The reticONE™ SYSTEM for EPICS® XL™ Flow Cytometry Systems combines a reagent kit consisting of a Coriphosphine-O dye and a Biological Calibrator, and software for automated analysis of reticulocytes in whole blood using EPICS® XL™ Flow Cytometry Systems with SYSTEM II™ Software. The system is intended "For In Vitro Diagnostic Use" and allows identification and enumeration of reticulocyte percentage and absolute count.

The reticulocyte count is diagnostically useful in discriminating between normal erythropoiesis. It can be useful in the diagnosis or detection of anemal hemorthaging, hemoglobinopathies and certain nutritional or vitamin deficiencies. Decreased or defective red cell production may result in a lower reticulocyte count such as in aplastic anemias. Elevated reticulocyte counts may be observed in clinical conditions where red cell destruction is increased (for example, hemolytic anemias and hypersplenism) or where there is increased red cell production (for example, erythropoietin drug therapy and a response to treated anemia).

Since the kidneys are a primary source of erythropoietin (a hormone that regulates erythroid development), the reticulosyte count is also affected in individuals with renal disease. In cases of renal atrophy, the reticulocyte count will be decreased. In cases of malignancy or hypersplenism, the reticulocyte count will be elevated.

Reticulocyte counts are also used as an indicator of bone marrow recovery following intensive or a bone marrow transplantation. Increased reticulocyte counts in these patients are indicative of bone marrow regeneration.

Device Description

AI/ML Overview

The reticONE™ SYSTEM for EPICS® XL™ Flow Cytometry Systems has demonstrated its performance through a series of studies. The acceptance criteria and device performance are outlined below, along with details regarding the study methodologies.

1. Table of Acceptance Criteria and Reported Device Performance

Acceptance Criteria Category	Specific Test/Metric	Acceptance Criteria	Reported Device Performance	Study Supporting Performance
Stability	Stored (Unstained) Whole Blood Specimens (72 Hours at 2-8 °C)	Reticulocyte percentage and standard deviation demonstrate stability over time.	Stability demonstrated, meeting claims for stored unstained whole blood.	Stability Studies (1.a)
	Stored (Unstained) Whole Blood Specimens (6 Hours at 20-25 °C)	Reticulocyte percentage and standard deviation demonstrate stability over time.	Stability demonstrated, meeting claims for stored unstained whole blood.	Stability Studies (1.b)
	Stored Prepared (Stained) Samples (6 Hours at 20-25 °C)	Reticulocyte percentage and standard deviation demonstrate stability over time.	Stability demonstrated, meeting claims for prepared stained samples.	Stability Studies (1.c)
Carryover	Carryover Percent	Minimal carryover compared to Flow Cytometer Specification.	Minimal carryover demonstrated.	Carryover Study (2)
Linearity	Range of 0.2% to 12.5% reticulocytes	Linearity demonstrated for recovered vs. expected reticulocyte percentage.	Linearity demonstrated over the defined (reportable) range.	Linearity Study (3)
Precision	Within Run (Intralaboratory) Precision	Reproducible measurements (mean reticulocyte percentage, SD, %CV).	Reproducibility demonstrated for all levels.	Precision Study (4.a)
	Interlaboratory Precision	Reproducible measurements across different labs (mean reticulocyte percentage, SD, %CV).	Reproducibility demonstrated across laboratories.	Precision Study (4.b)
	Site Precision	Reproducible measurements (mean reticulocyte percentage, SD, %CV).	Reproducibility demonstrated.	Precision Study (4.c)
Accuracy	Comparison to Retic-COUNT™ (Predicate Device)	Identifies and enumerates comparable numbers of reticulocytes.	Clearly demonstrated comparable identification and enumeration of reticulocytes to Retic-COUNT™.	Accuracy Study (5)

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

Stability Studies (1.a): 3 normal whole blood specimens. These were likely prospective as they involved specific handling and analysis over defined time points. The country of origin for these specimens is not explicitly stated but can be inferred as the USA, given the company's location and FDA submission.
Stability Studies (1.b): 10 normal whole blood specimens. Similar to above, likely prospective and from the USA.
Stability Studies (1.c): 5 normal whole blood specimens. Similar to above, likely prospective and from the USA.
Carryover Study (2): Not explicitly stated as "specimens" but "a single sample" was prepared for each of the High Level and Low Level Retic-Chex control products. These are commercial control products, not patient specimens.
Linearity Study (3): 2 whole blood specimens, serially diluted to create 10 data points. Likely prospective and from the USA.
Precision Study (4.a, 4.c): "Sample [normal or abnormal] whole blood specimens" for each of five levels of reticulocyte percentage. The exact number of specimens is not specified beyond "for each of five levels."
Precision Study (4.b): Samples from a single normal whole blood specimen.
Accuracy Study (5): "Normal and abnormal whole blood specimens were collected from geographically diverse populations of males and females unselected as to race and ranging in age from 18 to 85 years." The exact number of specimens is not specified. The provenance is described as "geographically diverse populations," implying a broader reach, but still likely within the USA given the submission context. These were likely prospective as they involved parallel processing and assaying with two systems.

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

The ground truth in these studies is primarily established by a predicate device (Retic-COUNT™) and established laboratory methods and controls (e.g., NCCLS Document H44-A). No human experts are explicitly mentioned as establishing a "ground truth" for individual cases in the way that, for example, a radiologist would interpret an image. The performance of the reticONE™ SYSTEM is compared against the established performance of the predicate device and expected biological/analytical behavior.

4. Adjudication Method for the Test Set

Not applicable. As described above, the ground truth is primarily based on comparison to a predicate device or established analytical control materials/standards, not on human interpretation that would require adjudication.

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

No, an MRMC comparative effectiveness study was not done. The studies focused on the analytical performance of the device itself (stability, linearity, precision, carryover) and its concordance with a predicate device, not on how human readers' performance might improve with or without AI assistance. This device is an automated reticulocyte analysis system, replacing manual microscopy, rather than an AI-assisted diagnostic tool for human interpretation.

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

Yes, the studies conducted demonstrate the standalone performance of the reticONE™ SYSTEM. The system is described as combining "a reagent kit...and software for automated analysis of reticulocytes." The performance metrics reported (stability, linearity, precision, accuracy) are inherent to the automated system and its reagents/software. There is no mention of a human-in-the-loop component for the analysis process itself as part of these performance studies.

7. The Type of Ground Truth Used

The primary type of "ground truth" used is:

Predicate Device Performance: The Retic-COUNT™ system (K872166 and K880636) served as the reference for accuracy, with the reticONE™ SYSTEM demonstrating comparable results.
Established Analytical Methods/Controls: The studies refer to NCCLS Document H44-A for procedures like carryover testing. Commercial control products (e.g., Streck Laboratories, Inc® multiple-level Reticulocyte Control product, Retic-Chex) were also used.
Expected Physiological/Analytical Behavior: For linearity, serial dilutions were used to achieve a "defined range of expected reticulocytes." Stability was assessed against the expectation that measurements would not significantly change over time under specified conditions.

8. The Sample Size for the Training Set

No explicit training set is mentioned in the provided document. As a medical device that automates a laboratory assay rather than a machine learning model that needs to be "trained" on data, the concept of a training set as understood in AI/ML is not directly applicable here. The system's "training" would be more akin to software development and calibration during its design and manufacturing.

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

Not applicable, as no training set (in the AI/ML sense) is described. The system's underlying principles are based on established flow cytometry technology, fluorescent staining, and standard hematological analysis.

Summary

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4|26|99

K990327

Image /page/0/Picture/2 description: The image shows the word "COULTER" in a bold, sans-serif font. Above the word is a logo consisting of two overlapping circles with a series of triangles pointing to the right. The logo is black and white, and the word "COULTER" is in black. There is a horizontal line below the word "COULTER".

COULTER CORPORATIONP.O. BOX 169015Miami, Florida 33116-9015 USA
Customer Service: (800) 526-7694Product Information: (800) 526-6932(800) 327-6531 (305) 327-6531www.coulter.com	Date:	January 29, 1999
	Title:	Summary of Safety and Effectiveness Information for 510(k) Premarket Notification
Coulter CorporationMiami, Florida USA	Product:	reticONE™ SYSTEM for EPICS® XL Flow Cytometry Systems
Coulter Leasing CorporationMiami, Florida USA	Company:	Coulter Corporation11800 SW 147 AvenueMiami, FL 33196-2500
Coulter Electronics, Pty. Ltd.Sydney, Australia	Contact:	Dr. Marion S. Gaide (M/C: 31-B06)Senior Regulatory Affairs SpecialistPremarket Regulatory Affairs
Coulter Electronics Ind. & Com., Ltda.Rio de Janeiro, Brazil
Coulter Electronics of Canada, Ltd.Burlington, Ontario, Canada	Telephone:	305-380-2594
Coulter Electronics, Ltd.Luton, Bedfordshire, England	Common or Usual or Classification Name:Reticulocyte Analysis System with Reagents andSoftware for Flow Cytometry
Coultronics France, S.A.Margency, France	Product Classification:Product Code: GKZ; C.F.R. Section: 864.5220; ClassificationPanel: Hematology and Pathology Devices; Device Class: II
Coulter Electronics GmbHKrefeld, Germany
Coulter Electronics (HK), Ltd.Hong Kong	Intended Use:The reticONE™ SYSTEM for EPICS® XL™ Flow Cytometry Systemscombines
Coulter K. K.Tokyo, Japan	a reagent kit consisting of a Coriphosphine-O dye and a Biological Calibrator,and software for automated analysis of reticulocytes in whole blood usingEPICS® XL™ Flow Cytometry Systems with SYSTEM II™ Software. Thesystem is intended "For In Vitro Diagnostic Use" and allows identificationand enumeration of reticulocyte percentage and absolute count.
Coulter de Mexico S.A., DE C.V.Mexico City, Mexico
Coulter Electronics, Ltd.Mijdrecht, Netherlands
Coulter Electronics, Pty. Ltd.Auckland, New Zealand
Coulter Electronics Sales of PR., Inc.San Juan, Puerto Rico	Substantial Equivalence: 510(k) Premarket Notification: K872166Retic-COUNT™ (Thiazole Orange*) Reagent
Coulter Electronics, Ltd.Johannesburg, South Africa		510(k) Premarket Notification: K880636Retic-COUNT™ Reticulocyte Enumeration Software
Coulter Electronics Ltd.Istanbul, Turkey	Product Differences:The reticONE™ SYSTEM and Retic-COUNT™ are essentiallyidentical with respect to features and principles of operation. Theintended use of the New System and the Predicate System is the same.Further, the New System and the Predicate System use the same, well-established, state-of the-art technologies of cell staining with afluorescent dye-containing reagent and flow cytometric analysis tomeasure reticulocytes in whole blood [NCCLS Document H44-A].Also, each system is designed for use with specific reagents,instruments and dedicated software for a) automated instrument set-up,standardization and operation; b) sample gating and data acquisition; c)flow cytometric analysis; and d) results generation and display. Finally,both systems are alternatives to standard microscopy for reticulocyte
Coulter Electronics S.A.Caracas, Venezuela

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Product testing to assess the performance of the reticONE™ SYSTEM is described below. Studies were Product Testing: designed in line with instructions for use given in the reticONE™ SYSTEM Guide, Package Inserts, Product Manuals, and performance specifications. Specimens were assayed with Retic-COUNT™ for comparison purposes. The results of product testing demonstrate that the retic-ONE™ SYSTEM meets all performance specifications and provides reticulocyte percentage and absolute count values comparable to those of Retic-COUNT™.

1. Stability Studies:

Three studies were carried out to demonstrate whole blood specimen and sample stability claims for the reticONE™ SYSTEM.

Stored (Unstained) Whole Blood Specimens Held for 72 Hours at 2-8 ℃: a.

Normal whole blood specimens (n = 3) were collected by venipuncture into K,EDTA and ten replicate samples prepared for each specimen for each of four time points: 0-hours; 48-hours; and 72-hours; and 72-hours after collection. The 0-hours samples were stained and analyzed for reticulocyte percentage within 30 minutes of specimen collection using the reticONE™ SYSTEM and EPICS® XL-MCL™ flow cytometer. The remaining samples were stained and analyzed for reticulocyte percentage after either 24-hours; or 72-hours of storage at 2-8°C. All analyses were carried out at 20-25° using the procedures described in the reticONE™ SYSTEM Guide. The results were analyzed in terms of reticulocyte percentage and standard deviation (± 1 SD) for each donor.

Stored (Unstained) Whole Blood Specimens Held for 6 Hours at 20-25 ℃: b.

Normal whole blood specimens (n = 10) were collected by venipuncture into K,EDTA and ten replicate samples prepared for each specimen for each of two time points: 0-hours after collection. The 0hours samples were stained and analyzed for reticulocyte percentage within 30 minutes of specimen collection using the reticONE™ SYSTEM and EPICS® XL-MCL™ flow cytometer. The remaining samples were stained and analyzed for reticulocyte percentage after 6-hours of storage at 20-25°C. All analyses were carried out at 20-25° using the procedures described in the reticONE™ SYSTEM. The results were analyzed in terms of reticulocyte percentage and standard deviation (± 1 SD) for each donor.

Stored Prepared (Stained) Samples Held for 6 Hours at 20-25 ℃: C.

Normal whole blood specimens (n = 5) were collected by venipuncture into K.EDTA and ten replicate samples prepared for each specimen. The samples (0-hours) were stained and analyzed for reticulocyte percentage within 30 minutes of specimen collection using the reticONE™ SYSTEM and EPICS® XL-MCL™ flow cytometer. These same stained samples were reanalyzed for reticulocyte percentage after 6-hours of storage at 20-25°C. All analyses were carried out at 20-25° using the procedures described in the reticONE™ SYSTEM Guide. The results were analyzed in terms of reticulocyte percentage and standard deviation (± 1 SD) for each donor.

The results for the three stability studies clearly demonstrated that the reticONE™ SYSTEM meets stability claims for both stored (unstained) whole blood specimens and prepared (stained) samples under the storage times and temperature conditions studied.

2. Carryover Study:

Carryover percent was evaluated on two EPICS® XL-MCL™ flow cvtometers by the approved NCCLS procedure for high-to-low carryover testing in a reticulocyte flow cytometric analysis system [NCCLS Document H44-A]. The High Level and the Low Level of the Streck Laboratories, Inc® multiple-level Reticulocyte Control product, Retic-Chex, were used in place of a whole blood specimen with an elevated RBC count and a whole blood specimen with a low RBC count. A single sample was prepared for each Level and stained with the COULTER® reticONE™ Reagent Kit. The stained High Level samples were then analyzed for reticulocyte percentage in three successive runs (Runs: 1), I, 1, 1, using the reticONE™ SYSTEM and EPICS® XL-MCL™ flow cytometer. The stained Low Level samples were next analyzed in the same manner (Runs: J, J, J.). All sample staining and analyses were carried out at 20-25° using the procedures described in the reticONE™ SYSTEM Guide. Carryover percent was calculated using the following equation: Carryover Percent (%) == [{J ] - J x 100] == [(1} - J}). The results for the carryover study clearly demonstrated that the reticONE™ SYSTEM exhibits minimal carryover compared to the Flow Cytometer Specification.

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3. Linearity Study:

Two (n = 2) whole blood specimens were collected by venipuncture into K,EDTA and serially diluted (n = 10 data points: low, high and 8 serial dilutions in between) to achieve a defined range of 0.2% to 12.5% reticulocytes. A sample was prepared for each data point and stained with the COULTER® reticONE™ Reagent Kit. Three replicate measurements of reticulocyte percentage were then made for each stained sample using the reticONE™ SYSTEM and EPICS® XL-MCL™ flow cytometer. All sample staining and analyses were carried out at 20-25° using the procedures described in the reticONE™ SYSTEM Guide. The results were analyzed in terms of regression and correlation analyses for recovered versus expected reticulocyte percentage. The results for the linearity study clearly demonstrated linearity over the defined (reportable) range for the reticONE™ SYSTEM.

Precision Study: 4.

Three studies were carried out to demonstrate the reproducibility of the reticONE™ SYSTEM.

Within Run (Intralaboratory) Precision: a.
Measurements were made on sample [normal or abnormal] whole blood specimens for each of five levels of reticulocyte percentage using one EPICS® XL-MCL™ flow cytometer. All measurements were made in replicates of ten determinations. Results were analyzed in terms of mean reticulocyte percentage, standard deviation (± 1 SD) and coefficient of variation (%CV).
b. Interlaboratory Precision:
Measurements were made on samples from a single [normal] whole blood specimen for one level of reticulocyte percentage on the same day by three laboratories using different EPICS® XL-MCL™ flow cvtometers. All measurements were made in replicates of ten determinations. Results were analyzed in terms of mean reticulocyte percentage, standard deviation (± 1 SD) and coefficient of variation (%CV).
C. Site Precision:
Measurements were made on sample [normal or abnormal] whole blood specimens for each of five levels of reticulocyte percentage using one EPICS® XL-MCL™ flow cytometer. All measurements were made in replicates of ten determinations. Results were analyzed in terms of mean reticulocyte percentage, standard deviation (± 1 SD) and coefficient of variation (%CV).

The results for the three precision studies clearly demonstrated the reproducibility of the reticONE™ SYSTEM.

ડ. Accuracv Study:

Normal and abnormal whole blood specimens were collected from geographically diverse populations of males and females unselected as to race and ranging in age from 18 to 85 years. Specimens were divided, processed and assayed in parallel with reticONE™ SYSTEM and Retic-COUNT™. Reticulocyte percentage (%) and absolute count (cells/uL) values were determined with EPICS® XL-MCL or FACScan™ flow cytometers (as applicable). Red blood cell counts were obtained using a COULTER® STKS™ Analyzer. The results were analyzed in terms of minimum, maximum, mean, standard deviation (± 1 SD), confidence intervals with 95% limits, regression and correlation analyses, and analyses of variance. The results for the accuracy study clearly demonstrated that the reticONE™ SYSTEM and Retic-COUNT™ identify and enumerate comparable numbers of reticulocytes in whole blood specimens.

Marion S. Gaiser

Marion S. Gaide, Ph.D. Senior Regulatory Affairs Specialist Premarket Regulatory Affairs

January 29, 1999
Date

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Public Health Service

Marion S. Gaide, Ph.D. Senior Regulatory Affairs Specialist Premarket Regulatory Affairs Coulter Corporation 11800 SW 147 Avenue Miami. Florida 33196-2500

APR 2 6 1999

Food and Drug Administration 9200 Corporate Boulevard Rockville MD 20850

Re: K990327

Trade Name: reticONE™ SYSTEM for EPICS® XL™ Flow Cytometry Systems Regulatory Class: II Product Code: GKZ Dated: January 29, 1999 Received: February 2, 1999

Dear Dr. Gaide:

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 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). 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 Current Good Manufacturing Practice requirements, as set forth in the Quality System Regulation (QS) for Medical Devices: General regulation (21 CFR Part 820) and that, through periodic QS 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.

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Page 2

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.

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 (301) 443-6597, or at its internet address "http://www.fda.gov/cdrh/dsma/dsmamain.html".

Sincerely yours,

Steven Dutman

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

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INDICATIONS FOR USE

199032

510(k) Number (if known): Not Yet Assigne

reticONE™ SYSTEM for EPICS® XL™ Flow Cytometry Systems Device Name:

Indications For Use:

Peter E. Mackin

(Division Sign-Off)
Division of Clinical Laboratory Devices
510(k) Number. K970321

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

Concurrence of CDRH, Office of Device Evaluation (ODE)

1 Prescription Use (Per 21 CFR 801.109)

Over-The-Counter Use

retI4Use

Regulation Number and Section

§ 864.5220 Automated differential cell counter.

(a)
Identification. An automated differential cell counter is a device used to identify one or more of the formed elements of the blood. The device may also have the capability to flag, count, or classify immature or abnormal hematopoietic cells of the blood, bone marrow, or other body fluids. These devices may combine an electronic particle counting method, optical method, or a flow cytometric method utilizing monoclonal CD (cluster designation) markers. The device includes accessory CD markers.(b)
Classification. Class II (special controls). The special control for this device is the FDA document entitled “Class II Special Controls Guidance Document: Premarket Notifications for Automated Differential Cell Counters for Immature or Abnormal Blood Cells; Final Guidance for Industry and FDA.”