good consistency in measuring spiked ferritin.
High Dose Hook Effect	No hook effect within a clinically relevant range.	No high dose hook effect occurs prior to 10,000 ng/mL.	This is an important safety and accuracy measure for immunoassays.
Intra-assay Precision (CV)	Not explicitly stated (e.g., <10% or <15% for acceptable precision).	368.4 ng/mL: 8.6% CV123.9 ng/mL: 3.6% CV40.5 ng/mL: 5.4% CV	Shows good within-run reproducibility.
Inter-assay Precision (CV)	Not explicitly stated.	50.9 ng/mL: 4.5% CV121.6 ng/mL: 3.7% CV363.2 ng/mL: 9.3% CV	Shows good day-to-day or run-to-run reproducibility.
Method Comparison (Correlation with Predicate Device)	Not explicitly stated (e.g., R-value > 0.95 and slope close to 1, y-intercept close to 0).	Correlation coefficient (R) = 0.98Slope = 0.72Y-intercept = 15.9	Indicates strong correlation with the predicate device (Chiron Diagnostics ACS:180 Ferritin Assay), though the slope suggests a slight systemic difference.
Linearity and Nonparallelism (Recovery)	Not explicitly stated (e.g., recovery within 80-120%).	Sample A: 96.6-119.5%Sample B: 109.5-121.9%Sample C: 86.4-92.3%	Generally good recovery across dilutions, demonstrating linearity.
Spike and Recovery	Not explicitly stated (e.g., recovery within 80-120%).	Sample 1: 86%Sample 2: 100%Sample 3: 84%	Good recovery of spiked ferritin, showing the assay can accurately measure ferritin اضافه.
Recovery in Serum and Plasma	Not explicitly stated (e.g., differences within a certain percentage, demonstrating matrix equivalence).	% Recovery in EDTA plasma vs. Serum: 85-103%	Demonstrates the assay performs similarly in both serum and EDTA plasma.
Effect of Common Interferents (Recovery)	Not explicitly stated (e.g., recovery within 90-110% in the presence of interferents).	Hemoglobin (500mg/dL): 98%Bilirubin (20mg/dL): 90%Triglycerides (3000mg/dL): 99%	Shows the assay is not significantly affected by these common interferents within the tested concentrations.

2. Sample Sizes and Data Provenance

Test Set Sample Sizes:
- Sensitivity: 20 replicates of a zero-level calibrator.
- Specificity: 2 human serum samples (spiked).
- Precision (Intra-assay): N = 20 per concentration level for commercial controls.
- Precision (Inter-assay): N = 20 (2 replicates x 10 assays) for commercial controls.
- Method Comparison: N = 99 patient samples.
- Linearity: 3 human serum samples, each diluted.
- Spike and Recovery: 3 normal human serum samples.
- Recovery in Serum and Plasma: 6 normal subjects.
- Effect of Common Interferents: Pooled normal human serum (spiked).
Data Provenance: The studies were conducted at SeaLite Sciences. All samples appear to be human (serum, plasma, human spleen ferritin for spiking). The document does not specify the country of origin for the human samples. The studies are by nature prospective for the device's validation, as they were specifically designed and executed to evaluate the AquaLite® Ferritin Assay.

3. Number of Experts and Qualifications for Ground Truth

This device is an in vitro diagnostic (IVD) assay for quantitative measurement. The "ground truth" is typically established instrumentally or by reference methods/materials rather than expert consensus on images or clinical assessments.

No "experts" in the sense of clinical reviewers (e.g., radiologists) were used to establish ground truth.
The "ground truth" for quantitative assays like this relies on:
- Reference materials: Commercial controls with known ferritin concentrations, calibrators traceable to international standards (e.g., WHO 2nd IS 80/578 for spike and recovery).
- Predicate device: The Chiron Diagnostics ACS:180 Ferritin Assay served as a reference for method comparison.

4. Adjudication Method for the Test Set

Not applicable. As a quantitative IVD assay, adjudication by multiple human readers (as in image-based diagnostic aids) is not relevant. The results are numerical measurements.

5. Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study

No, a multi-reader multi-case (MRMC) comparative effectiveness study was not done. Such studies are typically performed for diagnostic imaging devices or AI-powered clinical decision support tools where human interpretation plays a significant role. This document describes an automated laboratory assay.

6. Standalone Performance

Yes, the studies listed (Sensitivity, Specificity, Precision, High Dose Hook Effect, Linearity, Spike and Recovery, Recovery in Serum and Plasma, Effect of Common Interferents) describe the standalone performance of the AquaLite® Ferritin Assay. These evaluations assess the algorithm (the assay's chemical/biological reaction and measurement system) without human interpretation in the results generation process itself. The "Method Comparison" also shows the standalone performance relative to an existing predicate device.

7. Type of Ground Truth Used

The ground truth used is primarily based on:

Known concentrations: From commercial controls, calibrators, and spiked samples.
Reference methods/materials: Comparison against a legally marketed predicate device (Chiron Diagnostics ACS:180 Ferritin Assay) and internationally recognized standards (WHO 2nd IS 80/578 for spleen ferritin).

8. Sample Size for the Training Set

The document does not specify a separate "training set" sample size. For an immunoassay like this, the "training" typically refers to the development and optimization of the assay's reagents and parameters, which is an iterative process often involving experimental work rather than a distinct, labeled "training set" of patient data as seen in machine learning contexts. The calibrators and controls used would be part of the ongoing validation and quality control, but not a separate "training set" for an algorithm in the modern sense.

9. How the Ground Truth for the Training Set Was Established

Not explicitly stated in terms of a formal "training set ground truth." However, the equivalent process for an immunoassay involves:

Development and Optimization: During the development phase, reagents, incubation times, concentrations, and detection methods are optimized using known ferritin concentrations (potentially from purified ferritin, spiked samples, or commercial controls) to achieve desired performance characteristics.
Calibrators: The assay uses calibrators with established ferritin concentrations to generate its standard curve. These calibrators are usually traceable to primary reference materials.
Quality Control (QC) Materials: Commercial QC materials with defined ferritin values are used to monitor assay performance over time.

Summary

{0}------------------------------------------------

K970612

JUN 24 1997

510(k) SUMMARY

I. GENERAL INFORMATION

Trade or proprietary name - SeaLite Sciences, Inc. AquaLite® Ferritin Assay

Common or usual name - Bioluminescent immunoassay (BIA)

Classification name - FDA has classified ferritin test systems intended for the measurement of ferritin in the diagnosis of diseases affecting iron metabolism, such as hemochromatosis (iron overload) and iron deficiency anemia as Class II devices. (21 C.F.R. § 866.5340)

Submitter's Name and Address:	Cathryn C. CambriaDirector, Regulatory Affairs and QualityAssuranceSeaLite Sciences, Inc.3000 Northwoods ParkwaySuite 200Norcross, GA 30071(800) 874-4471, ext. 227
Submission Date:	February 14, 1997
Legally Marketed DeviceTo Which Claim SubstantialEquivalence:	Chiron Diagnostics ACS:180 Ferritin Assay

INDICATIONS FOR USE II.

{1}------------------------------------------------

DEVICE DESCRIPTION III.

IV. SUMMARY OF STUDIES AND TECHNOLOGICAL CHARACTERISTICS

Studies on the AquaLite® Ferritin Assay were conducted at SeaLite Sciences. The results are summarized below:

Performance Characteristics

1. Sensitivity
  The sensitivity or detection limit of the AquaLite® Ferritin Assay is 0.03 ne/mL. Sensitivity is determined by adding the mean signal of 20 replicates of the zero level calibrator plus two (2) standard deviations above this mean. The ferritin concentration (ng/mL) associated with this calculated signal is defined as the sensitivity of the assay.

{2}------------------------------------------------

2. Specificity

The AquaLite® Ferritin Assay measures intact ferritin. Cross reactivity of the AquaLite® Ferritin assay was determined by measuring the concentration of ferritin in 2 human serum samples spiked with 300 ng/mL liver ferritin. Percent crossreactivity (%) is reported below:

EndogenousFerritin(ng/mL)	FerritinExpected(ng/mL)	FerritinObserved(ng/mL)	Cross-Reactivity(%)
79	379	323	85
139	439	451	103

High Dose Hook Effect 3.

No high dose hook effect occurs prior to 10,000 ng/mL ferritin.

4. Precision

(a) Two serum commercial controls containing Intra-assay precision. ferritin at the following concentrations were assayed to determine intraassay precision. (Total N = 20 per concentration level.)

Ferritin Level (ng/mL)	% CV
368.4	8.6
123.9	3.6
40.5	5.4

{3}------------------------------------------------

(b) Inter-assay precision. Commercial controls containing ferritin at the following concentrations were assayed in duplicate repetitively. Ten (10) assays were performed and a new standard curve was generated for each assay. The inter-assay precision observed for the solutions (Total n = 2 x 10 = 20) are shown below.

Ferritin Level (ng/mL)	% CV
50.9	4.5%
121.6	3.7%
363.2	9.3%

5. Method Comparison

The AquaLite® Ferritin Assay was used to assay patient samples (N=99) that were previously assayed by a commercially available chemiluminometric immunoassay. A slope of 0.72 with a y-intercept of 15.9 was obtained. The correlation coefficient was 0.98.

6. Linearity and Nonparallelism

Three human serum samples containing the levels of endogenous ferritin shown below were diluted as indicated using Calibrator A (0 ng/mL) and assayed in duplicate. All concentrations are in ng/mL.

{4}------------------------------------------------

SAMPLEID	DILUTIONFACTOR	OBSERVED(ng/mL)	FERRITINEXPECTED(ng/mL)	RECOVERY(%)
A	Undiluted1:21:41:8	160.177.347.423.9	--804020	--96.6118.5119.5
B	Undiluted1:21:41:8	294.1160.984.445.1	--1477427	--109.5114.1121.9
C	Undiluted1:21:41:8	878383.0189.6101.4	--439219.5109.8	--87.286.492.3

7. Spike and Recovery

Three normal human serum samples were diluted with 120 ng/mL human spleen ferritin (WHO 2nd IS 80/578). The spiked samples were assayed using the AquaLite® Ferritin Assay. All values are in ng/mL.

Sample	Unspiked(ng/mL)	SpikeRecovered(ng/mL)	SpikedExpected(ng/mL)	%
1	1260	595	690	86
2	58	90	90	100
3	406	221	263	84

8. Recovery in Serum and Plasma

Blood samples from 6 normal subjects were prepared as sera (standard tubes) as Ferritin was quantified using the AquaLite® Ferritin well as EDTA plasma. Assay. Recovered ferritin was compared with the ferritin recovered in serum (standard technique). The data demonstrate that there are no significant differences among serum and EDTA plasma when using the AquaLite® Ferritin Assay.

{5}------------------------------------------------

Sample	Serum(ng/mL)	EDTA(ng/ml)	%
1	63.9	62.0	97
2	44.1	38.7	88
3	62.8	53.6	85
4	109.6	110.9	101
5	105.2	108.8	103
6	63.3	62.6	99

9. Effect of Common Interferents

Pooled normal human serum was spiked with preparations of hemoglobin, bilirubin, and triglycerides to the levels shown below. Ferritin was quantified using the AquaLite® Ferritin assay. Recovered ferritin was compared to the ferritin recovered in normal serum. The data demonstrate that the AquaLite® Ferritin assay is not significantly affected by hemoglobin, bilirubin, or triglycerides at the levels tested.

Analyte	Observed	Expected	%
Hgb at 500 mg/dL	113.3	115.6	98
Bilirbn at 20 mg/dL	114.2	115.6	90
Trig at 3000 mg/dL	116.2	128.9	99

V. POTENTIAL ADVERSE EFFECTS OF THE DEVICE ON HEALTH

Use Universal Precautions. No known test method can offer complete Caution: assurance that products derived from human serum are pathogen-free; therefore, handle all materials of human origin as though they were potentially infectious.

Sodium azide is used as a preservative. This preservative may react with metallic plumbing to from explosive metal azides. Flush with large volumes of water when disposing of materials containing sodium azide.

D-6

{6}------------------------------------------------

As an in vitro diagnostic test, there are not direct adverse effects on the health of a patient from the use of this product. However, failure of the device to perform as indicated, the contamination of reagents, the use of reagents past the labeled expiration dates, the use of improper specimens, or human error during the performance of the test may lead to erroneous results and possible improper patient management.

VI. CONCLUSIONS DRAWN FROM STUDIES

The data from the studies conducted demonstrate that the performance of SeaLite Sciences, Inc. AquaLite® Ferritin Assay is similar and substantially equivalent to that of other commercially available assays for ferritin.

{7}------------------------------------------------

Image /page/7/Picture/1 description: The image shows the logo for the U.S. Department of Health & Human Services. The logo features a stylized depiction of an eagle with its wings spread, rendered in black lines. The words "DEPARTMENT OF HEALTH & HUMAN SERVICES • USA" are arranged in a circular pattern around the eagle, also in black.

Food and Drug Administration 2098 Gaither Road Rockville MD 20850

Cathryn C. Cambria Director, Regulatory Affairs & Quality Assurance SeaLite Sciences, Inc. 3000 Northwoods Parkway -----Suite 200 Norcross, GA 30071

JUN 2 4 1997

Re: K970612/S001 Trade Name: SeaLite Sciences, Inc. AquaLite® Ferritin Assay Regulatory Class: II Product Code: DBF Dated: May 08, 1997 Received: May 12, 1997

Dear Ms. Cambria:

We have reviewed your Section 510(k) notification of intent to market the device referenced above and we have determined the device is substantially equivalent (for the indications for use stated in the enclosure) to devices marketed in interstate commerce prior to May 28, 1976, the enactment date of the Medical Device Amendments or to devices that have been reclassified in accordance with the provisions of the Federal Food, Drug, and Cosmetic Act (Act). You may, therefore, market the device, subject to the general controls provisions of the Act. The general controls provisions of the Act include requirements for annual registration, listing of devices, good manufacturing practice, labeling, and prohibitions against misbranding and adulteration.

If your device is classified (see above) into either class II (Special Controls) or class III (Premarket Approval), it may be subject to such additional controls. Existing major regulations affecting your device can be found in the Code of Federal Regulations, Title 21, Parts 800 to 895. A substantially equivalent determination assumes compliance with the Good Manufacturing Practice for Medical Devices: General (GMP) regulation (21 CFR Part 820) and that, through periodic GMP inspections, the Food and Drug Administration (FDA) will verify such assumptions. Failure to comply with the GMP regulation may result in regulatory action. In addition, FDA may publish further announcements concerning your device in the Federal Register.

Please note: this response to your premarket notification submission does not affect any obligation you might have under sections 531 through 542 of the Act for devices under the Electronic Product Radiation Control provisions, or other Federal Laws or Regulations.

Under the Clinical Laboratory Improvement Amendments of 1988 (CLIA-88), this device may require a CLIA complexity categorization. To determine if it does, you should contact the Centers for Disease Control and Prevention (CDC) at (770)488-7655.

{8}------------------------------------------------

Page 2

This letter will allow you to begin marketing your device as described in your 510(k) premarket notification. The FDA finding of substantial equivalence of your device to a legally marketed predicate device results in a classification for your device and thus, permits your device to proceed to the market.

If you desire specific advice for your device on our labeling regulation (21 CFR Part 801 and additionally 809.10 for in vitro diagnostic devices), please contact the Office of Compliance at (301) 594-4588. Additionally, for questions on the promotion and advertising of your device, please contact the Office of Compliance at (301) 594-4639. Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21 CFR 807.97). Other general information on your responsibilities under the Act may be obtained from the Division of Small Manufacturers Assistance at its toll free number (800) 638-2041 or at (301) 443-6597 or at its internet address "http://www.fda.gov/cdrh/dsmamain.html"

Sincerely yours,

Steven Autman

Steven I. Gutman, M.D., M.B.A. Director Division of Clinical Laboratory Devices Office of Device Evaluation Center for Devices and Radiological Health

Enclosure

{9}------------------------------------------------

Page 1 of _l

510(k) Number (if known): _

AquaLite® Ferritin Bioluminescent Immunoassay (BIA) Kit (or the Device Name: AquaLite® Ferritin Assay)

Indications for Use:

The AquaLite® Ferritin Bioluminescent Immunoassay (BIA) Kit (or AquaLite® Ferritin Assay) is an in vitro diagnostic product intended for use in clinical laboratories for the quantitative measurement of ferritin in human serum and plasma. Ferritin measurements are used in the diagnosis of diseases affecting iron metabolism.

Peter E. Mafini

(PLEASE DO NOT WRITE BELOW THIS LINE CONTINUE ON ANOTHER PAGE IF NEEDED)

Concurrence of CDRH, Office of Device Evaluation (ODE)

Prescription Use_V (Per 21 CFR 801.109)

Over-The-Counter Use

(Optional Format 1-2-96)

Regulation Number and Section

§ 866.5340 Ferritin immunological test system.

(a)
Identification. A ferritin immunological test system is a device that consists of the reagents used to measure by immunochemical techniques the ferritin (an iron-storing protein) in serum and other body fluids. Measurements of ferritin aid in the diagnosis of diseases affecting iron metabolism, such as hemochromatosis (iron overload) and iron deficiency amemia.(b)
Classification. Class II (performance standards).