The miDiagnostics HSV-1&2 CSF Test is an in vitro polymerase chain reaction (PCR) assay intended for use with the miDiagnostics PCR Platform for the qualitative detection and differentiation of HSV-1 and HSV-2 DNA in cerebrospinal fluid (CSF) samples from patients suspected of Herpes Simplex Virus (HSV) infection of the central nervous system (CNS). This test is intended as an aid in the diagnosis of HSV-1 and HSV-2 infection in the CNS.

Negative results do not preclude HSV-1 or HSV-2 infection and should not be used as the sole basis for treatment or other patient management decisions.

The test is not intended for use as a donor screening test. The test is for professional use only.

Device Description

The miDiagnostics HSV-1&2 CSF Test is a real-time polymerase chain reaction (PCR) assay that enables the direct amplification, detection, and differentiation of Herpes Simplex Virus 1 (HSV-1) and Herpes Simplex Virus 2 (HSV-2) DNA from cerebrospinal fluid (CSF) specimens, obtained via lumbar puncture, from individuals with signs and/or symptoms of HSV-1 or HSV-2 infection in the central nervous system (CNS).

After manual sample and PCR reaction preparation, the miDiagnostics HSV-1&2 CSF test consists of three major steps: (1) manual sample preparation, (2) manual PCR reaction preparation and (3) automated PCR run on the miDiagnostics PCR Platform which is an automated in vitro diagnostic system for the amplification and detection of the target nucleic acid(s) using real-time polymerase chain reaction (PCR).

Description of the different components of miDiagnostics HSV-1&2 CSF Test:

miDiagnostics HSV-1&2 CSF Detection Mix: contains a freeze-dried cake, stored in a tube, containing all the PCR components for amplification and fluorescent detection of HSV-1, HSV-2 and the Sample Processing Control (SPC). Each tube is individually packed in a pouch. The device consists of 24 pouches in a box.
miDiagnostics HSV-1&2 CSF Dilution Buffer: contains 300 µL HSV CSF Dilution Buffer for diluting the CSF specimen. It also contains a SPC (RPP30, a human housekeeping gene). The device consists of 24 tubes packed in one box.
miDiagnostics Concentration Device: consists of a combination of the miDiagnostics Concentration Column and 2 miDiagnostics Collection Tubes, to concentrate the viral material. The device consists of one bag with 24 miDiagnostics Concentration Columns and 48 miDiagnostics Collection Tubes.
miDiagnostics HSV-1&2 CSF Algorithm: The software system that interprets fluorescence signals and reports test results. The miDiagnostics HSV-1&2 CSF Algorithm is embedded in the miDiagnostics Software.

The miDiagnostics PCR Platform is an automated in vitro diagnostics medical device intended to be used in combination with a selected miDiagnostics test for the amplification and detection of the target nucleic acid(s) using real-time polymerase chain reaction (PCR). The miDiagnostics PCR Platform is intended to be used by healthcare professionals (trained laboratory personnel) in laboratory environment.

Description of the different components of miDiagnostics PCR Platform:

miDiagnostics PCR Reader 2.0 receives the miDiagnostics PCR Card 2.0 and is responsible for performing consecutive PCR cycles through thermocycling. The miDiagnostics PCR Reader 2.0 monitors the consequent polymerase chain reaction's progress with a four-channel camera system. It also contains a 2D barcode reader to identify the loaded miDiagnostics PCR Card 2.0 and is connected to the miDiagnostics Computer, from which it receives test-specific parameters for the PCR and to which it sends the camera images that contain the raw data of the PCR.

The firmware is an embedded software system running on the miDiagnostics PCR Reader 2.0 microprocessor with as main functionalities the thermocycling and the triggering of the camera.

miDiagnostics PCR Card 2.0 is a single-use, disposable test accessory. The miDiagnostics PCR Card 2.0 is manually loaded with a small volume of sample mixed with PCR components of the selected miDiagnostics test (so called reaction mixture). Once loaded, the miDiagnostics PCR Card 2.0 is manually inserted in the miDiagnostics PCR Reader 2.0. The miDiagnostics PCR Card 2.0 includes a fluidic chip with the reaction chamber that enables a very fast polymerase chain reaction driven by the miDiagnostics PCR Reader 2.0. Each miDiagnostics PCR Card 2.0 is individually packed in a pouch. The device consists of 24 pouches in a box.
miDiagnostics Computer connects to the miDiagnostics PCR Reader 2.0 via two USB-cables and can connect to the customer's network via ethernet or WiFi. It interacts with a user via a touchscreen, has an integrated barcode scanner that allows the user to scan the sample consumable identifiers, and it runs the miDiagnostics Software.

The miDiagnostics Software is the software of the miDiagnostics PCR Platform, that is installed on the miDiagnostics Computer. It controls the miDiagnostics PCR Reader 2.0, retrieves the camera captures, hosts the test specific algorithm that interprets the fluorescence signals, reports the test results, and provides a user interface for the operator to interact with the miDiagnostics PCR Platform.

AI/ML Overview

N/A

Summary

U.S. Food & Drug Administration 510(k) Clearance Letter

Page 1

U.S. Food & Drug Administration
10903 New Hampshire Avenue
Silver Spring, MD 20993
www.fda.gov

Doc ID # 04017.08.00

September 30, 2025

miDiagnostics nv
Katrin Martens
Lead Regulatory Submissions
Gaston Geenslaan 1
Leuven, Flemish Brabant 3001
Belgium

Re: K250050
Trade/Device Name: miDiagnostics HSV-1&2 CSF Test
Regulation Number: 21 CFR 866.3307
Regulation Name: Herpes Simplex Virus Nucleic Acid-Based Assay For Central Nervous System Infections
Regulatory Class: Class II
Product Code: PGH
Dated: September 4, 2025
Received: September 4, 2025

Dear Katrin Martens:

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 (the 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 available 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.

Additional information about changes that may require a new premarket notification are provided in the FDA guidance documents entitled "Deciding When to Submit a 510(k) for a Change to an Existing Device"

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K250050 - Katrin Martens Page 2

(https://www.fda.gov/media/99812/download) and "Deciding When to Submit a 510(k) for a Software Change to an Existing Device" (https://www.fda.gov/media/99785/download).

Your device is also subject to, among other requirements, the Quality System (QS) regulation (21 CFR Part 820), which includes, but is not limited to, 21 CFR 820.30, Design controls; 21 CFR 820.90, Nonconforming product; and 21 CFR 820.100, Corrective and preventive action. Please note that regardless of whether a change requires premarket review, the QS regulation requires device manufacturers to review and approve changes to device design and production (21 CFR 820.30 and 21 CFR 820.70) and document changes and approvals in the device master record (21 CFR 820.181).

Please be advised that FDA's issuance of a substantial equivalence determination does not mean that FDA has made a determination that your device complies with other requirements of the Act or any Federal statutes and regulations administered by other Federal agencies. You must comply with all the Act's requirements, including, but not limited to: registration and listing (21 CFR Part 807); labeling (21 CFR Part 801 and Part 809); medical device reporting (reporting of medical device-related adverse events) (21 CFR Part 803) for devices or postmarketing safety reporting (21 CFR Part 4, Subpart B) for combination products (see https://www.fda.gov/combination-products/guidance-regulatory-information/postmarketing-safety-reporting-combination-products); good manufacturing practice requirements as set forth in the quality systems (QS) regulation (21 CFR Part 820) for devices or current good manufacturing practices (21 CFR Part 4, Subpart A) for combination products; and, if applicable, the electronic product radiation control provisions (Sections 531-542 of the Act); 21 CFR Parts 1000-1050.

Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21 CFR 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-devices/medical-device-safety/medical-device-reporting-mdr-how-report-medical-device-problems.

For comprehensive regulatory information about medical devices and radiation-emitting products, including information about labeling regulations, please see Device Advice (https://www.fda.gov/medical-devices/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-devices/device-advice-comprehensive-regulatory-assistance/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,

Mayra Garcia, Ph.D., M.B.A. on behalf of
Jorge L. Munoz, Ph.D.
Deputy Branch Chief
Division of Microbiology Devices
OHT7: Office of In Vitro Diagnostics
Office of Product Evaluation and Quality
Center for Devices and Radiological Health

Digitally signed by MAYRA GARCIA -S
Date: 2025.09.30 16:17:02 -04'00'

Page 3

FORM FDA 3881 (8/23) Page 1 of 1 PSC Publishing Services (301) 443-6740 EF

DEPARTMENT OF HEALTH AND HUMAN SERVICES
Food and Drug Administration

Indications for Use

Form Approved: OMB No. 0910-0120
Expiration Date: 07/31/2026
See PRA Statement below.

510(k) Number (if known): K250050

Device Name: miDiagnostics HSV-1&2 CSF Test

Indications for Use (Describe)

Negative results do not preclude HSV-1 or HSV-2 infection and should not be used as the sole basis for treatment or other patient management decisions.

The test is not intended for use as a donor screening test. The test is for professional use only.

Type of Use (Select one or both, as applicable)
☒ Prescription Use (Part 21 CFR 801 Subpart D) ☐ Over-The-Counter Use (21 CFR 801 Subpart C)

CONTINUE ON A SEPARATE PAGE IF NEEDED.

This section applies only to requirements of the Paperwork Reduction Act of 1995.
DO NOT SEND YOUR COMPLETED FORM TO THE PRA STAFF EMAIL ADDRESS BELOW.

The burden time for this collection of information is estimated to average 79 hours per response, including the time to review instructions, search existing data sources, gather and maintain the data needed and complete and review the collection of information. Send comments regarding this burden estimate or any other aspect of this information collection, including suggestions for reducing this burden, to:

Department of Health and Human Services
Food and Drug Administration
Office of Chief Information Officer
Paperwork Reduction Act (PRA) Staff
PRAStaff@fda.hhs.gov

"An agency may not conduct or sponsor, and a person is not required to respond to, a collection of information unless it displays a currently valid OMB number."

Page 4

510(k) Summary

miDiagnostics HSV-1&2 CSF Test

Submitter

Applicant Name and Address	miDiagnostics nvGaston Geenslaan 13001 LeuvenBelgium
Contact Person	Katrin Martens
Date Prepared	09/30/2025

Device Information

Trade Name	Regulation Name	Regulation Number	Product code	Class
miDiagnostics HSV-1&2 CSF Test	Herpes simplex virus nucleic acid-based assay for central nervous system infections	21 CFR 866.3307	PGH	II

Intended use

Negative results do not preclude HSV-1 or HSV-2 infection and should not be used as the sole basis for treatment or other patient management decisions.

The test is not intended for use as a donor screening test. The test is for professional use only.

Device Description

Page 5

Description of the different components of miDiagnostics HSV-1&2 CSF Test:

miDiagnostics HSV-1&2 CSF Detection Mix: contains a freeze-dried cake, stored in a tube, containing all the PCR components for amplification and fluorescent detection of HSV-1, HSV-2 and the Sample Processing Control (SPC). Each tube is individually packed in a pouch. The device consists of 24 pouches in a box.
miDiagnostics HSV-1&2 CSF Dilution Buffer: contains 300 µL HSV CSF Dilution Buffer for diluting the CSF specimen. It also contains a SPC (RPP30, a human housekeeping gene). The device consists of 24 tubes packed in one box.
miDiagnostics Concentration Device: consists of a combination of the miDiagnostics Concentration Column and 2 miDiagnostics Collection Tubes, to concentrate the viral material. The device consists of one bag with 24 miDiagnostics Concentration Columns and 48 miDiagnostics Collection Tubes.
miDiagnostics HSV-1&2 CSF Algorithm: The software system that interprets fluorescence signals and reports test results. The miDiagnostics HSV-1&2 CSF Algorithm is embedded in the miDiagnostics Software.

Description of the different components of miDiagnostics PCR Platform:

miDiagnostics PCR Reader 2.0 receives the miDiagnostics PCR Card 2.0 and is responsible for performing consecutive PCR cycles through thermocycling. The miDiagnostics PCR Reader 2.0 monitors the consequent polymerase chain reaction's progress with a four-channel camera system. It also contains a 2D barcode reader to identify the loaded miDiagnostics PCR Card 2.0 and is connected to the miDiagnostics Computer, from which it receives test-specific parameters for the PCR and to which it sends the camera images that contain the raw data of the PCR.

The firmware is an embedded software system running on the miDiagnostics PCR Reader 2.0 microprocessor with as main functionalities the thermocycling and the triggering of the camera.
miDiagnostics PCR Card 2.0 is a single-use, disposable test accessory. The miDiagnostics PCR Card 2.0 is manually loaded with a small volume of sample mixed with PCR components of the selected miDiagnostics test (so called reaction mixture). Once loaded, the miDiagnostics PCR

Page 6

Card 2.0 is manually inserted in the miDiagnostics PCR Reader 2.0. The miDiagnostics PCR Card 2.0 includes a fluidic chip with the reaction chamber that enables a very fast polymerase chain reaction driven by the miDiagnostics PCR Reader 2.0. Each miDiagnostics PCR Card 2.0 is individually packed in a pouch. The device consists of 24 pouches in a box.

miDiagnostics Computer connects to the miDiagnostics PCR Reader 2.0 via two USB-cables and can connect to the customer's network via ethernet or WiFi. It interacts with a user via a touchscreen, has an integrated barcode scanner that allows the user to scan the sample consumable identifiers, and it runs the miDiagnostics Software.

The miDiagnostics Software is the software of the miDiagnostics PCR Platform, that is installed on the miDiagnostics Computer. It controls the miDiagnostics PCR Reader 2.0, retrieves the camera captures, hosts the test specific algorithm that interprets the fluorescence signals, reports the test results, and provides a user interface for the operator to interact with the miDiagnostics PCR Platform.

Principle of operations

The test consists of three major steps: (1) manual sample preparation, (2) manual PCR reaction preparation and (3) automated PCR run which includes targets amplification and detection by fluorescent probes for 1 target specific to HSV-1, 1 target specific to HSV-2 and the SPC. The SPC is a human DNA control target and is used to verify that miDiagnostics HSV-1&2 CSF Test was executed successfully and that no PCR inhibition has occurred.

The sample preparation starts out with adding 200 μL CSF to the miDiagnostics HSV-1&2 CSF Dilution Buffer. The diluted specimen is then transferred to the miDiagnostics Concentration Device and centrifuged at high speed for 5 minutes. The remaining volume in the miDiagnostics Concentration Column is washed with molecular grade water (DNase & RNase free) and centrifuged again at high speed for 5 minutes. The sample is then heated for 2 minutes at 95°C. Thereafter, an aliquot of the sample is mixed with the miDiagnostics HSV-1&2 CSF Detection Mix and loaded on the miDiagnostics PCR Card 2.0, to fill the reaction chamber of the fluidic chip.

The miDiagnostics PCR Card 2.0 is manually inserted in the miDiagnostics PCR Reader 2.0 for amplification and detection of HSV-1, HSV-2 and the SPC. The miDiagnostics PCR Reader 2.0 rapidly heats and cools the chip to drive a polymerase chain reaction inside the reaction chamber of the fluidic chip. Via the transparent optical window in the miDiagnostics PCR Card 2.0's bottom, the progress of the reaction is measured by a camera system in the miDiagnostics PCR Reader 2.0, that sends the raw images to the miDiagnostics Computer for analysis by the miDiagnostics Software. The miDiagnostics HSV-1&2 CSF Algorithm (hosted by miDiagnostics Software) automatically interprets the fluorescence signals and reports the test results.

Page 7

Substantial Equivalence

Table 1: Substantial Equivalence Comparison.

Device Trade Name	Predicate Device: (K133621)Simplexa HSV 1 & 2 Direct	New Device: (K250050)miDiagnostics HSV-1&2 CSF Test
General Device Characteristic Similarities
Intended Use /Indications for Use	The Focus Diagnostics Simplexa™ HSV 1 & 2 Direct is intended for use on the 3M Integrated Cycler instrument for the qualitative detection and differentiation of HSV-1 and HSV-2 DNA in cerebrospinal fluid (CSF) samples from patients suspected of Herpes Simplex Virus (HSV) infections of the central nervous system (CNS).This test is intended as an aid in the diagnosis of HSV-1 and HSV-2 infections of the CNS.Negative results do not preclude HSV-1 or HSV-2 infection and should not be used as the sole basis for treatment or other patient management decisions.The assay is not intended for use as a donor screening test.The assay is for professional use only.The Positive Control is intended to be used as a control with the Simplexa™ HSV 1 & 2 Direct. This control is not intended for use with other assays or systems.	The miDiagnostics HSV-1&2 CSF Test is an in vitro polymerase chain reaction (PCR) assay intended for use with the miDiagnostics PCR Platform for the qualitative detection and differentiation of HSV-1 and HSV-2 DNA in cerebrospinal fluid (CSF) samples from patients suspected of Herpes Simplex Virus (HSV) infection of the central nervous system (CNS). This test is intended as an aid in the diagnosis of HSV-1 and HSV-2 infection in the CNS.Negative results do not preclude HSV-1 or HSV-2 infection and should not be used as the sole basis for treatment or other patient management decisions.The test is not intended for use as a donor screening test.The test is intended for professional use only.
Targets	Herpes Simplex Virus 1 (HSV-1) DNAHerpes Simplex Virus 2 (HSV-2) DNA	Same
Specimen type	Cerebrospinal fluid (CSF)	Same
Analyte detected	DNA	Same

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Device Trade Name	Predicate Device: (K133621)Simplexa HSV 1 & 2 Direct	New Device: (K250050)miDiagnostics HSV-1&2 CSF Test
Technology	Real-time Polymerase Chain Reaction	Same
Result interpretation	Automated	Same
PCR Control	Assay contains Control to detect reagent failure and/or inhibition	Same
General Device Characteristic Differences
Instrumentation	3M Integrated Cycler Studio system	miDiagnostics PCR Platform
External Controls	The Positive Control is provided and intended to be used as a control with the Simplexa™ HSV 1 & 2 Direct. This control is not intended for use with other assays or systems.	External control materials are not provided with the miDiagnostics HSV-1&2 CSF Test. External controls are required to run with the test.

Performance

Analytical performance

Analytical Sensitivity (Limit of Detection)

The Limit of Detection (LOD) for the miDiagnostics HSV-1&2 CSF Test was determined using two HSV-1 strains (MacIntyre and HF strains) and two HSV-2 strains (MS and G strains). Serial dilutions of each strain were made in pooled negative human CSF matrix (negative CSF) near their LOD during feasibility. Tests were performed over the course of 4 days, with 2 reagent batches and 9 miDiagnostics PCR Platforms. Ten replicates at each dilution series were tested per viral strain per reagent batch with a total of 20 replicates per strain. The concentration at which ≥ 95% of replicates were detected was the preliminary LOD for that strain. The preliminary LOD for each strain was subsequently confirmed with 20 replicates diluted at the preliminary LOD concentration using the worst performing reagent batch. The confirmed LOD per strain is listed in Table 2.

Table 2: Limit of Detection (LOD) summary table for the miDiagnostics HSV-1&2 CSF Test.

Virus Strain	LOD(TCID50/mL)	LOD(copies/mL)	#Detected/#Total(% of Detection)
HSV-1 MacIntyre	8.84	800	20/20(100%)
HSV-1 HF	20.32	1000	20/20(100%)
HSV-2 MS	0.56	400	19/20(95%)
HSV-2 G	0.43	800	20/20(100%)

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Analytical Reactivity (Inclusivity)

The analytical reactivity of the miDiagnostics HSV-1&2 CSF Test was evaluated with four HSV-1 and three HSV-2 strains, see Table 3. Viral material with known concentration was spiked into negative CSF at near LOD concentration and assessed in triplicates. Testing with increasing concentrations was continued until all strains generated "Detected" results for all three replicates. The miDiagnostics HSV-1&2 CSF Test was able to detect all HSV strains.

Table 3: Analytical reactivity (Inclusivity) summary table for the miDiagnostics HSV-1&2 CSF Test.

Analyte	Virus strain	Spiked concentration
HSV-1	HSV-1 strain F	20.32 TCID50/mL
	HSV-1 strain KOS	40.64 TCID50/mL
	HSV-1 ATCC-2011-1	20.32 TCID50/mL
	HSV-1 strain 95 (95/1906)	20.32 TCID50/mL
HSV-2	HSV-2 strain 131596	1605 copies/mL
	HSV-2 strain HG52	3.36 TCID50/mL
	HSV-2 ATCC-2011-2	0.56 TCID50/mL

Analytical Specificity

Cross-reactivity

The cross-reactivity of the miDiagnostics HSV-1&2 CSF Test was evaluated by testing 46 microorganisms that may be present in CSF. The cross-reactants were spiked into negative CSF at high concentrations and assessed in triplicate. No cross-reactivity was observed in any of the microorganisms listed in Table 4.

Table 4: Summary table of microorganisms tested for cross reactivity for the miDiagnostics HSV-1&2 CSF Test.

Microorganism	Concentration tested
Adenovirus (type 1)	105 TCID50/mL
Adenovirus (type 7A)	105 TCID50/mL
BK polyomavirus	105 TCID50/mL
Citrobacter freundii	106 CFU/mL
Citrobacter koseri	106 CFU/mL
Cryptococcus neoformans	106 CFU/mL
Cytomegalovirus (AD169)	3.12 x 103 TCID50/mL

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Microorganism	Concentration tested
Dengue virus	105 copies/mL
Enterobacter aerogenes	106 CFU/mL
Enterovirus 71	7.78 x 103 TCID50/mL
Epstein Barr virus (B95-8)	105 TCID50/mL
Escherichia coli	106 CFU/mL
Haemophilus influenzae	106 CFU/mL
Haemophilus influenzae type b (MinnA)	106 CFU/mL
Hepatitis B	105 TCID50/mL
Hepatitis C1	105 TCID50/mL
HIV1 (type IIIB)	104 IU/mL
Human herpesvirus 6	105 TCID50/mL
Human herpesvirus 7	9.36 x 103 TCID50/mL
Human herpesvirus 8	1.45 x 105 TCID50/mL
Influenza B	2.80 x 104 TCID50/mL
Influenza A H1N1	1.46 x 105 TCID50/mL
John Cunningham virus	105 TCID50/mL
Klebsiella pneumoniae	106 CFU/mL
Listeria monocytogenes	106 CFU/mL
Measles	105 TCID50/mL
Mumps	105 TCID50/mL
Mycobacterium tuberculosis	106 copies/mL
Naegleria fowleri	106 copies/mL
Neisseria meningitides (serogroup A)	106 CFU/mL
Parainfluenza Virus 1	105 TCID50/mL
Parainfluenza Virus 2	8.34 x 104 TCID50/mL
Parainfluenza Virus 3	105 TCID50/mL
Parvovirus B19	1.28 x 102 TCID50/mL

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Microorganism	Concentration tested
Proteus mirabilis (Z050)	106 CFU/mL
Pseudomonas aeruginosa	106 CFU/mL
Rabies virus	Not available *
Rhinovirus (Type 1A)	2.82 x 104 TCID50/mL
Rotavirus (Type Wa)	105 TCID50/mL
Rubella	2.58 x 103 TCID50/mL
St. Louis Encephalitis	105 copies/mL
Staphylococcus aureus COL	106 CFU/mL
Streptococcus agalactiae	106 CFU/mL
Streptococcus pneumoniae Z022; 19F	106 CFU/mL
Varicella zoster virus	105 TCID50/mL
West Nile Virus	105 TCID50/mL

* No official concentration provided by the supplier. The maximum volume of the pathogen's RNA extract was spiked for testing without exceeding 20% of the total sample volume.

Cross-reactivity with Polio virus and La Crosse encephalitis virus was evaluated by an in-silico analysis using viral genome RNA sequences of these two viruses available in GenBank. There was no homology found between the primers and probes of the miDiagnostics HSV 1 &2 CSF test to either of the virus.

Microbial interference

The ability of the miDiagnostics HSV-1&2 CSF Test to detect HSV-1 and HSV-2 in the presence of other potentially inhibitory microorganisms was evaluated. A total of 46 microorganisms were spiked either individually or as a pool into HSV-1 and HSV-2 positive CSF (spiked at 3x LOD concentration). No interference was observed for either HSV-1 or HSV-2. The microorganisms and the tested concentration of each of them are listed in Table 4 and are the same that were evaluated for cross-reactivity.

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Interfering Substances

The potential interference of substances that may be present in CSF specimens was evaluated by testing eleven substances at the concentrations listed (above normal clinical concentrations) in the table below. Negative CSF was spiked with HSV-1 and HSV-2 at 3x LOD and was tested in triplicate for each substance individually. No interference was observed for any of the substances at the concentrations listed in Table 5.

Table 5: Summary table of potential interfering substances tested with miDiagnostics HSV-1&2 CSF Test.

Potential interfering substance	Interferent concentration	Clinically relevant concentration
Albumin	10 mg/mL	0.32-2.63 mg/mL
Lactate (L-, sodium salt)	2.2 mg/mL	0-0.6 mg/mL
D(+) Glucose	9.9 mg/mL	0.32-0.98 mg/mL
Casein	5 mg/mL	0.15-0.40 mg/mL
Hemoglobin	0.625 mg/mL	<0.03 mg/mL
Immunoglobulin (IgG)	10 mg/mL	0.8 mg/mL
White Blood Cells (WBC)	2 x 106 cells/ mL	0.009- 1 x 106 cells/mL
Whole Blood	5% (v/v)	<1% (v/v)
Betadine®	1% (w/v)	0.25% (v/v)
Acyclovir	2.5 mg/mL	0.022 mg/mL
Acyclovir Triphosphate	0.5 mg/mL	Metabolized from up to 0.022 mg/mL

Competitive Interference

The interference between HSV targets of the miDiagnostics HSV-1&2 CSF Test was evaluated by spiking one of the targets at high concentration (105 TCID50/mL) and the other target at low concentration (3x LOD) into negative CSF. No interference between the targets was observed.

Carry-over Contamination

The risk of carry-over contamination from one test to another was evaluated by alternately testing high positive sample (HSV-1, at ≥ 105 pfu/mL) followed by negative sample on a single Reader. Five operators processed a high positive sample followed by a negative sample, for a total of eight times per Reader across five different Readers. No carry-over contamination was observed throughout the study.

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Within-Laboratory Precision

The miDiagnostics HSV-1&2 CSF Test within-laboratory precision was evaluated including Intermediate precision (within lab) and within run repeatability. The within-laboratory precision study included 3 panel members 2x LoD (low positive), 5x LoD (moderate positive) and negative samples. The positive panel members were prepared by spiking HSV-1 (HF strain) and HSV-2 (MS strain) stocks into pooled negative human CSF matrix.

Each panel member was tested in 2 replicates by 2 operators on 2 miDiagnostics PCR Platforms for 12 days, with 2 runs per day per operator, using 2 different lots of reagents for a total of 96 replicates of each panel member (2 replicates x 2 runs per day x 2 operators x 12 days = 96 total replicates).

The within-laboratory precision results for HSV-1 and HSV-2 are summarized below in Table 6.

Table 6: Within-laboratory precision summary table of miDiagnostics HSV-1&2 CSF Test for HSV-1 and HSV-2

Analyte	Sample	Rate of Detection	Mean Cq	Within Run	Between Run	Between Day	Within Laboratory^a	Between Operator	Total^b
				SD	% CV	SD	% CV	SD	%CV
HSV-1	2x LoD	96/96 (100%)	34.84	0.61	1.75	1.06	3.05	0.00	0.00
	5x LoD	96/96 (100%)	32.94	0.51	1.54	0.98	2.97	0.00	0.00
	Negative	NA	NA	NA	NA	NA	NA	NA	NA
HSV-2	2x LoD	95/96 (98.96%)	35.92	0.79	2.19	0.99	2.75	0.00	0.00
	5x LoD	96/96 (100%)	34.53	0.71	2.07	1.11	3.21	0.00	0.00
	Negative	NA	NA	NA	NA	NA	NA	NA	NA

^a Within Laboratory includes Within Run, Between Run, and Between Day Components
^b Total includes Within Run, Between Run, Between Day, and Between Operator Components.

Reproducibility Study (multi-site precision)

The reproducibility of miDiagnostics HSV-1&2 CSF Test was evaluated by testing a 4 panel member panel, including a 2x LoD (low positive), 5x LoD (moderate positive), 10x LoD (high positive) and 1x LoD (high negative). The positive panel members were prepared by spiking HSV-1 (HF strain) and HSV-2 (MS strain) stocks into pooled negative human CSF matrix. Each panel member was tested with 3 replicates at 3 sites, 2 operators for 5 days (3 replicates x 3 sites x 2 operators x 5 days = 90 total replicates). The study was carried out at three sites (two external and one internal) with 2 operators, 2 lots of reagents, and 3 miDiagnostics PCR Platforms per site across 5 non-consecutive days. Each panel member was tested with 3 replicates at 3 sites, 2 operators for 5 days (3 replicates x 3 sites x 2 operators x 5 days = 90 total replicates). The reproducibility results of the miDiagnostics HSV-1&2 CSF Test for HSV-1 and HSV-2 are summarized below in the Table 7.

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Table 7: Reproducibility summary table of miDiagnostics HSV-1&2 CSF Test for HSV-1 and HSV-2

Analyte	Panel Member	Rate of Detection	Mean Cq	Within Run	Between Run	Between Day	Between Lot	Between Site	Total^a
				SD	% CV	SD	% CV	SD	% CV
HSV-1	2x LoD	90/90 (100%)	35.16	1.06	3.02	0.58	1.65	0.00	0.00
	5x LoD	90/90 (100%)	33.41	0.96	2.87	0.00	0.00	0.52	1.56
	10x LoD^b	88/88 (100%)	31.48	0.83	2.63	0.00	0.00	0.53	1.69
	High Negative	50/90 (55.56%)	38.16	1.34	3.50	1.20	3.15	0.00	0.00
HSV-2	2x LoD	90/90 (100%)	36.36	1.62	4.46	0.00	0.00	0.33	0.91
	5x LoD	90/90 (100%)	34.56	0.79	2.28	0.95	2.74	0.17	0.49
	10x LoD	88/88 (100%)	32.74	0.85	2.59	0.00	0.00	0.61	1.88
	High Negative	33/90 (36.67%)	38.72	2.11	5.45	0.00	0.00	0.00	0.00

^a Total includes Within Run/Day, Between Run/Day, Between Lot, and Between-Site Components.
^b For site 1 and site 2, one replicate of the high positive sample was excluded from the analysis due to operator errors.

Clinical performance characteristics

Performance characteristics of the miDiagnostics HSV-1&2 CSF Test were established in prospective (Cohort 1) and retrospective (Cohort 2) clinical studies conducted at 4 participating testing sites in the United States and United Kingdom. The specimens were tested with the miDiagnostics HSV-1&2 CSF Test and the comparator method.

A total of 362 specimens were enrolled and tested from August 2024 to December 2024 at 4 participating testing sites to evaluate the clinical performance of the miDiagnostics HSV-1&2 Test. A total of 325 specimens were included for the clinical performance of the miDiagnostics HSV-1&2 CSF Test. Thirtyseven (37) specimens did not meet the inclusion criteria and were excluded from the studies. The CSF specimens of individuals of all sexes, all ethnicities and all ages, with signs and/or symptoms of HSV-1 or HSV-2 infection in the CNS, that were eligible according to the inclusion and exclusion criteria, were enrolled. The CSF specimens were either collected prospectively (Cohort 1) or originating from a biorepository, and thus retrospective (Cohort 2), and each CSF specimen originated from a unique subject.

The gender and age demographic of individuals for prospective and retrospective studies are presented in Table 8 and Table 9 below.

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Table 8: Prospective Study - Age and Gender Distribution of individuals

Age Category	Female				Male
	HSV-1	HSV-2	Not detected	Total	HSV-1	HSV-2	Not detected	Total
From birth to 1 year	0	0	4	4	0	0	3	3
>1 to 2 years	0	0	2	2	0	0	0	0
>2 to 12 years	0	0	3	3	0	0	2	2
>12 to 21 years	0	0	4	4	0	0	4	4
>21 to 60 years	1	1	41	43	1	1	35	37
>60 years	0	0	35	35	0	1	35	36
Total	1	1	89	91	1	2	79	82

Table 9: Retrospective Study - Age and Gender Distribution of individuals

Age Category	Female				Male
	HSV-1	HSV-2	Not detected	Total	HSV-1	HSV-2	Not detected	Total
From birth to 1 year	0	0	2	2	0	0	2	2
>1 to 2 years	0	0	0	0	0	0	0	0
>2 to 12 years	0	0	2	2	0	0	1	1
>12 to 21 years	0	1	2	3	0	1	2	3
>21 to 60 years	1	19	34	54	4	5	24	33
>60 years	4	3	16	23	10	2	17	29
Total	5	23	56	84	14	8	46	68

Performance of the miDiagnostics HSV-1&2 Test is presented as Positive Percent Agreement (PPA) and Negative Percent Agreement (NPA) with the comparator method in the tables 10 to 13 below.

Table 10: HSV-1 Prospective Samples.

Candidate Device	Comparator Method		Total
	Detected	Not Detected
Detected	0	2*	2
Not Detected	0	171	171
Total	0	173	173

PPA: NA
NPA: 171/173 (98.84%); (95% CI: 95.88% - 99.68%)

* Two false positive specimens were reported to be HSV-1 negative by Standard of Care PCR testing. No clinical diagnoses were available.

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Table 11: HSV-1 Retrospective Samples.

Candidate Device	Comparator Method		Total
	Detected	Not Detected
Detected	16	3*	19
Not Detected	0	133	133
Total	16	136	152

PPA: 16/16 (100.00%); (95% CI: 80.64% - 100.00%)
NPA: 133/136 (97.79%); (95% CI: 93.72% - 99.25%)

* All 3 false positive specimens were reported to be HSV-1 negative by Standard of Care PCR testing. Two of them were clinically diagnosed with HSV-1 infection, and one subject was clinically diagnosed with Herpes Meningoencephalitis.

Table 12: HSV-2 Prospective Samples

Candidate Device	Comparator Method		Total
	Detected	Not Detected
Detected	2	1*	3
Not Detected	0	170	170
Total	2	171	173

PPA: 2/2 (100.00%); (95% CI: 34.24% - 100.00%)
NPA: 170/171 (99.42%); (95% CI: 96.76% - 99.90%)

* One false positive specimen was reported to be HSV-2 negative by Standard of Care PCR testing. No clinical diagnosis was available.

Table 13: HSV-2 Retrospective Samples

Candidate Device	Comparator Method		Total
	Detected	Not Detected
Detected	29	2*	31
Not Detected	0	121	121
Total	29	123	152

PPA: 29/29 (100.00%); (95% CI: 88.31% - 100.00%)
NPA: 121/123 (98.37%); (95% CI: 94.27% - 99.55%)

* The 2 false positive specimens were reported to be HSV-2 positive by Standard of Care PCR testing. One subject was clinically diagnosed with Herpes Meningitis. No clinical diagnosis was available for the other subject.

CONCLUSION STATEMENT

The analytical and clinical study results demonstrate that miDiagnostics HSV-1&2 CSF Test on the miDiagnostics PCR Platform performs comparably to the predicate device in detecting HSV-1 & 2 DNA and supports a substantial equivalence decision.

The information submitted in this premarket notification is complete and supports a substantial equivalence decision.

Regulation Number and Section

§ 866.3307 Herpes simplex virus nucleic acid-based assay for central nervous system infections.

(a)
Identification. A herpes simplex virus nucleic acid-based assay for central nervous system infections is a qualitative in vitro diagnostic device intended for the detection and differentiation of HSV-1 and HSV-2 in cerebrospinal fluid (CSF) samples from patients suspected of Herpes Simplex Virus (HSV) infections of the central nervous system (CNS). This test is intended as an aid in the diagnosis of HSV-1 and HSV-2 infections of the CNS. Negative results do not preclude HSV-1 or HSV-2 infection 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) Design verification and validation must include:
(i) Detailed documentation for the device description, including the device components, ancillary reagents required but not provided, and a detailed explanation of the methodology, including primer design and selection.
(ii) Detailed documentation from the following analytical and clinical performance studies: Analytical sensitivity (limit of detection), reactivity, inclusivity, precision, reproducibility, interference, cross reactivity, carryover, and cross contamination. Documentation must include reagent and sample stability recommendations.
(iii) Detailed documentation from a clinical study. The study, performed on a study population consistent with the intended use population, must compare the device performance to the results of two polymerase chain reaction methods followed by bidirectional sequencing.
(iv) Documentation of an appropriate end user device training program that will be offered as part of efforts to mitigate the risk of failure to correctly operate the instrument.
(v) Quality assurance protocols and detailed documentation for device software, including standalone software applications and hardware-based devices that incorporate software.
(2) The labeling required under § 809.10(b) of this chapter must include:
(i) A detailed explanation of the interpretation of results and acceptance criteria.
(ii) A limiting statement indicating that negative results do not preclude HSV-1 or HSV-2 infection and should not be used as the sole basis for treatment or other patient management decisions.