The A/C Enzymatic Homocysteine Assay is intended for the quantitative determination of total homocysteine (tHCY) in human plasma or serum.
The device can assist in the diagnosis and treatment of patients suspected of having hyperhomocysteinemia.
The A/C Enzymatic Homocysteine Assay Kit is only for in vitro diagnostic use.

Device Description

The A/C Enzymatic Homocysteine Assay is calibrated with A/C Enzymatic Homocysteine Assay Calibrators. The A/C Enzymatic Homocysteine Assay is assayed for the verification of the accuracy and precision on the Hitachi 912 Automatic Analyzer.

The A/C Enzymatic Homocysteine Assay measures tHCY. The principle of the assay is that recombinant homocysteinase (rHCYase) produces hydrogen sulfide (H2S) from tHCY, which is quantified by use of N,N-dibutyl phenylene diamine (DBPDA), the combination of which forms a chromophore.

The A/C Enzymatic Homocysteine Assay on the Hitachi 912 Automatic Analyzer used four reagents, a number compatible with implementation on the Hitachi 912 Automatic Analyzer. We used 30 uL of EDTA plasma in a dithiothreitol (DTT) reduction reaction (1 mmol/L DTT, 0.2% Triton X-100 in 40 mmol/L sodium phosphate buffer [pH 8.3]) for 1.5 minutes to release bound homocysteine. The rHCYase reaction (0.05 mg/ml in 40 mmol/L sodium phosphate buffer [pH 8.3] with 20 umol/L pyridoxal 5-phosphate [PLP]) is then run for 3.5 minutes. The DBPDA chromophore (12.5 mmol/L DBPDA in 1.5 N H2SO4) is then added and 5 minutes later, an oxidant, potassium ferricyanide (5 mmol/L K3Fe(CN), in 10 mmol/L sodium phosphate buffer [pH 7.6]), is added. Five minutes after addition of oxidant, the end-points are read at absorbances of 700 and 660 nm. As the assay is based on an increase in absorbance over baseline, no blank without enzyme was used. The detection limit of the assay is 1.5 umol/L defined by quantification of a serial dilution of a plasma sample of tHCY diluted to 0.77 umol/L. The limit of quantification is defined as the lowest concentration measured having a CV <20%. The linear range extends to at least 80 umol tHCY/L as determined by measuring varying amounts of homocysteine in phosphate buffered saline.

The within-run imprecisions (CV) run over 8 repeats were 4.8% at 8.9 umol/L tHCY; 3.0% for 14.9 umol/L tHCY; and 4.5% for 25 umol/L tHCY. Between-assay imprecisions (CV) over 10 days were 7.8% for 8.8 umol/L tHCY: 5.9% for 15 umol/L tHCY; and 4.9% for 25 umol/L tHCY. These imprecisions are with in ranges reported for currently-used assays including the Bio-Rad HCY assay on HPLC, which is used as a comparison method in the present study.

The results showed that L-cysteine (L-CYS) in the physiological concentrations (0-200 µmol/L) had less than 10% interference and L-methionine (L-MET) in the physiological concentrations (0-200 umol/L) had no interference in the A/C Enzymatic Homocysteine Assay on the Hitachi 912 Automatic Analyzer.

The A/C Enzymatic Homocysteine Assay on the Hitachi 912 Automatic Analyzer is a four-step reaction. The total assay takes 15 minutes, and the through-put is 360 tests per hour.

AI/ML Overview

Here's an analysis of the provided text regarding the A/C Enzymatic Homocysteine Assay, focusing on acceptance criteria and the supporting study:

1. Table of Acceptance Criteria and Reported Device Performance

The document doesn't explicitly state formal "acceptance criteria" in a clear, quantified manner for regulatory approval. Instead, it describes performance characteristics and compares them to a predicate device. The performance is deemed acceptable because it is within ranges reported for currently-used assays including the predicate.

Here's a table summarizing the performance characteristics presented, with an interpretation of implied acceptance based on the comparison to existing assays:

Performance Metric	Implied Acceptance Criteria (based on predicate equivalence)	Reported Device Performance (A/C Enzymatic Homocysteine Assay)
Linear Range	Similar to or better than predicate	Extends to at least 80 umol tHCY/L
Detection Limit	Not explicitly stated as comparative metric	1.5 umol/L
Limit of Quantification (LOQ) CV	Not explicitly stated as comparative metric	<20% for the lowest concentration measured (implied at 1.5 umol/L or lower)
Within-run Imprecision (CV)	Within ranges reported for currently-used assays (including Bio-Rad HCY assay on HPLC)	4.8% (8.9 umol/L), 3.0% (14.9 umol/L), 4.5% (25 umol/L)
Between-assay Imprecision (CV)	Within ranges reported for currently-used assays (including Bio-Rad HCY assay on HPLC)	7.8% (8.8 umol/L), 5.9% (15 umol/L), 4.9% (25 umol/L)
L-Cysteine Interference	Less than 10% interference	Less than 10% interference (0-200 µmol/L)
L-Methionine Interference	No interference	No interference (0-200 umol/L)
Method Comparison (Regression Eq.)	Close to y = x (slope ~1, intercept ~0)	y = 0.98 + 1.90 (r = 0.977)
Method Comparison (Mean Difference)	Not significantly different from predicate	-1.62 umol/L (SD=2.33)
Method Comparison (Correlation)	High correlation (r value close to 1)	Pearson r = 0.977; correlation of differences with concentration: r = 0.12, p = 0.185 (not significant)

2. Sample Size Used for the Test Set and Data Provenance

Sample Size for Test Set: 121 plasma samples.
Data Provenance: Not explicitly stated. The document refers to "plasma samples," but does not specify their country of origin or whether they were collected retrospectively or prospectively.

3. Number of Experts Used to Establish the Ground Truth for the Test Set and Qualifications

This device is an in vitro diagnostic (IVD) assay for quantitative determination of a biomarker (homocysteine), not an imaging or clinical decision support AI device that requires human expert review for "ground truth." Therefore, the concept of "experts establishing ground truth" as it applies to subjective assessments is not relevant here.

The "ground truth" for the test set is established by the predicate device, the Bio-Rad Homocysteine by HPLC (k 993107). The performance of the new device is compared directly to the measurements obtained from this established method.

4. Adjudication Method for the Test Set

Not applicable. As this is a quantitative chemical assay, there's no "adjudication" in the sense of resolving conflicting interpretations by multiple human readers. The comparison is statistical between two quantitative measurement methods.

5. If a Multi Reader Multi Case (MRMC) Comparative Effectiveness Study Was Done

No. An MRMC study is relevant for medical imaging or subjective interpretation tasks where multiple human readers assess cases. This document describes the performance of a chemical assay, so an MRMC study is not applicable.

6. If a Standalone (i.e. algorithm only without human-in-the loop performance) Was Done

Yes, in the sense that the A/C Enzymatic Homocysteine Assay operates as a standalone analytical system on the Hitachi 912 Automatic Analyzer. Its performance characteristics (detection limit, linearity, imprecision, interference) and its comparison to the predicate are presented as the algorithm's direct output. There is no human interpretation component being measured against a human-augmented performance.

7. The Type of Ground Truth Used

The primary "ground truth" used for comparison and establishing equivalence in this study is the measurements obtained from a legally marketed predicate device (Bio-Rad Homocysteine by HPLC). This is a common method for IVD assays seeking 510(k) clearance by demonstrating "substantial equivalence."

8. The Sample Size for the Training Set

Not applicable. This device is a chemical assay, not a machine learning or AI algorithm in the contemporary sense that typically requires a "training set." The assay's parameters would have been developed and optimized through laboratory work and internal studies, but not in the framework of a distinct "training set" and "test set" common in AI/ML validation.

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

Not applicable, as there isn't a "training set" in the context of an AI/ML algorithm. The assay's chemical reagents and reaction conditions were presumably optimized through standard chemical and enzymatic assay development processes based on biochemical principles and laboratory experimentation.

Summary

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Summary of Safety and Effectiveness Information Supporting a Substantially Equivalent Determination

Submitter Information 1.

A/C Diagnostics Division AntiCancer Inc. 7917 Ostrow Str. San Diego, California 92111 Phone: (858)654-2555 FAX: (858)268-4175 e-mail: all(@anticancer.com

Contact Person:

Submitter:

Yuying Tan, M.D. Research Manager AntiCancer Inc.

Date of Summary Preparation: February 18, 2003

2. Device Information

Device Name:

A/C Enzymatic Homocysteine Assay

Classification Name: Class: Product Code: Homocysteine Assay II LPS

3. Predicate Device Information

Device Name:	Bio-Rad Homocysteine by HPLCBio-Rad LaboratoriesDiagnostics Group4000 Alfred Nobel DriveHercules, California 94547
--------------	------------------------------------------------------------------------------------------------------------------------------------

510(k) Number: K993107

K030754

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Information of Manufacturer 4.

Manufacturer:
---------------

Bioserv Corporation 5340 Eastgate Mall San Diego, CA 92121 Telephone: (858) 450-3123 (858) 450-0785 FAX:

FDA establishment registration number: US FDA 2027352

Contact Person :

Mary Richardson Quality Assurance Manager Bioserv Corporation

న. Statement of Intended Use

The A/C Enzymatic Homocysteine Assay is intended for the quantitative determination . of total homocysteine (tHCY) in human plasma or serum.
The device can assist in the diagnosis and treatment of patients suspected of having . hyperhomocysteinemia.
The A/C Enzymatic Homocysteine Assay Kit is only for in vitro diagnostic use. .

Description of Device 6.

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buffer [pH 7.6]), is added. Five minutes after addition of oxidant, the end-points are read at absorbances of 700 and 660 nm. As the assay is based on an increase in absorbance over baseline, no blank without enzyme was used. The detection limit of the assay is 1.5 umol/L defined by quantification of a serial dilution of a plasma sample of tHCY diluted to 0.77 umol/L. The limit of quantification is defined as the lowest concentration measured having a CV <20%. The linear range extends to at least 80 umol tHCY/L as determined by measuring varying amounts of homocysteine in phosphate buffered saline.

The A/C Enzymatic Homocysteine Assay on the Hitachi 912 Automatic Analyzer is a four-step reaction. The total assay takes 15 minutes, and the through-put is 360 tests per hour.

7. Method Comparison

To establish equivalence to an existing device, and thus establish the safety and effectiveness. the A/C Enzymatic Homocysteine Assay on Hitachi 912 Automatic Analyzer is compared to the Bio-Rad Homocysteine by HPLC (k 993107).

We assayed 121 plasma samples with the A/C Enzymatic Homocysteine Assay on the Hitachi 912 Automatic Analyzer (y) and with the Bio-Rad Homocysteine Assay by HPLC (x). The regression equation was y = 0.98 + 1.90 (r = 0.977). The mean difference (SD) between the A/C Enzymatic Homocysteine Assay and the Bio-Rad Homocysteine by HPLC was -1.62 umol/L (SD=2.33). Differences were not significantly correlated with homocysteine concentration (Pearson r = 0.12, p = 0.185).

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Image /page/3/Picture/1 description: The image shows the logo for the U.S. Department of Health & Human Services. The logo is a circular seal with the words "DEPARTMENT OF HEALTH & HUMAN SERVICES - USA" arranged around the top half of the circle. Inside the circle is an abstract symbol that resembles an eagle or bird in flight, composed of three curved lines.

Food and Drug Administration 2098 Gaither Road Rockville MD 20850

JUL 1 1 2003

Yuying Tan, M.D. Research Manager AntiCancer Inc. A/C Diagnostics Division 7917 Ostrow Street San Diego, CA 92111

Re: K030754

Trade/Device Name: A/C Enzymatic Homocysteine Assay Regulation Number: 21 CFR 862.1377 Regulation Name: Urinary homocystine (non-quantitative) test system Regulatory Class: Class II Product Code: LPS Dated: May 15, 2003 Received: May 16, 2003

Dear Dr. Tan:

We have reviewed your Section 510(k) premarket notification of intent to market the device referenced above and have determined the device is substantially equivalent (for the indications for use stated in the enclosure) to legally marketed predicate 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) that do not require approval of a premarket approval application (PMA). 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 (PMA). it may be subject to such additional controls. Existing major regulations affecting your device can be found in Title 21, Code of Federal Regulations (CFR), Parts 800 to 895. In addition, FDA may publish further announcements concerning your device in the Federal Register.

Please be advised that FDA's issuance of a substantial equivalence determination does not mean that FDA has made a determination that your device complies with other requirements of the Act or any Federal statutes and regulations administered by other Federal agencies. You must comply with all the Act's requirements, including, but not limited to: registration and listing (21 CFR Part 807); labeling (21 CFR Parts 801 and 809); and good manufacturing practice requirements as set forth in the quality systems (QS) regulation (21 CFR Part 820).

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Page 2 - .

This letter will allow you to begin marketing your device as described in your Section 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 information about the application of labeling requirements to your device, or questions on the promotion and advertising of your device, please contact the Office of In Vitro Diagnostic Device Evaluation and Safety at (301) 594-3084. Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21CFR Part 807.97). You may obtain other general information on your responsibilities under the Act from the Division of Small Manufacturers, International and Consumer Assistance at its toll-free number (800) 638-2041 or (301) 443-6597 or at its Internet address http://www.fda.gov/cdrh/dsma/dsmamain.html.

Sincerely yours,

Steven Putman

Steven I. Gutman, M.D., M.B.A. Director Office of In Vitro Diagnostic Device Evaluation and Safety Center for Devices and Radiological Health

Enclosure

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Appendix-6

K 030754

Indications for Use:

Enzymatic Homocysteine Assay is intended A/C for the ● The quantitative determination of total homocysteine (tHCY) in human plasma or serum.
The device can assist in the diagnosis and treatment of patients . suspected of having hyperhomocysteinemia.
The A/C Enzymatic Homocysteine Assay is only for in vitro diagnostic ● use.

/ prescription use

Albert Juh
Division Sign-Off for Jean Cooper

Office of In Vitro Diagnostic Device Evaluation and Safety

510(k) 050754

Regulation Number and Section

§ 862.1377 Urinary homocystine (nonquantitative) test system.

(a)
Identification. A urinary homocystine (nonquantitative) test system is a device intended to identify homocystine (an analogue of the amino acid cystine) in urine. The identification of urinary homocystine is used in the diagnosis and treatment of homocystinuria (homosystine in urine), a heritable metabolic disorder which may cause mental retardation.(b)
Classification. Class II.