The DPP Zika IgM System is intended for the presumptive qualitative detection of Zika virus IgM antibodies in human serum (plain or separation gel), potassium-EDTA plasma, potassium-EDTA venous whole blood, or fingerstick whole blood specimens, collected from individuals meeting the CDC Zika virus clinical criteria (e.g., a history of clinical signs and symptoms associated with Zika virus infection) and/or CDC Zika virus epidemiological criteria (e.g., history of residence in or travel to a geographic region with active Zika transmission at the time of travel, or other epidemiological criteria for which Zika virus testing may be indicated). Specimens from symptomatic patients or returning travelers from endemic areas should not be collected prior to 8 days after symptom onset or after potential exposure as a sample collected earlier may return a negative result. If testing is needed after day 8 and results are negative, testing must be repeated one week later. Positive results must be confirmed by following the latest CDC guidelines for the diagnosis of Zika virus infection.

Results of this test are intended to be used in conjunction with clinical observations, patient history, epidemiological information, and other laboratory results. Zika IgM levels over the course of illness are not well characterized. Zika IgM levels are variable during the course of infection and may be detectable near day 4 post-onset of symptoms and persist up to approximately 12 weeks following initial infection.

Negative results may be seen in specimens collected before day four post-onset of symptoms or after the window of detectable IgM closes and therefore do not preclude the possibility of Zika virus infection, past or present.

The Chembio DPP Zika IgM System is not indicated for testing blood or plasma donors.

The test cannot be visually interpreted by the operator and must be read on the DPP Micro Reader.

DPP Zika IgM System Control Pack

The Chembio DPP Zika IgM System Control Pack is an external quality control kit for use with the DPP Zika IgM System only. The performance characteristics of the DPP Zika IgM System Control Pack have not been established for any other assay or instrument different from the DPP Micro Reader.

DPP Micro Reader

The DPP Micro Reader is a reflectance reader used to obtain test results from DPP Zika IgM System. The DPP Micro Reader is necessary to minimize errors from direct visual interpretation; the results of DPP Zika IgM System cartridges must be read exclusively with the DPP Micro Reader.

Device Description

Chembio' s DPP Zika IgM System is a qualitative immunochromatographic assay for the presumptive detection of IgM antibodies to Zika virus. The DPP Zika IgM System is composed of:

A single-use immunochromatographic test for the presumptive detection of ZIK V 1. IgM antibodies in human serum (plain or separation gel), potassium-EDTA plasma, potassium-EDTA venous whole blood, or fingerstick whole blood specimens.
1. The DPP Micro Reader to minimize errors from direct visual interpretation.

AI/ML Overview

The document describes the Chembio DPP Zika IgM System, DPP Zika IgM System Control Pack, and DPP Micro Reader. The system is intended for the presumptive qualitative detection of Zika virus IgM antibodies in human serum, plasma, and whole blood specimens.

Here's a breakdown of the acceptance criteria and study details:

1. Table of Acceptance Criteria and Reported Device Performance

The acceptance criteria for the DPP Zika IgM System are primarily demonstrated through its Positive Percent Agreement (PPA) and Negative Percent Agreement (NPA) compared to a predicate EUA or FDA-cleared comparator assay, across various sample types. Analytical performance (precision, cross-reactivity, interference, analytical sensitivity, and stability) also forms part of the acceptance.

Performance Metric	Acceptance Criteria (Implied by achieved performance)	Reported Device Performance	Comments
Positive Percent Agreement (PPA)
Serum (Overall)	N/A (Achieved PPA above 95%)	95.1% (39/41; 95% CI=83.9-98.7%)	Compared to EUA comparator assay. Note: A high percentage of false positive results was observed against the comparator, largely contributed to by the source of the samples, but this metric is for samples expected to be positive.
Potassium-EDTA Plasma (Manufacturer Study)	N/A (Achieved PPA 100% for most relevant days post-onset)	100.0% (288/288) (95% CI 98.7-100.0%) for Days 8-84 post-onset of symptoms.	Data for 0-7 days post-onset excluded from overall PPA due to anticipated false negatives.
Potassium-EDTA Plasma (External Sites)	N/A (Achieved PPA 100%)	100.0% (171/171) (97.8-100.0%) for Days 8-84 post-onset of symptoms.
Potassium-EDTA Venous Whole Blood (Combined Studies)	N/A (Achieved PPA above 92%)	92.4% (194/210) (88-95.3%) for Days 8-84 post-onset of symptoms.	Note: Data for 0-7 days post-onset excluded.
Potassium-EDTA Venous Whole Blood (Contrived)	N/A (Achieved PPA above 99%)	Low Reactive: 99.1% (115/116); Moderate Reactive: 100% (63/63). Combined PPA: 99.4% (178/179) (95% CI 96.9-100%).	Compared against expected results based on spiking level.
Fingerstick Whole Blood (Contrived)	N/A (Achieved PPA above 98%)	Low Reactive: 98.0% (146/149); Moderate Reactive: 100.0% (88/88). Combined PPA: 98.7% (234/237) (95% CI 96.4-99.6%).	Compared against predetermined reactivity of spiked sample vials.
FDA Plasma Zika Panel (Zika IgM Consensus Positive)	N/A (Achieved PPA 95.8%)	95.8% (23/24)	PPA for Zika IgM Consensus Positive samples.
Negative Percent Agreement (NPA)
Serum (Endemic Area)	N/A (Achieved NPA above 94%)	95.7% (202/211) (95%CI 92.1-97.7)	By comparison with both FDA Cleared and EUA tests.
Potassium-EDTA Plasma (Endemic Area)	N/A (Achieved NPA above 93%)	93.8% (198/211) (95%CI 89.8-96.4)	By comparison with both FDA Cleared and EUA tests.
Potassium-EDTA Venous Whole Blood (Endemic Area)	N/A (Achieved NPA above 94%)	94.8% (200/211) (95%CI 90.9-97.1)	By comparison with both FDA Cleared and EUA tests.
Capillary Whole Blood (Endemic Area)	N/A (Achieved NPA above 97%)	97.2% (205/211) (95%CI 93.9-98.7)	By comparison with both FDA Cleared and EUA tests.
Serum (Non-Endemic Area)	N/A (Achieved NPA above 98%)	98.2% (220/224) (95%CI 95.5-99.3)	By comparison with both FDA Cleared and EUA tests.
Potassium-EDTA Plasma (Non-Endemic Area)	N/A (Achieved NPA above 97%)	97.8% (219/224) (95%CI 94.9-99.0)	By comparison with both FDA Cleared and EUA tests.
Potassium-EDTA Venous Whole Blood (Non-Endemic Area)	N/A (Achieved NPA above 97%)	97.3% (218/224) (95%CI 94.3-98.8)	By comparison with both FDA Cleared and EUA tests.
Capillary Whole Blood (Non-Endemic Area)	N/A (Achieved NPA above 98%)	98.7% (221/224) (95%CI 96.1-99.5)	By comparison with both FDA Cleared and EUA tests.
FDA Plasma Zika Panel (Zika IgM Consensus Negative)	N/A (Achieved NPA 91.7%)	91.7% (11/12)	NPA for Zika IgM Consensus Negative samples.
Cross-Reactivity	N/A (Evaluated cross-reactivity)	Cytomegalovirus: 5.3%; Dengue Virus: 2.0%	Other tested organisms/conditions (Chikungunya, West Nile, Yellow fever, Malaria, Borrelia, EBV, Hepatitis B/C, HSV-1/2, Leptospira, ANA, Parvovirus B19, Rubella, Rheumatoid Factor, Varicella zoster, HAMA) showed 0% cross-reactivity.
Interference	No interference observed	No interference observed for tested substances (Hemoglobin, Bilirubin, Proteins, HAMA, Cholesterol, Rheumatoid Factor, Triglycerides).
Analytical Sensitivity	N/A (Established cut-off values)	Serum: 650 IU/mL; Potassium-EDTA plasma: 700 IU/mL; Potassium-EDTA venous whole blood: 725 IU/mL	Based on WHO 1st International standard for anti-Asian lineage Zika virus antibody (human) (NIBSC 16/352).
Precision/Reproducibility	Coefficients of Variation (CV) provided for different components of variability.	Total CV: Moderate Positive (13.7%), Low Positive (19.0%), High Negative (24.5%), Negative (N/A)	For the DPP Zika IgM System.
DPP Zika IgM Control Kit Precision	Coefficients of Variation (CV) provided.	Total CV: Moderate Positive Control (17.9%), Low Positive Control (22.7%), Negative Control (N/A)
Assay Cut-off	Established at a specific reader value.	20 (when analyzed by the DPP Micro Reader)	Based on evaluation using CLSI EP17-A2.

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

Serum:
- Positive Predictive Agreement:
  - 99 samples from symptomatic individuals in Peru (retrospective).
  - 11 samples from individuals in the Dominican Republic (retrospective).
  - 32 samples from 26 individuals in Brazil during arbovirus outbreaks (retrospective).
  - FDA Plasma Zika Panel: 24 Zika IgM positive, 12 negative.
- Negative Predictive Agreement (Endemic Area): 250 subjects from a Zika endemic area (prospectively collected).
- Negative Predictive Agreement (Non-Endemic Area): 250 subjects from a Zika non-endemic area (prospectively collected).
Potassium-EDTA Plasma:
- Positive Predictive Agreement (Manufacturer Study): 299 IgM antibody samples from 48 individuals (from 50 symptomatic subjects) in the Dominican Republic (archived, confirmed by NAT). Includes 12 pregnant women.
- Positive Predictive Agreement (External Sites): 171 comparator positive IgM antibody samples from 39 individuals in the Dominican Republic (archived). Also, 49 prospectively collected asymptomatic pregnant women from the continental United States (negative by comparator assay) were interspersed.
- Negative Predictive Agreement (Endemic Area): 250 subjects from a Zika endemic area (prospectively collected).
- Negative Predictive Agreement (Non-Endemic Area): 250 subjects from a Zika non-endemic area (prospectively collected).
Potassium-EDTA Venous Whole Blood:
- Positive Predictive Agreement (Internal/External Sites): 41 plasma samples (from a plasma replacement study) from 6 individuals in the Dominican Republic; 10 frozen natural whole blood samples from individuals in the Dominican Republic; 171 antibody positive plasma specimens (from plasma replacement study) plus 49 antibody negative specimens from asymptomatic pregnant women from the US.
- Positive Predictive Agreement (Contrived): 299 subjects for analysis (from 300 "all comers" at 3 US clinics); samples were contrived by potassium-EDTA plasma replacement.
- Negative Predictive Agreement (Endemic Area): 250 subjects from a Zika endemic area (prospectively collected).
- Negative Predictive Agreement (Non-Endemic Area): 250 subjects from a Zika non-endemic area (prospectively collected).
Fingerstick Whole Blood:
- Positive Predictive Agreement (Contrived): 372 subjects for analysis (from 375 adult subjects across 4 US near-patient sites, "all comers" basis); samples were pre-spiked to create contrived samples.
- Negative Predictive Agreement (Endemic Area): 250 subjects from a Zika endemic area (prospectively collected).
- Negative Predictive Agreement (Non-Endemic Area): 250 subjects from a Zika non-endemic area (prospectively collected).
Cross-Reactivity: 329 specimens for various organisms/conditions.
Interference: Low reactive (n=3) and normal human plasma samples (n=3) for each interfering substance.
Analytical Sensitivity: Dilution series of WHO 1st International standard.
Assay Cut-off: 569 natural serum samples (US, Mexico); 184 natural plasma samples (US, Peru); 215 natural venous whole blood samples (US); 102 natural capillary whole blood samples (US).
Precision/Reproducibility: A blinded panel of four plasma samples (negative, high negative, low positive, moderate positive) tested with 3 lots of the system.
DPP Zika IgM Control Kit Precision: Moderate positive, low positive, and negative control samples.

Data Provenance: Studies include samples from Peru, Dominican Republic, Brazil, and the United States. Many samples are retrospective (archived, historical data), while some are prospectively collected (e.g., negative predictive agreement studies).

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

The document does not explicitly state the "number of experts" or their specific "qualifications" used to establish ground truth in the traditional sense of multiple human readers independently adjudicating images/cases.

Instead, the ground truth for clinical performance studies (PPA and NPA) was established by:

Comparison with an EUA (Emergency Use Authorization) or FDA-cleared comparator assay.
In some cases, samples were confirmed positive by RT-PCR assay for Zika virus (e.g., Peruvian and Brazilian serum samples).
Consensus results were provided for the FDA Plasma Zika Panel, and these are stated to be the responsibility of the FDA.
For contrived samples (venous whole blood, fingerstick whole blood), the "ground truth" was based on expected results from spiking with known positive or negative material, corroborated by comparator testing.

4. Adjudication Method for the Test Set

The primary method for "adjudication" (or reference standard determination) appears to be comparison with another laboratory assay (EUA or FDA-cleared comparator assay). For Negative Percent Agreement, results were compared against an FDA Cleared Comparator and an Additional EUA Test, and "Both FDA Cleared and EUA tests" where results from both were in agreement. There is no mention of a traditional expert consensus or adjudication panel (e.g., 2+1, 3+1) for clinical performance in this document.

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

No, a Multi-Reader Multi-Case (MRMC) comparative effectiveness study was not done. The study focuses on the standalone performance of the DPP Zika IgM System against reference methods/comparator assays, not on how human readers improve with or without AI assistance. The DPP Micro Reader is an integral part of the device, reading results from the immunochromatographic assay, and is not an AI tool assisting human interpretation. The document explicitly states the test "cannot be visually interpreted by the operator and must be read on the DPP Micro Reader," meaning it's not a human-in-the-loop scenario.

6. If a Standalone Study Was Done

Yes, a standalone study was done. The document provides extensive data on the performance of the DPP Zika IgM System (algorithm only, as it's an automated reader) in detecting Zika virus IgM antibodies across various sample types and clinical scenarios, independently compared to established reference standards (comparator assays, RT-PCR, or expected reactivity from spiked samples).

7. The Type of Ground Truth Used

The ground truth used various forms:

Comparator Assay Results: Results from an EUA or FDA-cleared Zika IgM comparator assay.
RT-PCR Confirmation: For some positive samples, PCR confirmation for Zika virus was used.
Consensus Data: For the FDA Plasma Zika Panel, consensus results were utilized.
Expected Reactivity from Spiked Samples: For contrived samples (venous whole blood, fingerstick whole blood), the ground truth was based on the known positive or negative status of the spiked material.

8. The Sample Size for the Training Set

The document does not explicitly specify a "training set" in the context of machine learning. The DPP Micro Reader and assay are likely developed based on extensive R&D and optimization, but the provided performance data relates to validation studies, not an explicit training set for a distinct AI model. The product is an in-vitro diagnostic device that produces quantitative results read by a device, not a machine learning algorithm requiring a separate training dataset in the typical sense.

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

Since an explicit "training set" for a machine learning model is not described, the method for establishing its ground truth is not applicable or detailed in this document. The development of the assay and the Micro Reader's algorithm would have involved internal validation and optimization using various samples, but these are part of product development rather than a defined, separate "training set" for regulatory evaluation in the context of this 510(k) summary.

Summary

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June 3, 2020

Chembio Diagnostic Systems Louise Muscat Sigismondi R&D Director of Regulatory Affairs 3661 Horseblock Road Medford, New York 11763

Re: K200506

Trade/Device Name: DPP Zika IgM System, DPP Zika IgM System Control Pack, and DPP Micro Reader

Regulation Number: 21 CFR 866.3935 Regulation Name: Zika Virus Serological Reagents Regulatory Class: Class II Product Code: QFO, QCH, JJQ Dated: February 27, 2020 Received: March 2, 2020

Dear Louise Sigismondi:

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. Although this letter refers to your product as a device, please be aware that some cleared products may instead be combination products. The 510(k) Premarket Notification Database located at https://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfpmn/pmn.cfm identifies combination product submissions. 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. Please note: CDRH does not evaluate information related to contract liability warranties. We remind you, however, that device labeling must be truthful and not misleading.

If your device is classified (see above) into either class II (Special Controls) or class III (PMA), it may be subject to additional controls. Existing major regulations affecting your device can be found in the Code of Federal Regulations, Title 21, Parts 800 to 898. 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

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requirements, including, but not limited to: registration and listing (21 CFR Part 807); labeling (21 CFR Part 801 and Part 809); medical device reporting of medical device-related adverse events) (21 CFR 803) for devices or postmarketing safety reporting (21 CFR 4, Subpart B) for combination products (see https://www.fda.gov/combination-products/guidance-regulatory-information/postmarketing-safety-reportingcombination-products); good manufacturing practice requirements as set forth in the quality systems (OS) regulation (21 CFR Part 820) for devices or current good manufacturing practices (21 CFR 4, Subpart A) for combination products; and, if applicable, the electronic product radiation control provisions (Sections 531-542 of the Act); 21 CFR 1000-1050.

Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21 CFR Part 807.97). For questions regarding the reporting of adverse events under the MDR regulation (21 CFR Part 803), please go to https://www.fda.gov/medical-device-safety/medical-device-reportingmdr-how-report-medical-device-problems.

For comprehensive regulatory information about mediation-emitting products, including information about labeling regulations, please see Device Advice (https://www.fda.gov/medicaldevices/device-advice-comprehensive-regulatory-assistance) and CDRH Learn (https://www.fda.gov/training-and-continuing-education/cdrh-learn). Additionally, you may contact the Division of Industry and Consumer Education (DICE) to ask a question about a specific regulatory topic. See the DICE website (https://www.fda.gov/medical-device-advice-comprehensive-regulatoryassistance/contact-us-division-industry-and-consumer-education-dice) for more information or contact DICE by email (DICE@fda.hhs.gov) or phone (1-800-638-2041 or 301-796-7100).

Sincerely,

Steven Gitterman, M.D., Ph.D. Deputy Director Division of Microbiology Devices OHT7: Office of In Vitro Diagnostics and Radiological Health Office of Product Evaluation and Quality Center for Devices and Radiological Health

Enclosure

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Indications for Use

510(k) Number (if known) K200506

Device Name

DPP Zika IgM System, DPP Zika IgM System Control Pack, and DPP Micro Reader

Indications for Use (Describe)

DPP Zika IgM System

The Chembio DPP Zika IgM System is not indicated for testing blood or plasma donors.

The test cannot be visually interpreted by the operator and must be read on the DPP Micro Reader.

DPP Zika IgM System Control Pack

DPP Micro Reader

Type of Use (Select one or both, as applicable)

X Prescription Use (Part 21 CFR 801 Subpart D)

Over-The-Counter Use (21 CFR 801 Subpart C)

CONTINUE ON A SEPARATE PAGE IF NEEDED.

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510(k) SUMMARY

Assigned 510(k) Number: K200506

I.	Date Prepared:	June 1, 2020
II.	Applicant Information:	Louise Sigismondi, Ph.D.Research & Development Director of Regulatory AffairsChembio Diagnostic Systems Inc.3661 Horseblock Rd.Medford, NY 11763Phone: 631-924-1135Fax: 631-924-2065Email: lsigismondi@chembio.com
III.	Regulatory Information:
	Trade Name:	DPP® Zika IgM SystemDPP® Zika IgM System Control PackDPP® Micro Reader
	Common Name:	Zika virus assay
	Classification Names:	Zika Virus Serological Reagents: Class II, 21 CFR 866.3935; Microbiology (83)Assayed quality control material for clinical microbiology assays: Class II, 21CFR 866.3920; Microbiology (83)Colorimeter, Photometer, Spectrophotometer for Clinical Use: Class I, 21 CFR §862.2300; Clinical Chemistry (75)
	Product Code:	QFO - Zika Virus Serological ReagentsQCH - Assayed quality control material for clinical microbiology assaysJJQ - Colorimeter, Photometer, Spectrophotometer for Clinical Use
IV.	Predicate Device:	InBios ZIKV Detect™ 2.0 IgM Capture ELISA(DEN180069)

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V. Intended use/Indication for use:

DPP Zika IgM System

The DPP Zika IgM System is intended for the presumptive qualitative detection of Zika virus IgM antibodies in human serum (plain or separation gel), potassium-EDTA plasma, potassium-EDTA venous whole blood, or fingerstick whole blood specimens, collected from individuals meeting the CDC Zika virus clinical criteria (e.g., a history of clinical signs and symptoms associated with Zika virus infection) and/or CDC Zika virus epidemiological criteria (e.g., history of residence in or travel to a geographic region with active Zika transmission at the time of travel, or other epidemiological criteria for which Zika virus testing may be indicated). Specimens from symptomatic patients or returning travelers from endemic areas should not be collected prior to 8 days after symptom onset or after potential exposure as a sample collected earlier may return a negative result. If testing is needed after day 4 but before day 8 and results are negative, testing must be repeated one week later. Positive results must be confirmed by following the latest CDC guidelines for the diagnosis of Zika virus infection.

The Chembio DPP Zika IgM System is not indicated for testing blood or plasma donors.

The test cannot be visually interpreted by the operator and must be read on the DPP Micro Reader.

DPP Zika IgM System Control Pack

VI. Device Description:

Chembio' s DPP Zika IgM System is a qualitative immunochromatographic assay for the presumptive detection of IgM antibodies to Zika virus. The DPP Zika IgM System is composed of:

A single-use immunochromatographic test for the presumptive detection of ZIK V 1. IgM antibodies in human serum (plain or separation gel), potassium-EDTA

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plasma, potassium-EDTA venous whole blood, or fingerstick whole blood specimens.

1. The DPP Micro Reader to minimize errors from direct visual interpretation.

Materials provided

Each kit contains the reagents and tools to perform 20 tests:

20 individually pouched DPP Zika IgM Test Devices, each containing:

1 DPP Zika Test Device (membrane strip with immobilized recombinant Zika NS-● 1 antigen in the TEST (T) area and Protein A in the CONTROL (C) area.
1 Desiccant Pouch ●

20 Disposable 10 uL Microsafe Tubes

20 Sample vials

20 Transfer Pipets (100 µl)

1 DPP Zika IgM Buffer- YELLOW Cap
- 7.5 mL contains sodium phosphate, sodium chloride, EDTA, NP-40, Tween 20, ● Urea, chicken serum, gentamicin, streptomycin, and sodium azide as preservative.
1 Product Insert for the DPP Zika IgM System

1 Quick Reference Guide for the DPP Zika IgM System

Materials required but not provided (system related)

Chembio DPP Micro Reader. ● Each kit contains:
- DPP Micro Reader with Zika IgM RFID sticker
  - o 3 Lithium-ion, type CR2032 (3 V/230 mAh), coin cell batteries (installed)
- Custom power adapter cable (USB to 2.0 mm jack) -
- Power plug adaptor -
- DPP Cartridge Holder -
- Microfiber cloth -
- -User Manual

Additional required materials (assay related)

Chembio DPP Zika IgM System Control Pack (Catalog #62-1001-1) ●
- 1 DPP Zika Reactive Control (300 µl): undiluted, naturally occurring Zika IgM positive plasma samples.
- 1 DPP Zika Non-Reactive Control (300 µ1): undiluted, naturally occurring Zika -IgM negative plasma samples.
- 1 DPP Zika Diluent (300 µl): undiluted, naturally occurring Zika IgM negative plasma samples.
- 1 Product Insert -

All reagents are supplied ready to use.

Clock, watch, or other timing device ●
Pipettor capable of delivering 10-100 µL of sample may be used in lieu of the ● disposable 10 uL MicroSafe Tube and 100 uL Transfer Pipets supplied with the Kit (for serum, potassium-EDTA plasma, and potassium-EDTA venous whole blood specimens or with the Chembio DPP Zika IgM System Control Pack)

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●

Microcentrifuge Tubes
Disposable gloves
Antiseptic wipes ●
Biohazard disposal container
For fingerstick whole blood specimens:
- Sterile gauze
- Sterile Safety Lancets for fingerstick whole blood specimens
- For venous whole blood or serum/plasma specimens:
- Collection devices

For a fingerstick whole blood sample, sample collection uses a 10 ul Microsafe Tube. For human serum (plain or separation gel), potassium-EDTA plasma, potassium-EDTA venous whole blood specimens, 10 ul of sample is collected using a calibrated laboratory pipet. Once the sample and buffer combination are added to the DPP Zika IgM Test Device, results are read by using the Chembio DPP Micro Reader, a portable, maintenance-free, batterypowered instrument that is operated by a single, multi-function button. The results of the DPP Zika IgM Test Device are presented through a 14-segment liquid crystal display (LCD) on the top of the DPP Micro Reader. The DPP Micro Reader is necessary to minimize errors from direct visual interpretation, therefore the results must not be visually interpreted by the operator. Controls are kit lot specific and must not be interchanged between different DPP Zika IgM System lots.

VII. Purpose for Submission:

The purpose of this premarket notification is to submit a new device (DPP Zika IgM System) to FDA for consideration for clearance as a 510(k).

VIII. Comparison with predicate:

A comparison of the similarities and differences between the DPP Zika IgM System and the predicate is provided in the table below:

Item	DPP Zika IgM System(K200506)	ZIKV DetectTM 2.0 IgM Capture ELISA(DEN 180069)
Similarities
Analyte	Human IgM antibodies	Human IgM antibodies
Intended Use	The DPP Zika IgM System is intended for the presumptive qualitative detection of Zika virus IgM antibodies in human serum (plain or separation gel), potassium-EDTA plasma, potassium-EDTA venous whole blood, or fingerstick whole blood specimens, collected from individuals meeting the CDC Zika virus clinical criteria (e.g., a history of clinical signs and symptoms associated with Zika virus infection) and/or CDC Zika virus epidemiological criteria (e.g., history of residence in or travel to a	The ZIKV Detect 2.0 IgM Capture ELISA is intended for the qualitative detection of Zika virus IgM antibodies in human sera for the presumptive clinical laboratory diagnosis of Zika virus infection. The assay is intended for use only in patients with clinical signs and symptoms consistent with Zika virus infection, and/or CDC Zika virus epidemiological criteria (e.g., history of residence in or travel to a geographic region with active Zika transmission at the time of

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	geographic region with active Zika transmission atthe time of travel, or other epidemiological criteriafor which Zika virus testing may be indicated).Specimens from symptomatic patients or returningtravelers from endemic areas should not becollected prior to 8 days after symptom onset orafter potential exposure as a sample collectedearlier may return a negative result. If testing isneeded after day 4 but before day 8 and results arenegative, testing must be repeated one week later.Positive results must be confirmed by followingthe latest CDC guidelines for the diagnosis of Zikavirus infection.Results of this test are intended to be used inconjunction with clinical observations, patienthistory, epidemiological information, and otherlaboratory results. Zika IgM levels over thecourse of illness are not well characterized. ZikaIgM levels are variable during the course ofinfection and may be detectable near day 4 post-onset of symptoms and persist up toapproximately 12 weeks following initialinfection.Negative results may be seen in specimenscollected before day four post-onset of symptomsor after the window of detectable IgM closes andtherefore do not preclude the possibility of Zikavirus infection, past or present.The Chembio DPP Zika IgM System is notindicated for testing blood or plasma donors.The test cannot be visually interpreted by theoperator and must be read on the DPP MicroReader.	travel, or other epidemiological criteria forwhich Zika virus testing may be indicated).Assay results are for the presumptivedetection of IgM antibodies to Zika virus(ZIKV). Positive results must be confirmedby following the latest CDC guidelines forthe diagnosis of Zika virus infection.Results of this test are intended to be used inconjunction with clinical observations,patient history, epidemiological information,and other laboratory evidence to makepatient management decisions. Zika IgMlevels are variable over the course of theinfection, and may be detectable near dayfour post onset of symptoms and persist upto approximately 12 weeks following initialinfection.Negative results may be seen in specimenscollected before day four post onset ofsymptoms or after the window of detectableIgM closes, and therefore do not preclude thepossibility of Zika virus infection, past orpresent.This assay is not indicated for testing bloodor plasma donors.
Differences
Sample Type	Human serum (plain or separation gel), potassium-EDTA plasma, potassium-EDTA venous wholeblood, or fingerstick whole blood specimens	Human serum
Sample Size	Minimum of 10 µL before dilution	Minimum of 4 µL before dilution
AntibodyTarget	NS-1 protein	Zika envelope glycoproteins
Type of Test	Lateral cross-flow dual path platform (DPP)immunochromatographic assay	IgM Capture enzyme-linked immunosorbentassay (ELISA)
ExternalQuality Control	Provided Separately	Included
Interpretation ofResults	Qualitative analysis by instrument: reactive ornon-reactive for Zika IgM antibodies; invalid	Qualitative analysis by instrumentPresumptive Zika Positive, PresumptiveOther Flavivirus Positive (non-Zika);negative

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	Interpretation of Results is performed by the DPPMicroreader. No calculations are required by theuser.	Interpretation requires calculationsperformed by the user prior to reporting theresult
Time to Result	Time to result is 15 minutes after addition ofBuffer into Well#2.	Time to result is approximately 3.5 hrs.
ReagentStorage	2-8°C Refrigerator or up to 23°C	2-8°C Refrigerator only

IX. Performance Standard:

Standard	Title
CLSI EP05-A3	Evaluation of Precision Performance of Quantitative Measurement Methods; ApprovedGuideline
CLSI EP07-3rd Ed.	Interference Testing in Clinical Chemistry; Approved Guideline—Third Edition
CLSI EP12-A2	User Protocol for Evaluation of Qualitative Test Performance; Approved Guideline, SecondEdition
CLSI EP15-A3	User of Performance for Precision and Trueness, Approved Guideline—Third Edition
CLSI EP17-A2	Evaluation of Detection Capability for Clinical Laboratory Measurement Procedures; ApprovedGuideline - Second Edition
CLSI EP25-A	Evaluation of Stability of In Vitro Diagnostic Reagents; Approved Guideline.
CLSI EP37	Supplemental Tables For Interference Testing In Clinical Chemistry
IEC 60601-1-2	Medical electrical equipment - Part 1-2: General requirements for basic safety and essentialperformance - Collateral Standard: Electromagnetic disturbances - Requirements and tests
21CFR 820	Quality System Regulation
21CFR 809.10	Labeling for in vitro diagnostic products
21 CFR §866.3935	Special controls, Zika virus serological reagents
21 CFR §862.2300	Colorimeter, photometer, or spectrophotometer for clinical use

X. Performance Characteristics:

a. Clinical Studies

Positive Predictive Agreement

Positive Percent Agreement (PPA) was estimated for serum (plain or separation gel), potassium-EDTA plasma, potassium-EDTA venous whole blood, or fingerstick whole blood specimens by comparison with either the EUA or the FDA-cleared comparator assay.

Serum

Positive percent agreement for serum samples was evaluated using the following samples collected from patients residing in flavivirus areas:

. 99 serum samples collected from symptomatic individuals living in Peru, collected within 5-7 days after the onset of symptoms. All samples tested positive by a RT-PCR assay for Zika virus. Only 98 were tested in the EUA comparator assay due to a volume shortage in one sample.

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. 11 serum samples sera from individuals living in the Dominican Republic, 6 being Zika-positive by a RT-PCR assay.
32 serum samples collected from 26 individuals during a time of arbovirus . outbreaks in a flavivirus endemic region of Brazil. All 26 individuals tested positive by a RT-PCR assay for Zika virus. The samples collected from individuals living in Brazil consisted of an initial serum specimen collected from 26 individuals on 1 to 17 days following the onset of fever symptoms, and a second serum specimen collected from 6 of these individuals after 6 to 9 days following the initial blood collection, for a total of 32 specimens.

For samples 'expected' to be positive, forty-one (41) samples were positive by the comparator assay. Overall PPA for serum samples with the EUA comparator assay was 95.1% (39/41; 95% CI=83.9-98.7%). A high percentage of false positive results was observed against the comparator, largely contributed to by the source of the samples.

Overall Serum Agreement
		Comparator1Zika IgM Reactive		Comparator1Zika IgM Non-Reactive
CollectionSite	Total(n)	DPP Zika IgMSystemReactive(R)2	DPP Zika IgMSystemNon-Reactive(NR)2	PositivePercentAgreement(95% CI)	DPP Zika IgMSystemReactive(R)2	DPP Zika IgMSystemNon-Reactive(NR)2	NegativePercentAgreement
DominicanRepublic	11	8	23	80%(8/10)	14	0	0%(0/1)
Peru	98	23	0	100%(23/23)5	68	7	9.3%(7/755,6)
Brazil	32	87	0	100%(8/8)	4	20	83.3%(20/24)
Total	141	39	2	95.1%(39/41)8(83.9-98.7%)	73	27	27%(27/100)

Overall Serum Agreement

1 EUA comparator assay reactive samples include Possible and Presumptive Zika Positive Specimens. EUA comparator assay non-reactive samples include Negative and Presumptive Other Flavivirus Positive specimens.

2This study was performed with an older version of the DPP Zika IgM System that was determined equivalent to the one under clearance.

3Negative for Zika IgM antibody by both the DPP Zika IgM System and a second authorized Zika IgM assay, while positive by an EUA Zika virus rtPCR assay and the EUA test used as a comparator.

"Negative for Zika IgM antibody by both the EUA test used as a comparator and a second authorized Zika IgM assay, while positive by an EUA Zika virus rtPCR assay.

5 All samples tested positive by a rtPCR assay for Zika virus.

6 All samples were < 7 days post onset of symptoms. Even when positive by PCR, depending on the senstivity of the assay, the level of antibodies may be low for detection.

There were 8 positive samples in total in the 32. Two out of eight (2/8) samples were a single sample picked in both the first and second blood draw; 1/8 was only picked on the first blood draw; 5/8 were picked only on the second blood draw. 8Two of the eight (2/8) Brazilian positive sample picked in both the first and second blood draw and counted twice for final performance calculations. Without double counting the specimen, the total PPA would be 95.0% (83.5-98.6%).

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Potassium-EDTA Plasma

Positive percent agreement for potassium-EDTA plasma was evaluated using archived samples consisting of eight serial samples collected from 50 symptomatic subjects confirmed positive for Zika virus by nucleic acid testing from the Dominican Republic. Of these, 299 IgM antibody samples from 48 individuals tested positive by the comparator. Samples included 12 pregnant women. Testing was performed at the manufacturer's laboratory. The results from this study demonstrate 100% positive agreement between the DPP Zika IgM Assay and the comparator for Zika IgM-positive potassium-EDTA plasma samples. Results are presented in the following table:

Days Post-Onset of Symptoms (Manufacturer Study)
Days Post onset ofSymptoms	Comparator1Zika IgM Reactive2Positive Percent Agreement	Comparator1Zika IgM Non-Reactive3Negative Percent Agreement
	0-7*	100.0% (39/39)
8-14	54.5% (6/11)	72.5% (29/40)
15-28	100.0% (82/82)	0% (0/4)
29-42	100.0% (79/79)	0% (0/7)
43-56	100.0% (61/61)	0% (0/21)
57-70	100.0% (19/19)	10% (1/10)
71-84	100.0% (8/8)	0% (0/3)
Total	100.0% (288/288)(95% CI 98.7-100.0%)	2.2% (1/45)

Potassium-EDTA Plasma Panel, Stratified by vs Post-Onset of Symptoms (Manufacturer Stu

Data from Days 0 – 7 is not included in the calculation of total PPA because obtaining samples prior to 8 days after symptom onset or exposure is not recommended and false negative results are anticipated. 1 EUA comparator assay

2 Comparator reactive samples include Possible and Presumptive Zika Positive Specimens.

3 Comparator non-reactive samples include Negative and Presumptive Other Flavivirus Positive specimens. NT: Not tested.

Positive percent agreement was also evaluated at external sites using 171 comparator positive IgM antibody potassium-EDTA plasma specimens from 39 individuals living in the Dominican Republic. Forty-nine (49) potassium-EDTA plasma samples prospectively collected from asymptomatic pregnant woman living in the continental United States that were found to be negative in the cleared comparator assay were interspersed for blinding. The 220 samples were divided and sent to two external clinical sites. The combined data are summarized in the following table.

Days Post-Onset of Symptoms (External Sites)
Days Post-Onset of Symptoms	Comparator Assay Zika IgM Reactive1		Positive Percent Agreement (95% CI)
	DPP Zika IgM System Reactive	DPP Zika IgM System Non-Reactive
8-14	29	0	100.0% (29/29)(88.3-100.0%)
15-28	48	0	100.0% (48/48)(92.6-100.0%)
29-42	46	0	100.0% (46/46)(92.3-100.0%)

Potassium-EDTA Plasma Panel, Stratified by

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43-56	29	0	100.0% (29/29)(88.3-100.0%)
57-70	12	0	100.0% (12/12)(75.8-100.0%)
71-84	7	0	100.0% (7/7)(64.6-100.0%)
Total	171	0	100.0% (171/171)(97.8-100.0%)
NonendemicNormal /Pregnant	Comparator Assay Zika IgM Non-Reactive2		Negative PercentAgreement (95% CI)
Asymptomatic	DPP Zika IgM SystemReactive	DPP Zika IgM SystemNon-Reactive
	1	48	98.0% (483/49)(89.3-99.6%)

1 EUA comparator assay reactive samples include Possible and Presumptive Zika Positive Specimens.

2 Cleared comparator assay non-reactive samples include Negative and Presumptive Other Flavivirus Positive (non-Zika) specimens.

3 For one sample, the microreader value indicated a negative result, but the end-user recorded an interpretation of "R" for Reactive.

Potassium-EDTA Venous Whole Blood

The PPA of the DPP Zika IgM System for potassium-EDTA venous whole blood was evaluated by two studies. For the first study, samples were tested at either an internal site or at external sites:

Samples tested at the internal site: ●
- 41 plasma samples prepared from a plasma replacement study with o serial bleeds over days from symptom onset from 6 individuals residing in the Dominican Republic. All donors were confirmed positive for Zika virus antibodies by the comparator assay.
- Frozen natural potassium-EDTA venous whole blood samples from ten o (10) antibody positive individuals from the Dominican Republic by the comparator assay.
Samples tested at external sites: ●
- 171 antibody positive plasma specimens from a plasma replacement O study using the same subset described above for potassium-EDTA plasma plus the 49 antibody negative potassium-EDTA plasma specimens from asymptomatic pregnant women from the US as described above. Results were confirmed by the comparator assay.

Results are summarized in the following table:

1 Plasma replacement is used here to describe a procedure by which venous whole blood was centrifuged, the plasma portion was removed and the pellet carefully suspended in an equal volume of plasma from another source positive for Zika antibodies.

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Comparator1			Comparator
Days Postonset ofSymptoms	Zika IgM Reactive		Zika IgM Non-Reactive
	PositivePercentAgreement	95%ConfidenceInterval	Negative PercentAgreement
Negative2	0%(0/1)	N/A	98%(483/49)
0-7*	83.3%(10/12)	55.2-95.3%	NT
8-14	97.1%(334/34)	85.1-99.5%	NT
15-28	94.7%(54/57)	85.6-98.2%	NT
29-42	96.4%(53/55)	87.7-99.0%	NT
43-56	84.2%(32/38)	69.6-92.6%	NT
57-70	80.0%12/15	54.8-93.0%	0%(0/1)1,5
71-84	90.9%(10/11)	62.3-93.4%	NT
Total			96.0%(48/50)
Total8-84	92.4%(194/210)	88-95.3%

Potassium-EDTA Venous Whole Blood Agreement Across Internal and Clinical Sites

1 EUA comparator assay reactive samples include Possible and Presumptive Zika Positive Specimens. 2 Cleared comparator assay non-reactive samples include Negative and Presumptive Other Flavivirus Positive (non-Zika) specimens.

3 For one sample, the microreader value indicated a negative result, but the end-user recorded an interpretation of "R" for Reactive.

4 For one sample, the microreader value indicated a reactive result, but the end-user recorded an interpretation of "NR" for Non-Reactive.

5 One sample was negative by 2nd authorized Zika IgM serology test.

Data from Days 0 - 7 is not included in the calculation of total PPA because obtaining samples prior to 8 days after symptom onset or exposure is not recommended and false negative results are anticipated. NT: Not Tested, and not included in the calculations.

The second study was conducted at 3 clinics in the US using prospectively-collected potassium-EDTA venous whole blood from subjects enrolled in an "all comers" fashion. One of the 300 subjects was excluded because their naïve whole blood tested reactive in the DPP Zika IgM System without being spiked, leaving 299 subjects for analysis. In order to assess positive agreement, samples were contrived by potassium-EDTA plasma replacement into venous whole blood samples to yield negative, low positive, and moderate positive specimens. The contrived samples were tested in a blinded fashion across three

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sites. Results were compared against the expected results based on the spiking level for each sample.

Sample	Agreement	Score95% CI	Combined Agreement
Negative	100%(120/120)	96.9-100.0%	NPA100% (120/120)(95%CI 96.9-100%)
Low Reactive(2.5x - < 3x Cut-Off)	99.1%(115/116)	95.3-99.9%	PPA99.4% (178/179)
Moderate Reactive(≥3x - 5x Cut-Off)	100%(63/63)	94.3-100.0%	PPA99.4% (178/179)(95%CI 96.9-100%)

Venous Whole Blood Results

Fingerstick Whole Blood

The performance of the DPP Zika IgM System for fingerstick whole blood was evaluated at 4 near-patient sites in the US. A total of 375 adult subjects across the four sites were enrolled on an "all comers" basis. Fresh seronegative fingerstick whole blood samples (non-anticoagulated) were collected using sample vials that were pre-spiked with Zika IgM antibody positive plasma or potassium-EDTA plasma to create negative, low positive and moderate positive Zika samples. Presence or absence of Zika IgM antibodies in the spiking material was corroborated by comparator testing. Of the 375 subjects, two (2) were eliminated due to protocol deviations in data transcription and one (1) subject was excluded because their naïve whole blood tested reactive in the DPP Zika IgM System without being spiked, leaving 372 subjects for analysis. The vials were coded so that study staff and the two trained operators at the clinical laboratory were not aware of the predetermined reactivity of the spiked sample vials. The results are summarized below.

Sample	Agreement	Score 95%CI	Combined Agreement
Negative	97.8%(132/135)	93.7-99.2%	NPA97.8% (132/135)(95%CI 93.7-99.2%)
Low Reactive(2.5x - <3x Cut-Off)	98.0%(146/149)	92.3-99.3%	PPA98.7% (234/237)(95%CI 96.4-99.6%)
Moderate Reactive(≥3x - 5x Cut-Off)	100.0%(88/88)	95.8-100.0%

Fingerstick Whole Blood Results

Negative Predicative Agreement

Negative Percent Agreement of the DPP Zika IgM System was established using prospectively collected potassium-EDTA venous and fingerstick whole blood samples from 250 subjects living in a Zika endemic area and 250 subjects living in a Zika nonendemic area. Potassium-EDTA venous samples and fingerstick whole blood were tested

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at seven (7) point-of-care sites including four (4) sites in areas endemic for mosquito-borne flaviviruses and three (3) sites in non-endemic areas.

Paired serum and potassium-EDTA plasma were forwarded to a reference laboratory where they were tested for Zika virus antibodies with the DPP Zika IgM System and the comparator method. Samples were expected to be negative for Zika IgM antibodies due to the low prevalence of Zika virus during the study. For those samples found positive, results were expressed as a percentage of positive results in the total population under study. Samples were tested using the DPP Zika IgM System, the cleared comparator assay, and an additional serology EUA test. Results are summarized in the following tables.

Endemic Area

		NPA# of DPP Zika IgM System byComparison with Samples Negative By		Percent of Positive Results in Population
	FDAClearedComparator¹	AdditionalEUA Test²	Both FDAClearedand EUAtests	DPP ZikaIgMSystem³	FDAClearedComparator¹	AdditionalEUA Test²
Serum	94.3%(213/226)95%CI90.4-98.6	94.8%(218/230)95%CI91.1-97.0	95.7%(202/211)95%CI92.1-97.7	8.0%(20ᵃ/250)	9.6%(24/250)	7.6%(19/250)
Potassium-EDTAPlasma	93.4%(211/226)95%CI89.3-95.9	93.0%(214/230)95%CI89.0-95.7	93.8%(198/211)95%CI89.8-96.4	8.8%(22ᵇ/250)	9.6%(24/250)	7.6%(19/250)
Potassium-EDTAVenous WholeBlood	93.8%(212/226)95%CI89.9-96.3	94.4%(217/230)95%CI90.6-96.7	94.8%(200/211)95%CI90.9-97.1	7.6%(19ᶜ/250)	9.6%(24/250)	7.6%(19/250)
Capillary WholeBlood	96.0%(217/226)95%CI92.6-97.9	96.5%(222/230)95%CI93.3-98.2	97.2%(205/211)95%CI93.9-98.7	5.6%(14ᵈ/250)	9.6%(24/250)	7.6%(19/250)

#Calculated using the score method.

1 Detects IgM antibodies against envelope

2 Detects IgM and IgG antibodies anti NS-1

3 Detects IgM antibodies anti NS-1

ª 15 agree in 3 or all matrices, the remaining in 2 or less

b 14 agree in 3 or all matrices, the remaining in 2 or less

c 14 agree in 3 or all matrices, the remaining in 2 or less

d 11 agree in 3 or all matrices, the remaining in 2 or less

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Non-Endemic Area

Non-Endemic Area
	NPA# of DPP Zika IgM System byComparison with Samples Negative By			Percent of Positive Results in Population
	FDAClearedComparator¹	AdditionalEUATest²	Both FDAClearedand EUAtests	DPP ZikaIgMSystem³	FDAClearedComparator¹	AdditionalEUATest²
Serum	98.0%(239/244)95%CI95.3-99.1	98.3%(225/229)95%CI95.6-99.3	98.2%(220/224)95%CI95.5-99.3	2.0%(5ᵃ/250)	2.4%(6/250)	8.4%(21/250)
Potassium-EDTAPlasma	96.7%(236/244)95%CI93.7-98.3	97.8%(224/229)95%CI95.0-99.1	97.8%(219/224)95%CI94.9-99.0	3.2%(8ᵇ/250)	2.4%(6/250)	8.4%(21/250)
Potassium-EDTAVenous WholeBlood	97.1%(237/244)95%CI94.2-98.6	97.4%(223/229)95%CI94.4-98.8	97.3%(218/224)95%CI94.3-98.8	2.8%(7ᶜ/250)	2.4%(6/250)	8.4%(21/250)
Capillary WholeBlood	98.4%(239/243)⁴95%CI95.9-99.4%	98.7%(226/229)95%CI96.2-99.6	98.7%(221/224)95%CI96.1-99.5	1.6%(4ᵈ/250)	2.4%(6/250)	8.4%(21/250)

Calculated using the score method.

1 Detects IgM antibodies against envelope

² Detects IgM and IgG antibodies anti NS-1

3 Detects IgM antibodies anti NS-1

4 FS result from one subject excluded due to protocol deviation (Test read < 15-minutes)

a 4 agree in 3 or all matrices, the remaining in 2 or less

b 4 agree in 3 or all matrices, the remaining in 2 or less

c 4 agree in 3 or all matrices, the remaining in 2 or less

d 3 agree in 3 or all matrices, the remaining in 2 or less

Performance Against the FDA Plasma Zika Panel

Performance of the DPP Zika IgM System was evaluated by testing a panel of samples provided to the sponsor by FDA. The panel consists of plasma samples from individuals infected with Zika, West Nile, or Dengue viruses. Samples were collected at different time points from Zika infection documented by PCR; samples taken early or late after infection may be anticipated to be antibody negative. Samples were randomized and blinded for testing. Results were as follows:

		DPP Zika IgM System
		PresumptiveZika Positive	Negative
Zika IgMConsensus	Positive(n=24)	23	1
	Negative(n=12)	1	11

PPA= 23/24 = 95.8% NPA= 11/12 = 91.7%

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		DPP Zika IgM System
		PresumptiveZika Positive(FalsePositives)	Negative
Cross-reactivityEvaluation	West Nile*(n=10)	1	9
	Dengue*(n=10)	0	10

Samples were antibody positive for West Nile or Dengue viruses, and negative for Zika virus.

This evaluation was performed using samples provided by Blood Systems Research Institute (BSRI, now Vitalant Research Institute) from a study supported by Contract No. HHSN2682011000011 from the National Heart, Lung, and Blood Institute (NHLBI), National Institutes of Health. The panel composition and consensus results are the responsibility of the FDA and do not necessarily represent the official views of BSRI, the NHLBI, or the National Institutes of Health.

b. Analytical performance:

Precision/Reproducibility of the DPP Zika IgM System: i.
DPP Zika IgM System reproducibility was evaluated at 3 different US sites with two runs per day, each run performed by one operator with one unique instrument (6 operators and 6 instruments in total) on 5 separate days. All samples were run as three replicates using three (3) lots of the DPP Zika IgM System. A blinded panel consisting of four plasma samples, including a 'negative', 'high negative', 'low positive,' and 'moderate positive,' were tested in this study.

Precision estimates were derived for reproducibility.

Sites, Days, Operators			Within-Run		Between-Operator/Instrument		Between-Day		Between-Lot		Between-Sites		Total*
Sample ID	MeanValue	N	SD	%CV	SD	%CV	SD	%CV	SD	%CV	SD	%CV	SD	%CV
ModeratePositive	110.0	270	13.4	12.2	2.8	2.5	0.0	0.0	6.3	5.8	0.0	0.0	15.1	13.7
Low Positive	36.5	270	5.5	15.0	2.4	6.7	0.0	0.0	3.6	1.0	0.0	0.0	7.0	19.0
HighNegative	15.1	270	3.3	21.5	1.5	9.8	0.0	0.0	1.0	6.6	0.0	0.0	3.7	24.5
Negative	1.7	270	1.3	n/a	0.0	n/a	0.0	n/a	0.0	n/a	1.3	n/a	1.8	n/a
%CV	coefficient of variation expressed as a percentage: SD standard deviation.

Reproducibility of the DPP Zika IgM System

Total = Within-Run + Between-Operator/Instrument + Between-Day + Between-Lot + Between-Site

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Precision/Reproducibility of the DPP Zika IgM Control Kit ii.

Internal precision for the DPP Zika IgM Control Kit was tested at a single site over the course of 5 days by two different technicians using two instruments per operator. All samples were run in duplicate per run with 2 runs per day, morning and afternoon. One lot of the DPP Zika IgM System and one lot of the DPP Zika IgM Control Kit consisting of a DPP Zika Reactive Control, a DPP Zika Non-Reactive Control and a DPP Zika Diluent was used. The DPP Zika Reactive Control and DPP Zika Diluent was used in a ratio of 1:3 to make the low-reactive control as per the instructions for use fresh, each day of testing.

		Within-Run		Between-Run		Between-Instrument		Between-Operator		Between-Day		Total*
SampleID	MeanReaderValue	N	SD	%CV	SD	%CV	SD	%CV	SD	%CV	SD	%CV	SD	%CV
ModeratePositiveControl	54.1	80	8.5	15.7	0.0	0.0	4.7	8.6	0.0	0.0	0.0	0.0	9.7	17.9
LowPositiveControl	31.9	80	5.6	17.5	0.9	3.0	1.8	5.6	4.2	13.0	0.0	0.0	7.2	22.7
NegativeControl	3.4	80	1.3	n/a	0.3	n/a	0.6	n/a	0.0	n/a	0.3	n/a	1.5	n/a

5-day Precision Results for DPP Zika IgM System Control Pack

Total = Within-Run + Between-Run + Between-Instrument + Between-Operator + Between-Day

iii. Linearity/assay reportable range:

N/A

Analytical specificity: iv.

a. Cross-Reactivity:

Cross-reactivity of the DPP Zika IgM System was evaluated by testing a total of 329 specimens from patients with IgM antibodies to closely related viruses as well as pathogens where infection produces symptoms similar to those observed during Zika virus infection, and autoantibodies which could potentially cause false positive results. A total of eighteen (18) different organisms/conditions per the supplier's Certificate of Analysis for various disease states, using FDA cleared/ approved devices when possible, were assessed. Of those, two (2) were found to be cross-reactive on the DPP Zika IgM System. The % Cross Reactivity for Cytomegalovirus and Dengue Virus on the DPP Zika IgM System is 5.3% and 2%, respectively. Babesia cross-reactivity was not evaluated.

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Summary of DPP Zika IgM System Cross Reactivity

	Organism/Condition	# of Specimens	# ZikaNegative	# ZikaPositive	% Cross-reactivity
Flavivirus	Anti-Chikungunya virus IgM	26	26	0	0%
	Anti-Dengue Virus IgM	48	47	1*	2.0%
	Anti-West Nile Vile IgM	28	28	0	0%
	Yellow fever virus post-immunization1	32	32	0	0%
OtherViruses/diseases	Anti- Malaria/anti-plasmodium falciparum2	20	20	0	0%
	Anti-Borrelia sp. (Lyme Disease) IgM	10	10	0	0%
	Anti-Cytomegalovirus (CMV) IgM	38	36	2**	5.3%
	Anti-Epstein Barr Virus (EBV) IgM	10	10	0	0%
	Anti-Hepatitis (B) virus (Total)	13	13	0	0%
	Anti-Hepatitis (C) virus (Total)	10	10	0	0%
	Anti-Herpes simplex virus 1 and 2 (HSV-1, HSV-2) IgM	23	23	0	0%
	Anti-Leptospira (Lepotospirosis) IgM3	16	16	0	0%
	Anti-nuclear Antibodies (ANA)	11	11	0	0%
	Anti-Parvovirus B19 IgM	7	7	0	0%
	Anti-Rubella virus IgM	12	12	0	0%
	Rheumatoid Factor	12	12	0	0%
	Anti-Varicella zoster virus IgM	3	3	0	0%
	Human Anti-mouse Antibody (HAMA)	10	10	0	0%
	Total	329	326	3

Negative by an FDA cleared Zika Assay. West Nile equivocal.

** Negative by an FDA cleared Zika Assay.

1 IgM levels could not be established.

2 Specimens were confirmed positive for Malaria infection by Giemsa and Microscopy but serological status is not known.

3 Pretreated with rheumatoid factor-absorbent prior to IgM detection.

b. Interference:

Potentially interfering endogenous substances were evaluated with the DPP Zika IgM System. Interfering substances were spiked into low reactive (n=3) and normal human plasma samples (n=3) to evaluate their impact on assay performance. Only one operator was involved in interpreting results in these studies. No interference was observed.

11 1 2 1 2 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1A CONSULERSON FOR ULL BALL RATE LETTE LETTE U FOUNDER
		DPP Zika IgM system
	(#reactive/#tested)
Interfering Substance	Concentration	Negative DiluentZika IgM Spike
		(Non-Reactive)	(2x LoD)
Hemoglobin	10 mg/mL	0/3	3/3
Bilirubin, Conjugated	0.4 mg/mL	0/3	3/3

Interfering Substances for the DPP Zika IgM System

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Bilirubin, Un-Conjugated	0.4 mg/mL	0/3	3/3
Serum Proteins	60 mg/mL	0/3	3/3
HAMA	732 ng/mL	0/3	3/3
Cholesterol	4 mg/mL	0/3	3/3
Rheumatoid Factor	2000 IU/mL	0/3	3/3
Triglycerides	15 mg/mL	0/3	3/3

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

a. Traceability/Analytical Sensitivity:

The analytical sensitivity at the cut-off values for the DPP Zika IgM System was established using WHO 1st International standard for anti-Asian lineage Zika virus antibody (human) (NIBSC 16/352). This preparation contains antibodies reactive to Dengue virus. The standard was used to prepare a dilution series. The concentration of the reference reagent that corresponds to average reader values just above the cut-off of 20 (average values of <30 when analyzed by the DPP Micro Reader) and where at least 95% of all replicates tested reactive was 650 IU/mL in serum, 700 IU/mL in potassium-EDTA plasma, and 725 IU/mL in potassium-EDTA venous whole blood.

b. Stability:

Evaluations to determine the stability of the Chembio DPP Zika IgM System, Chembio DPP Zika IgM controls and the storage stability of human plasma and serum specimens are summarized below.

Summary of Stability Claims for the DPP Zika IgM Assay System Reagents and Controls
Stability Type	Claim
DPP Zika IgM Assay System Device Shelf-Life Stability (up to 23 ℃)	6 months
DPP Zika IgM Assay System Device Shelf-Life Stability (2-8 °C)	12 months
DPP Zika IgM System Reagent In Use (up to 23 ℃)	1 month when used within the expiration date
DPP Zika IgM Assay System Control Shelf-Life Stability (-20 ℃)	6 months
DPP Zika IgM System Control In Use Stability (-20 °C)	N/A*
DPP Zika IgM System Control Freeze-Thaw Cycles	3

It is recommended that the end user divide the controls into smaller aliquots that are then frozen back. The end user dilutes the controls only before use, and immediately discards any remaining material after use. Frozen aliquots are acceptable if they are both within the freeze-thaw cycles and the shelf life claimed above.

The user must not open the pouch until ready to perform a test.

The Chembio DPP Zika IgM System must be performed on human serum (plain or separation gel), potassium-EDTA plasma, potassium-EDTA venous whole blood, or fingerstick whole blood specimens. Studies were performed to determine the stability of plasma and serum samples at storage temperatures of 2 to 8℃ (36 to 46°F). A multiple freeze/thaw (F/T) study was also performed. Testing was performed with reactivities across the dynamic range of the assay including negative and low positive samples. Stability of samples was determined to be as follows:

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Storage	Serum	Plasma
Refrigerated at 2- 8 °C	3 days	3 days
Freeze-thaw cycles	2 cycles	5 cycles

Potassium-EDTA venous whole blood and fingerstick whole blood should be tested at the time of specimen collection.

Detection limit: vi.
See Section v.a. Traceability/Analytical Sensitivity
vii. Assay cut-off:
Assay cut-off was evaluated following CLSI EP17-A2 describing the calculation of the Limit of Blank (LoB), using a non-parametric method based on the % cumulative frequency of the results. Samples tested included:
569 natural serum samples sourced from the United States and Mexico, both ● non-endemic populations;
184 natural plasma samples sourced from a non-endemic population from ● the United States (n=95) and an endemic population from Peru (n= 89);
215 natural venous whole blood samples sourced from a non-endemic population from the United States;
102 natural capillary whole blood samples sourced from a non-endemic population from the United States.

Assay cut-off was set at 20 when analyzed by the DPP Micro Reader.

viii. Prozone/ Hook Effect:
Chembio's proprietary DPP technology differs from classical lateral flow tests by operating in a manner similar to that of the sequential ELISA format which is not sensitive to the "Hook Effect". On the primary flow path of DPP devices, the sample migrates towards the immobilized immunoreagents on the horizontal strip that captures the analyte of interest, if present. Following a brief incubation to maximize analyte binding efficiency, the detector nanoparticles are released from the conjugate pad via the secondary flow path. This sequential approach resembles the traditional ELISA assay process and minimizes the potential of the prozone (or hook) effect. Formal Hook Effect testing was not performed.
ix. Matrix Comparison:
Matrix equivalency was established in comparison to natural matched potassium-EDTA plasma samples or expected reactivity after positive potassium-EDTA plasma spiking. Matrices tested were:
11 positive and negative natural matched frozen potassium-EDTA whole

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blood, potassium-EDTA plasma and serum samples

49 whole blood samples prepared by plasma-replacement using positive ● potassium-EDTA plasma specimens
30 samples for each serum and potassium-EDTA plasma prepared from 5 ● individual negative sera and plasma sources and spiked with up to a 10% volume of positive potassium-EDTA plasma specimens to obtain high negative, low positive, and 4 values across the dynamic range of the DPP assay run in duplicate
50 matched negative sample sets for serum and potassium-EDTA plasma ● samples sources and spiked with up to a 10% volume of positive potassium-EDTA plasma specimens to obtain high negative, low positive, mid positive, and high positive specimens run in duplicate. Un-spiked negative samples were also included.

Of the multiple combinations of antibody levels and negative individual matrix; disagreement between plasma and serum was observed only for high negative specimens prepared by spiking. Above the cut-off of the assay, there was 100% agreement between matrices.

Summary Table for Matrix Equivalency of the DPP Zika IgM System with Known Positive and Negative Zika Specimens

Matrix	Potassium EDTA Whole Blood	Serum^
Potassium EDTA Plasma	100% (60/60)(% CI 94.0 - 100%)	97.3% (88.5/91)(%CI 91.5-99.2%)

^ Average of mean agreement values for each experiment were used. Some studies have 1 while others 2 replicates per sample. When one replicate agreed and one disagreed, it was counted as 0.5 sample agreement.

X. Class Specificity

To determine if cross reactivity with IgG is a potential assay interferent, a study was performed where the reducing agent Dithiothreitol (DTT) was added to three (3) clinical specimens containing various levels of Zika virus IgM antibodies and high levels of Zika virus IgG antibodies.

All IgM positive samples resulted in non-reactive DPP Zika IgM System results after treatment with DTT. The study results support IgM class specificity.

xi. Carry-Over:
Not applicable as a single sample is run per device.

XI. Conclusion:

The submitted information in this premarket notification is complete and supports a substantial equivalence decision. The labelling is sufficient and it satisfies the requirements of 21CFR 809.10

Regulation Number and Section

§ 866.3935 Zika virus serological reagents.

(a)
Identification. Zika virus serological reagents arein vitro diagnostic devices that consist of antigens or antibodies for the detection of Zika virus or Zika antibodies in human specimens from individuals who have signs and symptoms consistent with Zika virus infection and/or epidemiological risk factors. The detection aids in the diagnosis of current or recent Zika virus infection or serological status. Negative results obtained with this test do not preclude the possibility of Zika virus infection, past or present. Positive results should be interpreted with consideration of other clinical information and laboratory findings and should not be used as the sole basis for treatment or other patient management decisions.(b)
Classification. Class II (special controls). The special controls for this device are:(1) The labeling required under § 809.10(b) of this chapter must include:
(i) An intended use with a detailed description of what the device detects (Zika IgM antibodies, other Zika antibodies, or Zika antigens), the type of results provided to the user, the specimen type for which testing is indicated (
e.g., serum, whole blood), the clinical indications appropriate for test use, and the specific population(s) for which the test is intended.(ii) Performance characteristics from analytical and clinical studies required under paragraphs (b)(2)(ii) and (iii) of this section.
(iii) A detailed explanation of the interpretation of results and criteria for validity of results (
e.g., criteria that internal or external quality controls must meet in order for a test/test run to be valid, minimum signal strength that the sample has to yield to be interpretable as a valid result).(iv) Limiting statements indicating that:
(A) Results are not intended to be used as the sole basis for diagnosis, treatment, or other patient management decisions. The test results should be interpreted in conjunction with clinical observations, patient history, epidemiological information, and other laboratory evidence.
(B) Device results are intended to be followed up according to the latest professional guidelines (
e.g., recommendations from the Centers for Disease Control and Prevention) for the diagnosis of Zika virus infection.(C) Negative test results do not preclude the possibility of Zika virus infection, past or present.
(D) Specimens can result in false negative results on the device if collected outside of the appropriate response window for specific Zika virus antigens or antibodies, as determined by scientific evidence (
e.g., for IgM