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K Number
DEN130010
Device Name
VYSIS EGR1 FISH PROBE KIT - SC (SPECIMEN CHARACTERIZATION)
Manufacturer
ABBOTT MOLECULAR, INC.
Date Cleared
2013-07-29

(111 days)

Product Code
PDO
Regulation Number
864.1870
AI/MLSaMDIVD (In Vitro Diagnostic)TherapeuticDiagnosticis PCCP Authorized
Intended Use
The Vysis EGR1 FISH Probe Kit - SC detects the LSI EGR1 probe target on chromosome 5q in bone marrow specimens. The Vysis EGR1 FISH Probe Kit -SC assay results characterize bone marrow specimens from patients with acute myeloid leukemia or myelodysplastic syndrome. The assay results are intended to be interpreted by a qualified pathologist or cytogeneticist. This device is not intended for high-risk uses such as selecting therapy, predicting therapeutic response or disease screening. The use of this product for diagnosis, monitoring or risk assessment has not been established.
Device Description
The Vysis EGR1 FISH Probe Kit - Specimen Characterization uses fluorescence in situ hybridization (FISH) DNA probe technology to detect probe target LSI EGR1 (containing early growth response 1 gene; location chromosome 5g31). The LSI D5S23, D5S721 probe (location chromosome 5p15.2) serves as a control. The Vysis EGR1 FISH Probe Kit – SC (List No. 04N37-001) consists of the following components which are sufficient to process 20 assays: 1. Vysis LSI EGR1 SpectrumOrange/D5S23, D5S721 Spectrum Green Probes 2. Vysis LSI/WCP Hybridization Buffer 3. DAPI II Counterstain 4. NP-40 5. 20X SSC Salt Items 2 through 5 above are general purposes reagents. DNA Probes: Vysis LSI EGR1 SpectrumOrange/ D5S23, D5S721 SpectrumGreen. a. The SpectrumOrange-labeled LSI EGR1 probe, approximately 209 kb in length (chr5:137654208-137862738; February 2009 Assembly; UCSC Human Genome Browser) is located at 5g31 and contains the complete EGR1 gene. b. The SpectrumGreen-labeled LSI D5S23, D5S721 probe, approximately 561 kb in length (chr5:9397109-9958407; February 2009 Assembly; UCSC Human Genome Browser) is located at 5p15.2.
More Information

No predicate device exists for this intended use

Not Found

No
The device description and performance studies focus on traditional FISH probe technology and manual interpretation by qualified personnel. There is no mention of automated image analysis, AI, or ML.

No
This device is clearly stated as "not intended for high-risk uses such as selecting therapy, predicting therapeutic response or disease screening." It is used for characterizing bone marrow specimens from patients for diagnostic purposes with results interpreted by a qualified professional.

No

The "Intended Use / Indications for Use" section explicitly states: "The use of this product for diagnosis, monitoring or risk assessment has not been established." It also clarifies that the device "characterize bone marrow specimens" and is "not intended for high-risk uses such as selecting therapy, predicting therapeutic response or disease screening."

No

The device description clearly lists physical components such as DNA probes, hybridization buffer, counterstain, NP-40, and SSC salt, indicating it is a kit containing chemical reagents, not software.

Yes, this device is an IVD (In Vitro Diagnostic).

Here's why:

  • Intended Use: The intended use explicitly states that the device "detects the LSI EGR1 probe target on chromosome 5q in bone marrow specimens" and that the "assay results characterize bone marrow specimens from patients with acute myeloid leukemia or myelodysplastic syndrome." This clearly indicates the device is used to examine specimens derived from the human body to provide information for diagnostic purposes.
  • Device Description: The device description details the use of "fluorescence in situ hybridization (FISH) DNA probe technology" to detect specific genetic targets in bone marrow specimens. This is a common technique used in IVD devices for genetic analysis.
  • Performance Studies: The document includes detailed descriptions of analytical performance studies (Precision/Reproducibility, Kit stability, Detection limit, and Analytical specificity) and references clinical validity studies. These types of studies are required for IVD devices to demonstrate their performance characteristics.
  • Intended User: The intended user is a "qualified pathologist or cytogeneticist," which are healthcare professionals who interpret laboratory results for diagnostic purposes.

While the intended use explicitly states it's "not intended for high-risk uses such as selecting therapy, predicting therapeutic response or disease screening" and that "The use of this product for diagnosis, monitoring or risk assessment has not been established," the core function of characterizing bone marrow specimens from patients with specific conditions using an in vitro assay firmly places it within the definition of an In Vitro Diagnostic device. The limitations on its use relate to the specific claims and clinical utility that have been established, not its fundamental nature as an IVD.

N/A

Intended Use / Indications for Use

The Vysis EGR1 FISH Probe Kit - SC detects the LSI EGR1 probe target on chromosome 5q in bone marrow specimens. The Vysis EGR1 FISH Probe Kit -SC assay results characterize bone marrow specimens from patients with acute myeloid leukemia or myelodysplastic syndrome. The assay results are intended to be interpreted by a qualified pathologist or cytogeneticist. This device is not intended for high-risk uses such as selecting therapy, predicting therapeutic response or disease screening. The use of this product for diagnosis, monitoring or risk assessment has not been established.

Product codes (comma separated list FDA assigned to the subject device)

PDO

Device Description

The Vysis EGR1 FISH Probe Kit - Specimen Characterization uses fluorescence in situ hybridization (FISH) DNA probe technology to detect probe target LSI EGR1 (containing early growth response 1 gene; location chromosome 5g31). The LSI D5S23, D5S721 probe (location chromosome 5p15.2) serves as a control.

The Vysis EGR1 FISH Probe Kit – SC (List No. 04N37-001) consists of the following components which are sufficient to process 20 assays:

    1. Vysis LSI EGR1 SpectrumOrange/D5S23, D5S721 Spectrum Green Probes
    1. Vysis LSI/WCP Hybridization Buffer
    1. DAPI II Counterstain
    1. NP-40
    1. 20X SSC Salt

Items 2 through 5 above are general purposes reagents.

DNA Probes: Vysis LSI EGR1 SpectrumOrange/ D5S23, D5S721 SpectrumGreen.

  • The SpectrumOrange-labeled LSI EGR1 probe, approximately 209 kb in a. length (chr5:137654208-137862738; February 2009 Assembly; UCSC Human Genome Browser) is located at 5g31 and contains the complete EGR1 gene.
  • b. The SpectrumGreen-labeled LSI D5S23, D5S721 probe, approximately 561 kb in length (chr5:9397109-9958407; February 2009 Assembly; UCSC Human Genome Browser) is located at 5p15.2.

Mentions image processing

Not Found

Mentions AI, DNN, or ML

Not Found

Input Imaging Modality

Not Found

Anatomical Site

bone marrow

Indicated Patient Age Range

Not Found

Intended User / Care Setting

qualified pathologist or cytogeneticist

Description of the training set, sample size, data source, and annotation protocol

Not Found

Description of the test set, sample size, data source, and annotation protocol

Not Found

Summary of Performance Studies (study type, sample size, AUC, MRMC, standalone performance, key results)

  1. Analytical performance:
    a. Precision/Reproducibility: Repeatability and reproducibility of the Vysis EGR1 FISH Probe Kit were tested.
    Study: Precision: Intra-Day and Inter-Day
    Study Protocol:
    - 6 Bone marrow specimens
    - 2 high positive (HP1 & 2) (69.3%, 49.9% positivity)
    - 2 low positive (LP1 & 2) (12% positivity)
    - 2 negative (Neg1 & 2)
    - 3 test sites
    - 5 different testing days
    - 1 lot of probe
    - Red and green signal patterns of 200 nuclei evaluated by 2 technologists (each technologist evaluated 100 nuclei per panel member)
    - Pre-specified acceptance criteria: Agreement with the expected result of greater than or equal to 90% for the high positive specimen category for each site and 90% for the negative specimen category across all sites with no more than 3 discordant results occurring at one site
    Conclusion: Acceptance criteria met

     Study: Reproducibility: Inter-Site
 Study Protocol:
 - Testing configuration same as above
 - In addition, 2 replicates of each panel member were tested
 - Pre-specified acceptance criteria: Same as above
 Conclusion: Acceptance criteria met

 Study: Lot to Lot Reproducibility
 Study Protocol:
 - 6 Bone marrow specimens
 	- 2 high positive (HP1 & 2) (69.3%, 49.9% positivity)
 	- 2 low positive (LP1 & 2) (12% positivity)
 	- 2 negative (Neg1 & 2)
 - 3 lots of probe
 - 4 replicates
 - 1 site
 - 2 operators per site
 - Pre-specified acceptance criteria: Same as above
 Conclusion: Acceptance criteria met

 Between Site Analysis of Variance:
 - High Positive 1 (N=30, Mean %=70.0, SD=5.44)
 - High Positive 2 (N=30, Mean %=47.6, SD=0.74)
 - Low Positive 1 (N=30, Mean %=18.1, SD=1.03)
 - Low Positive 2 (N=30, Mean %=14.9, SD=0.00)
 - Negative 1 (N=30, Mean %=0.7, SD=0.99)
 - Negative 2 (N=30, Mean %=0.9, SD=1.5)

 Within-day/Between-day Analysis of Variance:
 - High Positive 1 (N=30, Mean %=70.0, Within-Day Component SD=3.28, Between-Day Component SD=4.01)
 - High Positive 2 (N=30, Mean %=47.6, Within-Day Component SD=5.56, Between-Day Component SD=0.00)
 - Low Positive 1 (N=30, Mean %=18.1, Within-Day Component SD=3.00, Between-Day Component SD=3.82)
 - Low Positive 2 (N=30, Mean %=14.9, Within-Day Component SD=3.25, Between-Day Component SD=1.54)
 - Negative 1 (N=30, Mean %=0.7, Within-Day Component SD=0.71, Between-Day Component SD=0.00)
 - Negative 2 (N=30, Mean %=0.9, Within-Day Component SD=0.66, Between-Day Component SD=1.42)

 Lot-to-Lot Analysis of Variance:
 - High Positive 1 (N=12, Mean %=66.2, SD=0.00)
 - High Positive 2 (N=12, Mean %=47.4, SD=3.04)
 - Low Positive 1 (N=12, Mean %=12.7, SD=0.00)
 - Low Positive 2 (N=12, Mean %=12.3, SD=1.12)
 - Negative 1 (N=12, Mean %=0.0, SD=0.00)
 - Negative 2 (N=12, Mean %=0.1, SD=0.20)

    c. Kit stability:
    Study: Real-Time Stability
    Study Protocol: Attributes evaluated: signal intensity, target background, cross-hybridization, specificity, overall readability; 3 lots of the device; 3 specimens; The Vysis EGRI FISH Probe Kit dating is determined by the component with the shortest expiration dating. Pre-specified acceptance criteria: Acceptable quality of all attributes for all samples tested
    Conclusion: Acceptance criteria met for 12 month stability

     Study: In-Use Freeze-Thaw Stability
 Study Protocol: A series of 20 freeze-thaw cycles was performed on the probes, hybridization buffer and DAPI II counterstain. 1 lot of the device; 3 specimens; Attributes evaluated: Same as above; Pre-specified acceptance criteria: Same as above
 Conclusion: Acceptance criteria met throughout and at the end of 20 freeze-thaw cycles

 Study: Transport and Temperature Extreme Stability
 Study Protocol: Device components were removed from -20°C and cycled for 48 hours on dry ice, 72 hours at 25°C, 72 hours at 40°C, and -20°C for 24 hours prior to testing; 1 lot of the device; 3 specimens; Attributes evaluated: Same as above; Pre-specified acceptance criteria: Same as above
 Conclusion: Acceptance criteria met

 Study: Post-hybridization Signal Stability
 Study Protocol: A single slide from each of 3 bone marrow specimens was tested at baseline and then stored at -20°C ± 10°C protected from light; The samples were tested at three time points -Day 7, Day 14, and Day 25; 1 lot of the device; Attributes evaluated: Same as above; Pre-specified acceptance criteria: Same as above
 Conclusion: Acceptance criteria met for Post-hybridization stability of 3 weeks

 Study: Probe Photostability
 Study Protocol: The device was exposed continuously under white fluorescent light (to mimic standard laboratory conditions) at 15-30°C for different time lengths (0 hours, 3 hours, 8 hours, 24 hours and 48 hours); 1 lot of the device; Attributes evaluated: Same as above; Pre-specified acceptance criteria: Same as above
 Conclusion: Acceptance criteria met for Photo-stability of 48 hours

    d. Detection limit:
    The analytical sensitivity of the Vysis LSI EGRI SpectrumOrange/D5S23, D5S721SpectrumGreen probes was established using interphase nuclei prepared from 25 bone marrow specimens that were either karyotypically normal or 5pl5 and 5q31 deletion-free. The orange and green signal patterns of nuclei for these 25 specimens were evaluated by two technologists. Each technologist evaluated 100 nuclei per specimen for a total of 200 nuclei per specimen and 5000 scoreable nuclei from normal specimens. The analytical sensitivity was calculated as the percentage of scoreable interphase nuclei with the expected 2 red/2 green signal pattern.
    Probe: Vysis EGR1/D5S23, D5S721; Total Number of Nuclei Scored: 5000; Number of Nuclei With Expected Signal Pattern: 4979; Analytical Sensitivity (95% Confidence Interval): 99.6% (99.4, 99.7)

    e. Analytical specificity:
    The analytical specificity of the Vysis LSI EGR1 SpectrumOrange/D5S23, D5S721 SpectrumGreen probes for their respective chromosome target loci was established using metaphase chromosomes prepared from peripheral blood cultures of five karyotypically normal males that were pooled prior to dropping on microscope slides. The hybridization location of each FISH signal on chromosomes of 100 consecutive metaphase nuclei was evaluated by one technologist for a total of 200 target loci.
    The analytical specificity of each probe was calculated as the number of metaphase chromosome FISH signals hybridized to the correct locus divided by the total number of metaphase chromosome FISH signals hybridized and multiplied by 100 to give a percentage. The analytical specificity of the Vysis LSI EGR1 SpectrumOrange/D5S23, D5S721 SpectrumGreen Probes was 100%.
    Probe: D5S23, D5S721; Target: 5p15.2; Total No. of Metaphase Chromosome Hybridized: 200; No. of Metaphase Chromosome Hybridized to Correct Locus: 200; Analytical Specificity (95% Confidence Intervals): 100% (98, 100)
    Probe: EGR 1; Target: 5q31; Total No. of Metaphase Chromosome Hybridized: 200; No. of Metaphase Chromosome Hybridized to Correct Locus: 200; Analytical Specificity (95% Confidence Intervals): 100% (98, 100)

  2. Clinical studies:
    c. Other clinical supportive data: The sponsor provided three peer-reviewed published papers to support the clinical validity of the device in characterizing bone marrow specimens from patients with AML. All three papers used the Abbott Vysis EGR1 FISH Probe Kit.

    Data Source 1 (Sun et. al):

    • Was the specific device under review in the submission used in the study?: Yes
    • Was the specimen type in the study representative of the claimed specimen type(s)?: Yes
    • Target population (disease status): Known or suspected del(5q) having MDS or AML
    • Upper reference limit (percentage and per 200 nuclei): 6% or 12 1R2G patterns per 200 scoreable interphase nuclei
    • Total Number of specimens tested for each claimed type: 320 bone marrow specimens
    • Number of specimens with a positive probe result [5q- (1R2G)]: 66
    • Range of positive probe results: Not available

    Data Source 2 (Galvan et. al):

    • Was the specific device under review in the submission used in the study?: Yes
    • Was the specimen type in the study representative of the claimed specimen type(s)?: Yes
    • Target population (disease status): MDS, therapy-related MDS (t-MDS) and AML patients
    • Upper reference limit (percentage and per 200 nuclei): 6% or 12 1R2G patterns per 200 scoreable interphase nuclei
    • Total Number of specimens tested for each claimed type: 28 bone marrow specimens
    • Number of specimens with a positive probe result [5q- (1R2G)]: 23 (MDS - 6, t-MDS - 3, t-MDS&MM – 1, AML - 13)
    • Range of positive probe results: MDS: 35 - 81.5%, t-MDS: 25 - 75%, t-MDS&MM: 76%, AML: 20 - 99%

    Data Source 3 (Vance et. al):

    • Was the specific device under review in the submission used in the study?: Yes
    • Was the specimen type in the study representative of the claimed specimen type(s)?: Yes
    • Target population (disease status): AML
    • Upper reference limit (percentage and per 200 nuclei): 6% or 12 1R2G patterns per 200 scoreable interphase nuclei
    • Total Number of specimens tested for each claimed type: 181 bone marrow specimens* (* Line data unpublished, but this data was provided in the 510(k) submission)
    • Number of specimens with a positive probe result [5q- (1R2G)]: 8 (AML - 8)
    • Range of positive probe results: 45 - 91 %

Key Metrics (Sensitivity, Specificity, PPV, NPV, etc.)

Analytical Sensitivity: 99.6% (95% Confidence Interval: 99.4, 99.7)
Analytical Specificity: 100% (95% Confidence Intervals: 98, 100) for both D5S23, D5S721 and EGR 1 probes.

Predicate Device(s): If the device was cleared using the 510(k) pathway, identify the Predicate Device(s) K/DEN number used to claim substantial equivalence and list them here in a comma separated list exactly as they appear in the text. List the primary predicate first in the list.

No predicate device exists for this intended use

Reference Device(s): Identify the Reference Device(s) K/DEN number and list them here in a comma separated list exactly as they appear in the text.

Not Found

Predetermined Change Control Plan (PCCP) - All Relevant Information for the subject device only (e.g. presence / absence, what scope was granted / cleared under the PCCP, any restrictions, etc).

Not Found

§ 864.1870 Early growth response 1 (EGR1) gene fluorescence in-situ hybridization (FISH) test system for specimen characterization.

(a)
Identification. An early growth response 1 (EGR1) gene fluorescence in-situ hybridization (FISH) test system for specimen characterization is a device intended to detect the EGR1 probe target on chromosome 5q in bone marrow specimens from patients with acute myeloid leukemia (AML) or myelodysplastic syndrome (MDS). The assay results are intended to be interpreted only by a qualified pathologist or cytogeneticist. These devices do not include automated systems that directly report results without review and interpretation by a qualified pathologist or cytogeneticist. These devices also do not include any device intended for use to select patient therapy, predict patient response to therapy, or to screen for disease as well as any device with a claim for a particular diagnosis, prognosis, monitoring, or risk assessment.(b)
Classification. Class II (special controls). The special controls for this device are:(1) Premarket notification submissions must also include the following information:
(i) A detailed description of all probes included in the kit;
(ii) Purpose of each probe;
(iii) Probe molecular specificity;
(iv) Probe specificity;
(v) Probe limits;
(vi) Probe sensitivity;
(vii) Specification of required ancillary reagents, instrumentation, and equipment;
(viii) Specification of the specimen collection, processing, storage and slide preparation methods;
(ix) Specification of the assay procedure;
(x) Specification of control elements that are incorporated into the recommended testing procedures;
(xi) Specification of risk mitigation elements: Description of all additional procedures, methods, and practices incorporated into the directions for use that mitigate risks associated with testing;
(xii) Specification of the criteria for test result interpretation and reporting;
(xiii) Device analytical sensitivity data;
(xiv) Device analytical specificity data;
(xv) Device reference limit data;
(xvi) Device precision/reproducibility data;
(xvii) Device stability data to include:
(A) Real-time stability,
(B) Freeze-thaw stability,
(C) Transport and temperature stability,
(D) Post-hybridization signal stability,
(E) Photostability of probe, and
(xviii) Documentation that demonstrates the clinical validity of the device. The documentation must include data from clinical studies, a minimum of two peer-reviewed published literature references using the specific device seeking marketing clearance, or both. Documentation for the clinical studies and peer-reviewed published literature references cited must include the following elements:
(A) Documentation that the sponsor's probe was used in the literature reference,
(B) Number and type of specimens,
(C) Target population studied,
(D) Upper reference limit, and
(E) Range of positive probe results.
(2) Your § 809.10(b)(12) of this chapter compliant labeling must include a statement summarizing the data identified in paragraphs (b)(1)(xiii) through (xviii) of this section and a description of the studies supporting the information, including the pre-specified acceptance criteria for these performance studies, justification for the pre-specified acceptance criteria, and whether the pre-specified acceptance criteria were met.
(3) Your § 809.10 of this chapter compliant labeling must include:
(i) A warning that reads “The assay results are intended to be interpreted only by a qualified pathologist or cytogeneticist.”
(ii) A warning that reads “This device is not for high-risk uses such as selecting therapy, predicting therapeutic response or disease screening.”
(iii) A warning that reads “The use of this device for diagnosis, monitoring or risk assessment has not been established.”

0

EVALUATION OF AUTOMATIC CLASS III DESIGNATION FOR VYSIS EGR1 FISH PROBE KIT - SC (SPECIMEN CHARACTERIZATION)

Decision Summary

A. 510(k) NumberK123951
B. Purpose of the Submission:Clearance of new assay
C. Measurand:LSI EGR1 probe target on chromosome 5q in
bone marrow specimens
D. Type of Test:Flourescence in-situ hybridization (FISH)
E. Applicant:Abbott Molecular Inc.
F. Device Name:Vysis EGR1 FISH Probe Kit - SC (Specimen
Characterization)

G. Regulatory Information:

FDA identifies this type of device as:

An early growth response 1 (EGR1) gene fluorescence in-situ hybridization (FISH) test system for specimen characterization is a device intended to detect the EGR1 probe target on chromosome 5q in bone marrow specimens from patients with acute myeloid leukemia (AML) or myelodysplastic syndrome (MDS). The assay results are intended to be interpreted only by a qualified pathologist or cytogeneticist. These devices do not include automated systems that directly report results without review and interpretation by a qualified pathologist or cytogeneticist. These devices also do not include any device intended for use to select patient therapy, predict patient response to therapy or to screen for disease as well as any device with a claim for a particular diagnosis, prognosis, monitoring or risk assessment.

1. New Regulation Number:21 CFR 864.1870
2. Classification:Class II (special controls)
3. Product code:PDO
4. Panel:Hematology and Pathology Devices Panel

H. Intended Use:

1

1. Intended use(s):

The Vysis EGR1 FISH Probe Kit - SC detects the LSI EGR1 probe target on chromosome 5q in bone marrow specimens. The Vysis EGR1 FISH Probe Kit -SC assay results characterize bone marrow specimens from patients with acute myeloid leukemia or myelodysplastic syndrome. The assay results are intended to be interpreted by a qualified pathologist or cytogeneticist. This device is not intended for high-risk uses such as selecting therapy, predicting therapeutic response or disease screening. The use of this product for diagnosis, monitoring or risk assessment has not been established.

2. Indication(s) for use:

Same as intended use

    1. Special conditions for use statement(s):
      Warnings included in the labeling are as follows:

  • The assay results are intended to be interpreted only by a qualified pathologist or cytogeneticist.

  • · The Vysis EGR1 FISH Probe Kit SC is not for high-risk uses such as selecting therapy, predicting therapeutic response or disease screening.

  • · The use of this product for diagnosis, monitoring or risk assessment has not been established.

  • · Caution: Federal law restricts this device to sale by or on the order of a physician or other practitioner licensed by the law of the State in which he practices, to use or order the use of the device.

    1. Special instrument requirements:

Fluorescence microscope equipped with appropriate excitation and emission filters

I. Device Description:

The Vysis EGR1 FISH Probe Kit - Specimen Characterization uses fluorescence in situ hybridization (FISH) DNA probe technology to detect probe target LSI EGR1 (containing early growth response 1 gene; location chromosome 5g31). The LSI D5S23, D5S721 probe (location chromosome 5p15.2) serves as a control.

The Vysis EGR1 FISH Probe Kit – SC (List No. 04N37-001) consists of the following components which are sufficient to process 20 assays:

    1. Vysis LSI EGR1 SpectrumOrange/D5S23, D5S721 Spectrum Green Probes
    1. Vysis LSI/WCP Hybridization Buffer
    1. DAPI II Counterstain
    1. NP-40

2

5. 20X SSC Salt

Items 2 through 5 above are general purposes reagents.

DNA Probes: Vysis LSI EGR1 SpectrumOrange/ D5S23, D5S721 SpectrumGreen.

  • The SpectrumOrange-labeled LSI EGR1 probe, approximately 209 kb in a. length (chr5:137654208-137862738; February 2009 Assembly; UCSC Human Genome Browser) is located at 5g31 and contains the complete EGR1 gene.
  • b. The SpectrumGreen-labeled LSI D5S23, D5S721 probe, approximately 561 kb in length (chr5:9397109-9958407; February 2009 Assembly; UCSC Human Genome Browser) is located at 5p15.2.

J. Substantial Equivalence Information:

    1. Predicate device name(s):
      No predicate device exists for this intended use
    1. Predicate 510(k) number(s):
      Not applicable
    1. Comparison with predicate:
      Not Applicable

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

Not Applicable

L. Test Principle:

Alterations in chromosome 5 are common aberrations in myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML). Chromosome band 5q31 contains the early growth response 1 gene (EGR1). The Vysis locus specific identifier (LSI) EGR1 SpectrumOrange Probe and D5S23, D5S721 SpectrumGreen Probe are components of the Vysis EGR1 FISH Probe Kit, and are used to detect the target EGR 1 using FISH DNA probe technology. The results of this assay are used to characterize bone marrow specimens.

Bone marrow cells from AML patients attached to microscope slides using standard cytogenetic procedures are used for this assay. The resulting cellular DNA is denatured to single-stranded form and subsequently allowed to hybridize with the LSI EGR1 and LSI D5S23, D5S721 probes. Following hybridization, the unbound probe is removed by a series of washes, and the nuclei are counterstained with DAPI, a DNA-specific stain that fluoresces blue. Hybridization of the LSI EGR1 and LSI D5S23, D5S721 probes is viewed using a fluorescence microscope equipped with

3

appropriate excitation and emission filters, allowing visualization of the orange and green fluorescent signals.

The expected signal pattern in a cell with two copies of the EGR1 gene covered by the probes using the Vysis EGR1 FISH Probe Kit is 2 orange and 2 green signals (2R2G). In a cell with the 5q alteration, one orange signal (LSI EGR1) and two green signals (LSI D5S23, D5S721) will be expected. Enumeration of the orange LSI EGR 1 and green LSI D5S23, D5S721 signals provide a mechanism for determining absolute copy number of the probe targets and the presence of chromosomal alterations of interest.

M. Performance Characteristics (if/when applicable):

The sponsor provided the following information to support the analytical performance of the device:

    1. Analytical performance:
    • a. Precision/Reproducibility: Repeatability and reproducibility of the Vysis EGR1 FISH Probe Kit were tested as shown in the table below:

4

StudyStudy ProtocolConclusion
Precision:
Intra-Day and
Inter-Day• 6 Bone marrow specimens
  • 2 high positive (HP1 & 2)
    (69.3%, 49.9% positivity)
  • 2 low positive (LP1 & 2)
    (12% positivity)
  • 2 negative (Neg1 & 2)

• 3 test sites
• 5 different testing days
• 1 lot of probe

• Red and green signal
patterns of 200 nuclei
evaluated by 2
technologists (each
technologist evaluated 100
nuclei per panel member)

• Pre-specified acceptance criteria:
Agreement with the expected result
of greater than or equal to 90% for
the high positive specimen
category for each site and 90% for
the negative specimen category
across all sites with no more than 3
discordant results occurring at one
site | Acceptance criteria met |

5

StudyStudy ProtocolConclusion
Reproducibility:
Inter-Site• Testing configuration same
as above
• In addition, 2 replicates of
each panel member were
tested
• Pre-specified acceptance
criteria: Same as aboveAcceptance criteria met
Lot to Lot
Reproducibility• 6 Bone marrow specimens
  • 2 high positive (HP1 & 2)
    (69.3%, 49.9% positivity)
  • 2 low positive (LP1 & 2)
    (12% positivity)
  • 2 negative (Neg1 & 2)
    • 3 lots of probe
    • 4 replicates
    • 1 site
    • 2 operators per site
    • Pre-specified acceptance
    criteria: Same as above | Acceptance criteria met |

Between Site Analysis of Variance

Sample TypeNMean %aSDb
High Positive 13070.05.44
High Positive 23047.60.74
Low Positive 13018.11.03
Low Positive 23014.90.00
Negative 1300.70.99
Negative 2300.91.5

ª Percentage of cells with 1R2G signal pattern

bSD = standard deviation

| Sample Type | N | Mean %a | Within-Day
Component
SDb | Between-Day
Component
SD |
|-----------------|----|---------|--------------------------------|--------------------------------|
| High Positive 1 | 30 | 70.0 | 3.28 | 4.01 |
| High Positive 2 | 30 | 47.6 | 5.56 | 0.00 |
| Low Positive 1 | 30 | 18.1 | 3.00 | 3.82 |
| Low Positive 2 | 30 | 14.9 | 3.25 | 1.54 |
| Negative 1 | 30 | 0.7 | 0.71 | 0.00 |
| Negative 2 | 30 | 0.9 | 0.66 | 1.42 |

Within-day/Between-day Analysis of Variance

ª Percentage of cells with 1R2G signal pattern

bSD = standard deviation

6

Sample TypeNMean %aSDb
High Positive 11266.20.00
High Positive 21247.43.04
Low Positive 11212.70.00
Low Positive 21212.31.12
Negative 1120.00.00
Negative 2120.10.20

Lot-to-Lot Analysis of Variance

ª Percentage of cells with 1R2G signal pattern

bSD = standard deviation

  • b. Linearity/assay reportable range:
    Not applicable

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

Kit stability

The kit stability was assessed as detailed in the table below:

StudyStudyProtocolConclusion
Real-Time
Stability• Attributes evaluated: signal
intensity, target
background, cross-hybridization,
specificity, overall readability
• 3 lots of the device
• 3 specimens
• The Vysis EGRI FISH Probe Kit
dating is determined by the
component with the shortest
expiration dating.
• Pre-specified acceptance criteria:
Acceptable quality of all attributes
for all samples testedAcceptance criteria met for
12 month stability
In-Use Freeze-
Thaw Stability• A series of 20 freeze-thaw cycles
was performed on the probes,
hybridization buffer and DAPI II
counterstain.
• 1 lot of the device
• 3 specimens
• Attributes evaluated: Same
as above
• Pre-specified acceptance criteria:
Same as aboveAcceptance criteria met
throughout and at the end
of 20 freeze-thaw cycles
Transport and
Temperature
Extreme• Device components were removed
from -20°C and cycled for 48 hours
on dry ice, 72 hours at 25°C, 72Acceptance criteria met

7

| Stability | hours at 40°C, and -20°C for 24
hours prior to testing
• 1 lot of the device
• 3 specimens
• Attributes evaluated: Same as
above
• Pre-specified acceptance criteria:
Same as above | |
|--------------------------------------------|-------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|---------------------------------------------------------------------------|
| Post-
hybridization
Signal Stability | • A single slide from each of 3 bone
marrow specimens was tested at
baseline and then stored at -20°C ±
10°C protected from light
• The samples were tested at three
time points -Day 7, Day 14, and
Day 25
• 1 lot of the device
• Attributes evaluated: Same
as above
• Pre-specified acceptance criteria:
Same as above | Acceptance criteria met for
Post-hybridization stability
of 3 weeks |
| Probe
Photostability | • The device was exposed
continuously under white
fluorescent light (to mimic
standard laboratory conditions) at
15-30°C for different time lengths
(0 hours, 3 hours, 8 hours, 24 hours
and 48 hours)
• 1 lot of the device
• Attributes evaluated: Same as
above
• Pre-specified acceptance criteria:
Same as above | Acceptance criteria met for
Photo-stability of 48 hours |

Controls:

The Vysis EGR1 FISH Probe Kit also contains the LSI D5S23, D5S721 probe (location chromosome 5p15.2) which serves as a control. Observation of at least 1R signal and 1G signal, assures that the assay conditions are adequate to allow both the orange and green labeled probes to bind properly to their respective targets.

  • d. Detection limit:
    The analytical sensitivity of the Vysis LSI EGRI SpectrumOrange/D5S23, D5S721SpectrumGreen probes was established using interphase nuclei prepared from 25 bone marrow specimens that were either karyotypically normal or 5pl5 and 5q31 deletion-free. The orange and green signal patterns of nuclei for these 25 specimens were evaluated by two technologists. Each

8

technologist evaluated 100 nuclei per specimen for a total of 200 nuclei per specimen and 5000 scoreable nuclei from normal specimens. The analytical sensitivity was calculated as the percentage of scoreable interphase nuclei with the expected 2 red/2 green signal pattern.

| Probe | Total Number
of Nuclei
Scored | Number of Nuclei
With Expected
Signal Pattern | Analytical Sensitivity
(95% Confidence Interval) |
|--------------------------------|-------------------------------------|-----------------------------------------------------|-----------------------------------------------------|
| Vysis
EGR1/D5S23,
D5S721 | 5000 | 4979 | 99.6% (99.4, 99.7) |

e. Analytical specificity:

Analytical specificity is defined as the percentage of signals that hybridize to the correct locus and no other location. The analytical specificity of the Vysis LSI EGR1 SpectrumOrange/D5S23, D5S721 SpectrumGreen probes for their respective chromosome target loci was established using metaphase chromosomes prepared from peripheral blood cultures of five karyotypically normal males that were pooled prior to dropping on microscope slides. The hybridization location of each FISH signal on chromosomes of 100 consecutive metaphase nuclei was evaluated by one technologist for a total of 200 target loci.

For each probe and sample, the number of metaphase chromosome FISH signals hybridized to the correct locus and the number of metaphase chromosome FISH signals hybridized to the incorrect locus were enumerated.

The analytical specificity of each probe was calculated as the number of metaphase chromosome FISH signals hybridized to the correct locus divided by the total number of metaphase chromosome FISH signals hybridized and multiplied by 100 to give a percentage. The analytical specificity of the Vysis LSI EGR1 SpectrumOrange/D5S23, D5S721 SpectrumGreen Probes was 100%.

| Probe | Target | Total No. of
Metaphase
Chromosome
Hybridized | No. of
Metaphase
Chromosome
Hybridized to
Correct Locus | Analytical Specificity
(95% Confidence
Intervals) |
|---------------|--------|-------------------------------------------------------|---------------------------------------------------------------------|---------------------------------------------------------|
| D5S23, D5S721 | 5p15.2 | 200 | 200 | 100%
(98, 100) |
| EGR 1 | 5q31 | 200 | 200 | 100%
(98, 100) |

f. Assay cut-off:

Same as reference range (see #5 below)

9

2. Comparison studies:

  • a. Method comparison with predicate device:
    Not applicable

  • b. Matrix comparison:
    Not applicable

    1. Clinical studies:
    • a. Clinical Sensitivity:

Not applicable

  • b. Clinical specificity:
    Not applicable

  • c. Other clinical supportive data (when a. and b. are not applicable):
    The sponsor provided three peer-reviewed published papers to support the clinical validity of the device in characterizing bone marrow specimens from patients with AML. All three papers used the Abbott Vysis EGR1 FISH Probe Kit. The information is listed in the table below:

10

| Conditions | Data Source 1
Sun et. al | Data Source 2
Galvan et. al | Data Source 3
Vance et. al |
|-----------------------------------------------------------------------------------------------|---------------------------------------------------------------------|----------------------------------------------------------------------|---------------------------------------------------------------------|
| Was the specific
device under
review in the
submission used in
the study? | Yes | Yes | Yes |
| Was the specimen
type in the study
representative of
the claimed
specimen type(s) | Yes | Yes | Yes |
| Target population
(disease status) | Known or
suspected del(5q)
having MDS or
AML | MDS, therapy-
related MDS (t-MDS) and AML
patients | AML |
| Upper reference
limit (percentage
and per 200 nuclei) | 6% or 12 1R2G
patterns per 200
scoreable
interphase nuclei | 6% or 12 1R2G
patterns per 200
scoreable
interphase nuclei | 6% or 12 1R2G
patterns per 200
scoreable
interphase nuclei |
| Total Number of
specimens tested
for each claimed
type | 320 bone marrow
specimens | 28 bone marrow
specimens | 181 bone marrow
specimens* |
| Number of
specimens with a
positive probe
result [5q- (1R2G)] | 66 | 23
MDS - 6
t-MDS - 3
t-MDS&MM – 1
AML - 13 | 8
AML - 8 |
| Range of positive
probe results | Not available | MDS: 35 - 81.5%
t-MDS: 25 - 75%
t-MDS&MM: 76%
AML: 20 - 99% | 45 - 91 % |

MDS – Myelodysplastic syndrome

AML – Acute myeloid leukemia

t-MDS – Therapy related MDS

MM – Multiple Myeloma

  • Line data unpublished, but this data was provided in the 510(k) submission

Sun Y, Cook JR. Fluorescence in situ hybridization for del(5q) in myelodysplasia/acute myeloid lukemia: Comparison of EGR1 vs. CSF1R probes and diagnostic yield over metaphase cytogenetics alone. Leuk Res 2010; 34: 340-343.

Galvan AB, Mallo M, Arenillas L, et al. Does monosomy 5 really exist in myelodysplastic syndromes and acute myeloid leukemia? Leuk Res 2010; 34: 1242– 1245.

Vance GH, Haesook K, Hicks GA, et al. Utility of interphase FISH to stratify patients into cytogenetic

11

risk categories at diagnosis of AML in an Eastern Cooperative Oncology Group (ECOG) clinical trial (E1900). Leuk Res 2007; 31:605-609.

    1. Clinical cut-off:
      Not applicable
    1. Expected values/Reference range:
      The upper reference limit is defined as the maximum quantity of scoreable interphase nuclei with an altered signal pattern from either karyotypically normal specimens or 5p15.2 and 5g31 deletion-free specimens. The upper reference limit is expressed in terms of a percentage or the actual number of atypical nuclear FISH signal pattern per the standard number of nuclei tested.

The upper reference limit for this assay is 6% or 12 1R2G patterns per 200 scoreable interphase nuclei. Published literature was used to establish the reference range for this assay.

In order to validate the 6% upper reference limit of the Vysis EGR 1 FISH Probe Kit, the assay was performed on interphase nuclei from 25 bone marrow specimens from either karyotypically normal specimens or 5p15.2 and 5q31 deletion-free specimens. The signal patterns of 200 nuclei were evaluated by counting the number of orange and green signals. Each of two technologists evaluated 100 nuclei per specimen. Among the 25 normal specimens, none produced 1R2G signals at or above the 6% upper reference limit.

N. Other Supportive Instrument Characteristics Data Not Covered In The "Performance Characteristics" Section above:

None

O. Proposed Labeling:

Labeling satisfies the requirements of 21 CFR 809.10, 21 CFR 801.109, including an appropriate prescription statement as required by 21 CFR 801.109(b), and the special controls for this type of device.

Identified Potential RiskRequired Mitigations
False negative result1) Premarket notification submissions must
also include the following information:
i) A detailed description of all probes
included in the kit
ii) Purpose of each probe
iii) Probe molecular specificity
iv) Probe specificity
v) Probe limits
vi)Probe sensitivity
vii)Specification of required ancillary
reagents, instrumentation and
equipment
viii)Specification of the specimen
collection, processing, storage and
slide preparation methods
ix)Specification of the assay procedure
x)Specification of control elements
that are incorporated into the
recommended testing procedures
xi)Specification of risk mitigation
elements: description of all
additional procedures, methods, and
practices incorporated into the
directions for use that mitigate risks
associated with testing
xii)Specification of the criteria for test
result interpretation and reporting
xiii)Device analytical sensitivity data
xiv)Device analytical specificity data
xv)Device reference limit data
xvi)Device precision/reproducibility
data
xvii)Device stability data to include:
A) Real-time Stability
B) Freeze-Thaw Stability
C) Transport and Temperature
Stability
D)Post-Hybridization Signal
Stability
E) Photostability of Probe
xviii)Documentation that demonstrates the
clinical validity of the device. The
documentation must include data
from clinical studies, a minimum of
two peer-reviewed published
literature references using the
specific device seeking marketing
clearance, or both. Documentation
for the clinical studies and peer-
reviewed published literature
references cited must include the
following elements:
A) Documentation that the
literature reference
B) Number & type of specimens
C) Target population studied
D) Upper reference limit
E) Range of positive probe results
2) Your 21 CFR 809.10(b)(12) compliant labeling must include a statement summarizing the data identified in subparagraphs (1)(xiii)-(xviii) and a description of the studies supporting the information, including the pre-specified acceptance criteria for these performance studies, justification for the pre-specified acceptance criteria, and whether the pre-specified acceptance criteria were met.
3) Your 809.10 compliant labeling must include:
i) A warning that reads “The assay results are intended to be interpreted only by a qualified pathologist or cytogeneticist.”
ii) A warning that reads “This device is not for high-risk uses such as selecting therapy, predicting therapeutic response or disease screening.”
iii) A warning that reads “The use of this device for diagnosis, monitoring or risk assessment has not been established.”
False Positive Result1) Premarket notification submissions must also include the following information:
i) A detailed description of all probes included in the kit
ii) Purpose of each probe
iii) Probe molecular specificity
iv) Probe specificity
v) Probe limits
vi) Probe sensitivity
vii) Specification of required ancillary reagents, instrumentation and equipment
viii) Specification of the specimen collection, processing, storage and slide preparation methods
ix)Specification of the assay procedure
x)Specification of control elements that are incorporated into the recommended testing procedures
xi)Specification of risk mitigation elements: description of all additional procedures, methods, and practices incorporated into the directions for use that mitigate risks associated with testing
xii)Specification of the criteria for test result interpretation and reporting
xiii)Device analytical sensitivity data
xiv)Device analytical specificity data
xv)Device reference limit data
xvi)Device precision/reproducibility data
xvii)Device stability data to include:
A) Real-time Stability
B) Freeze-Thaw Stability
C) Transport and Temperature Stability
D) Post-Hybridization Signal Stability
E) Photostability of Probe
xviii)Documentation that demonstrates the clinical validity of the device. The documentation must include data from clinical studies, a minimum of two peer-reviewed published literature references using the specific device seeking marketing clearance, or both. Documentation for the clinical studies and peer-reviewed published literature references cited must include the following elements:
A) Documentation that the sponsor's probe was used in the literature reference
B) Number & type of specimens
C) Target population studied
D) Upper reference limit
E) Range of positive probe results
2)Your 21 CFR 809.10(b)(12) compliant labeling must include a statement summarizing the data identified in
description of the studies supporting the information, including the pre-specified acceptance criteria for these performance studies, justification for the pre-specified acceptance criteria, and whether the pre-specified acceptance criteria were met.
3) Your 809.10 compliant labeling must include:
i)A warning that reads “The assay results are intended to be interpreted only by a qualified pathologist or cytogeneticist.”
ii)A warning that reads “This device is not for high-risk uses such as selecting therapy, predicting therapeutic response or disease screening.”
iii)A warning that reads “The use of this device for diagnosis, monitoring or risk assessment has not been established.”

P Risks to Health and Mitigation Measures:

12

13

14

15

Q. Benefit/Risk Analysis:

| Summary of the
Benefits | Summary of
the Risks | Summary of Other
Factors | Conclusions |
|-----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|----------------------------------------------------------------------------------------------------------------------------------------------------------------------------|-----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|
| AML and MDS
patients may
potentially
benefit in the
intended use
population by use
of the device to
characterize bone
marrow
specimens with
assay results | Erroneous
device results
could adversely
influence
expectation of
more or less
favorable
clinical course
of AML or
MDS patients
due to false | In addition to cited
clinical studies using
the device probes,
analytical evaluation
and labeling supports
the intended use. De
novo regulatory
approach leverages
device use by a
qualified pathologist or
cytogeneticist in the | Yes. Based on
the supporting
clinical studies
for the
diagnostic
device along
with review of
the analytical
performance and
labeling, the
probable |
| interpreted by a
qualified
pathologist or
cytogeneticist. | negative or
false positive
results. | context of
histopathological
evaluation (e.g.,
immunohistochemistry). | benefits
outweigh the
probable risks. |

R. Conclusion:

16

The request for Evaluation of Automatic Class III Designation for this device is granted. The device is classified as Class II under regulation 21 CFR 864.1870 with special controls. The device is classified under the following:

Product Code:PDO
Device Type:Early growth response 1 (EGR1) gene fluorescence in-situ
hybridization (FISH) test system for specimen
characterization
Class:II (special controls)
Regulation:21 CFR 864.1870
(a) IDENTIFICATION: An early growth response 1
(EGR1) gene fluorescence in-situ hybridization (FISH) test
system for specimen characterization is a device intended
to detect the EGR1 probe target on chromosome 5q in bone
marrow specimens from patients with acute myeloid
leukemia (AML) or myelodysplastic syndrome (MDS).
The assay results are intended to be interpreted only by a
qualified pathologist or cytogeneticist. These devices do
not include automated systems that directly report results
without review and interpretation by a qualified pathologist
or cytogeneticist. These devices also do not include any
device intended for use to select patient therapy, predict
patient response to therapy or to screen for disease as well
as any device with a claim for a particular diagnosis,
prognosis, monitoring or risk assessment.
(b) CLASSIFICATION: Class II (special controls). EGR1
Gene FISH test system for specimen characterization must
comply with the following special controls:
  1. Premarket notification submissions must include the
    following information:
    i) A detailed description of all probes included in the kit
    ii) Purpose of each probe
    iii) Probe molecular specificity
    iv) Probe specificity
    v) Probe limits
    vi) Probe sensitivity
    vii) Specification of required ancillary reagents,
    instrumentation and equipment |

17

  • viii) Specification of the specimen collection, processing, storage and slide preparation methods
    • Specification of the assay procedure ix)
  • Specification of control elements that are x) incorporated into the recommended testing procedures
  • xi) Specification of risk mitigation elements: description of all additional procedures. methods, and practices incorporated into the directions for use that mitigate risks associated with testing
  • xii) Specification of the criteria for test result interpretation and reporting
  • xiii) Device analytical sensitivity data
  • xiv) Device analytical specificity data
  • XV) Device reference limit data
  • Device precision/reproducibility data xvi)
  • Device stability data to include: xvii)
    • A) Real-time Stability
    • B) Freeze-Thaw Stability
    • C) Transport and Temperature Stability
    • D) Post-Hybridization Signal Stability
    • E) Photostability of Probe
    • xviii) Documentation that demonstrates the clinical validity of the device. The documentation must include data from clinical studies, a minimum of two peer-reviewed published literature references using the specific device seeking marketing clearance, or both. Documentation for peer-reviewed published literature references cited must include the following elements:
      • A) Documentation that the sponsor's probe was used in the literature reference
      • B) Number & type of specimens
      • C) Target population studied. Target population must include the intended use population
      • D) Upper reference limit
      • F) Range of positive probe results
    1. 21 CFR 809.10(b)(12) compliant labeling must include a statement summarizing the data identified in subparagraphs (1)(xiii)-(xviii) and a description of the studies supporting the information, including

18

the pre-specified acceptance criteria for these performance studies, justification for the prespecified acceptance criteria, and whether the prespecified acceptance criteria were met.

    1. 21 CFR 809.10 compliant labeling must include:
    • i) A warning that reads "The assay results are intended to be interpreted only by a qualified pathologist or cytogeneticist."
    • ii) A warning that reads "This device is not for high-risk uses such as selecting therapy, predicting therapeutic response or disease screening."
    • iii) A warning that reads "The use of this device for diagnosis, monitoring or risk assessment has not been established."