(212 days)
Not Found
No
The summary describes a mass spectrometry-based assay and associated software for quantitative measurement of enzyme activity. There is no mention of AI or ML in the device description, intended use, performance studies, or key metrics. The software mentioned (MassLynx and NeoLynx) are standard data acquisition and processing software for mass spectrometry, not AI/ML platforms.
No.
This device is for the quantitative measurement of enzyme activity in dried blood spots to aid in screening newborns for certain lysosomal storage disorders; it is a diagnostic device, not a therapeutic one.
Yes
The intended use explicitly states that the analysis of enzymatic activity is "intended as an aid in screening newborns for the following lysosomal storage disorders (LSD) respectively; Gaucher Disease, Nieman-Pick A/B Disease, Pompe Disease, Fabry Disease, and MPS I Disease." Screening for diseases falls under the definition of a diagnostic device.
No
The device description explicitly lists hardware components such as a Waters TQD MSMS instrument, sample pump, autosampler, power cables, tubing, syringes, and connection cables, in addition to software.
Based on the provided information, the NeoLSD MSMS Kit is an IVD (In Vitro Diagnostic).
Here's why:
- Intended Use: The intended use explicitly states that the kit is for the "quantitative measurement of the activity of the enzymes... in dried blood spots (DBS) from newborn babies." This measurement is intended as an "aid in screening newborns for the following lysosomal storage disorders (LSD)". This clearly indicates that the device is used to examine specimens derived from the human body (dried blood spots) to provide information for the diagnosis or screening of a disease or condition (LSDs).
- Device Description: The description details a system that analyzes biological samples (dried blood spots) using chemical reactions (enzyme activity with substrates) and a mass spectrometry instrument to produce quantitative results. This aligns with the definition of an in vitro diagnostic device.
- Patient Population: The device is intended for use with specimens from "newborn babies," a specific patient population.
- Intended User/Care Setting: The intended user is a "Newborn screening laboratory," which is a clinical laboratory setting where in vitro diagnostic tests are performed.
All these factors strongly indicate that the NeoLSD MSMS Kit is an In Vitro Diagnostic device.
N/A
Intended Use / Indications for Use
The NeoLSD MSMS Kit is intended for the quantitative measurement of the activity of the enzymes acid-pglucocerebrosidase (ABG), acid-sphingomyelinase (ASM), acid-a-glucosidase (GAA), B-galactocerebrosidase (GALC), α-galactosidase A (GLA) and α-L-iduronidase (IDUA) in dried blood spots (DBS) from newborn babies. The analysis of the enzymatic activity is intended as an aid in screening newborns for the following lysosomal storage disorders (LSD) respectively; Gaucher Disease, Nieman-Pick A/B Disease, Pompe Disease, Fabry Disease, and MPS I Disease.
Product codes (comma separated list FDA assigned to the subject device)
POW, POT, POT, POV, OCL, OCM
Device Description
The NeoLSD MSMS test system uses mass spectrometry to quantitatively measure the activity of six lysosomal enzymes simultaneously from a dried blood spot sample. The NeoLSD MSMS test system is comprised of: NeoLSD MSMS kit, including substrates, internal standards, solutions and controls; Waters TQD MSMS instrument comprised of a Waters 1525 sample pump, Waters 2777c autosampler, Waters MassLynx v4.1 firmware, power cables, tubing, syringes, connection cables; Waters NeoLynx v4.1 software and computer with monitor; PerkinElmer MSMS Workstation Software. The NeoLSD MSMS kit evaluates enzyme activities by measuring the product generated when an enzyme reacts with a synthesized substrate to create a specific end product. The activities of the six lysosomal enzymes present in a 3.2 mm punch from a dried blood spot (DBS) are simultaneously measured by the NeoLSD MSMS kit. The punches are incubated with the assay reagent mixture which contains: six substrates, one corresponding to each lysosomal enzyme; six stable-isotope mass-labeled internal standards (IS) each designed to chemically resemble each product generated; a buffer to maintain the reaction pH, and to carry inhibitors to limit activity from competing enzymes if present and additives to enhance the targeted enzyme reactions.
Mentions image processing
Not Found
Mentions AI, DNN, or ML
Not Found
Input Imaging Modality
Not Found
Anatomical Site
Dried blood spots (DBS) from newborn babies.
Indicated Patient Age Range
Newborn babies
Intended User / Care Setting
Not Found
Description of the training set, sample size, data source, and annotation protocol
Not Found
Description of the test set, sample size, data source, and annotation protocol
A prospective clinical study of four-year-old retrospective routine samples was conducted at an EU newborn screening laboratory. Over the two-month study duration, 4041 newborn samples were tested from newborns ≤ 29 days old. All 4011 routine samples were 4 years old, enabling the follow-up of clinical status at 4 years. Due to the low incidence of LSDs, the study population was enriched with confirmed LSD positive newborn DBS specimens obtained from the site's biobank, these samples ranged from 5.8 to 17.6 years of age. Sample inclusion criteria were the same as in the reference range study.
The 4011 routine samples were tested according to the defined algorithm to determine the enzyme activities for sample interpretation. Site specific cut-off percentages determined prior to the clinical study were applied as percent of daily median activity. The routine specimens having enzyme activities below the cut-off values in the initial testing were re-tested in duplicate. The final results (normal, presumptive positive, invalid result) were classified after the initial testing for samples above the cut-off values and after the final testing for samples below the initial cut-off level.
Confirmed LSD positive newborn specimens were tested once, and assessed using the retest cut-off values for the enzymes. Given the known status of the confirmed LSD positive newborn specimens, processing the samples through the full testing algorithm provided no additional information nor changed the sample interpretation.
Clinical outcome was used as a comparator for all samples, including the 4011 routine screening samples, as derived from the civil registry status and national hospital registry. Subject's survival at 4 years of age without LSD diagnosis or clinical signs suggestive of an LSD was used as clinical confirmation of an unaffected newborn. Subjects who did not have follow-up information available up to 4 years were classified as lost to follow-up.
Summary of Performance Studies (study type, sample size, AUC, MRMC, standalone performance, key results)
Reproducibility Study:
- Study Type: Reproducibility study.
- Sample Size: 75 determinations for each level of a six-member panel for each enzyme across three sites.
- Key Results: Reproducibility, between- and within-laboratory precisions were determined for ABG, ASM, GALC, IDUA, GLA, and GAA. The overall reproducibility CV% ranged from 5.6% to 21.0%.
Limit of Blank, Limit of Detection, and Limit of Quantitation Study:
- Study Type: Determination of analytical limits according to CLSI document EP17-A2.
- Sample Size: 120 determinations for LoB, 150 determinations for LoD for the TQD MSMS instrument.
- Key Results:
- LoB (µmol/L/h): ABG (0.165), ASM (0.110), GALC (0.106), IDUA (0.059), GLA (0.476), GAA (0.073).
- LoD (µmol/L/h): ABG (0.63), ASM (0.27), GALC (0.34), IDUA (0.24), GLA (0.97), GAA (0.39).
- LoQ (µmol/L/h): ABG (0.69), ASM (0.90), GAA (0.63), GALC (0.34), GLA (0.97), IDUA (0.44).
- CV% at LoQ: ABG (21.7%), ASM (20.0%), GAA (17.5%), GALC (20.6%), GLA (17.5%), IDUA (18.2%).
Linearity Study:
- Study Type: Linearity determination according to CLSI document EP06-A.
- Key Results: The method was demonstrated to be linear within the following ranges:
- ABG: 0.69 - 20.1 µmol/L/h
- ASM: 0.90 - 20.5 µmol/L/h
- GALC: 0.63 - 6.3 µmol/L/h
- IDUA: 0.34 - 17.2 µmol/L/h
- GLA: 0.97 - 20.9 µmol/L/h
- GAA: 0.44 - 24.2 µmol/L/h
Interference Study:
- Study Type: Evaluation of interference according to CLSI document EP07-A2.
- Key Results:
- Metabolites and drugs found not to interfere at tested concentrations.
- Identified interferences:
- Glucose interferes with GAA activity (decreases measured activity).
- Hematocrit interferes with ABG (decreases at 35%, increases at 65%) and GALC (increases at 35%, decreases at 65%).
- Hemoglobin increases ABG, ASM, and IDUA activity. Concluded not pronounced enough to impair separation of affected/unaffected.
- Triglyceride (Intralipid) increases GAA, GLA, and IDUA activity.
- EDTA interferes with ABG (increases) and ASM (decreases).
Reference Range Study:
- Study Type: Prospective clinical study using retrospective routine newborn screening samples to establish cut-off values.
- Sample Size: 5041 newborn samples from 0-30 days of age at a European site. Additional 5251 samples from ≤ 4 days and 5053 samples from ≤ 7 days at US sites A and B respectively.
- Key Results: Established cut-off values based on percentiles of enzyme activity distribution for ABG, ASM, GALC, IDUA, GLA, and GAA. Descriptive statistics (mean, median, percentiles) were provided for all enzymes for the EU and two US sites.
Screening Performance Study:
- Study Type: Prospective clinical study of four-year-old retrospective routine samples.
- Sample Size: 4041 specimens (4011 routine newborn specimens + 30 confirmed positive LSD cases).
- Key Results:
- Performance estimates (invalid and lost-to-follow-up excluded) for all 6 enzymes:
- Sensitivity: 92.9 % (76.5%-99.1%)
- False-negative rate: 7.1%* (0.9% - 23.5%) *includes 2 Fabry females
- Specificity: 99.4 % (99.1% - 99.6%)
- False-positive rate: 0.6 % (0.4% - 0.9%)
- With female Fabry subjects excluded, there were no false negative results for any enzyme.
- False positive rates for individual enzymes: ABG (0.07%), ASM (0%), GALC (0.10%), IDUA (0.07%), GLA (0.27%), GAA (0.10%).
- Retest rate: 6.6%, with 2.2% due to test failure and 4.4% due to screening algorithm.
- Performance estimates (invalid and lost-to-follow-up excluded) for all 6 enzymes:
Key Metrics (Sensitivity, Specificity, PPV, NPV, etc.)
Performance estimates (invalid and lost-to-follow-up excluded) for all 6 enzymes:
Sensitivity: 92.9 % (76.5%-99.1%)
False-negative rate: 7.1%* (0.9% - 23.5%) (* includes 2 Fabry females)
Specificity: 99.4 % (99.1% - 99.6%)
False-positive rate: 0.6 % (0.4% - 0.9%)
With the female Fabry subjects excluded the NeoLSD MSMS test system has no false negative results for any of the enzymes; and a false positive range of ABG (0.07%), ASM (0%), GALC (0.10%), IDUA (0.07 %), GLA (0.27 %), GAA ( 0.10%).
Predicate Device(s): If the device was cleared using the 510(k) pathway, identify the Predicate Device(s) K/DEN number used to claim substantial equivalence and list them here in a comma separated list exactly as they appear in the text. List the primary predicate first in the list.
Baebies SEEKER 4-Plex Assay (IDUA/GAA/GBA/GLA) (DEN150035)
Reference Device(s): Identify the Reference Device(s) K/DEN number and list them here in a comma separated list exactly as they appear in the text.
Not Found
Predetermined Change Control Plan (PCCP) - All Relevant Information for the subject device only (e.g. presence / absence, what scope was granted / cleared under the PCCP, any restrictions, etc).
Not Found
§ 862.1488 Lysosomal storage disorder newborn screening test system.
(a)
Identification. A lysosomal storage disorder newborn screening test system is intended to measure lysosomal enzyme levels obtained from dried blood spot specimens on filter paper from newborns as an aid in screening newborns for a lysosomal storage disorder.(b)
Classification. Class II (special controls). The special controls for this device are:(1) Design verification and validation must include information that demonstrates the performance characteristics of the device, including:
(i) Study results that adequately demonstrate the clinical validity of the device, which must include information supporting the link between the analyte being measured and the condition being screened. The clinical validity of the device must be demonstrated in a clinical validation study using either well-characterized prospectively or retrospectively obtained clinical specimens from the intended use population. Testing in the clinical validation study must be performed by operators representative of the types of operators intended to use the test. The study design of the clinical validation study must assess the effects of sample collection and processing steps on test performance. Confirmed positive specimens must have a diagnosis based on confirmatory diagnostic methods or clinically meaningful information regarding the status of the subject must be obtained.
(ii) The reference interval in the normal newborn population for the analyte or analytes measured by the device.
(iii) Study results demonstrating the level of carryover or drift affecting the device performance.
(iv) Study results demonstrating the concentrations of the limit of blank, limit of detection, and limit of quantitation of the device. Sample concentrations below the limit of quantitation must not be reported by the device.
(v) Study results, which must be collected using sample panels from at least three reagent lots and at least three instruments over more than 20 testing days, demonstrating the imprecision of the device. The sample panels must consist of blood spot specimens with a range of analyte concentrations that span the reportable range of the device and must include samples with concentrations in the screen positive range, samples with concentrations at each cutoff, and samples with concentration in the normal range.
(2) The labeling required under § 809.10(b) of this chapter must include:
(i) A warning that indicates that the test is not intended to diagnose lysosomal storage disorders.
(ii) A warning that indicates that test results are intended to be used in conjunction with other clinical and diagnostic findings, consistent with professional standards of practice, including confirmation by alternative methods, and clinical evaluation as appropriate.
(iii) Detailed information on device performance, including the false positive rate and the false negative rate observed in the clinical study.
(iv) Information on device performance in any relevant subgroup (
e.g., age of newborn at time of sample collection, birth weight, sex, gestational age, race, ethnicity) observed in the clinical study.
0
Image /page/0/Picture/0 description: The image shows the logo of the U.S. Food and Drug Administration (FDA). On the left is the Department of Health & Human Services logo. To the right of that is the FDA logo, which consists of the letters "FDA" in a blue square, followed by the words "U.S. FOOD & DRUG" in blue, with the word "ADMINISTRATION" underneath in a smaller font.
July 18, 2018
Wallac Oy, a subsidiary of PerkinElmer Kay Taylor VP, Regulatory, Quality, Medical & Scientific Affairs 940 Winter Street Waltham, MA 02451
Re: K173829
Trade/Device Name: NeoLSD MSMS kit Regulation Number: 21 CFR 862.1488 Regulation Name: Lysosomal storage disorder newborn screening test system Regulatory Class: Class II Product Code: POW, POT, POT, POV, OCL, OCM Dated: July 9, 2018 Received: July 10, 2018
Dear Kay Taylor:
We have reviewed your Section 510(k) premarket notification of intent to market the device referenced above and have determined the device is substantially equivalent (for the indications for use stated in the enclosure) to legally marketed predicate devices marketed in interstate commerce prior to May 28, 1976, the enactment date of the Medical Device Amendments, or to devices that have been reclassified in accordance with the provisions of the Federal Food. Drug, and Cosmetic Act (Act) that do not require approval of a premarket approval application (PMA). You may, therefore, market the device, subject to the general controls provisions of the Act. The general controls provisions of the Act include requirements for annual registration, listing of devices, good manufacturing practice, labeling, and prohibitions against misbranding and adulteration. 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 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); good manufacturing practice requirements as set forth in the quality systems (QS) regulation (21 CFR
1
Part 820); 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 http://www.fda.gov/MedicalDevices/Safety/ReportaProblem/default.htm.
For comprehensive regulatory information about medical devices and radiation-emitting products, including information about labeling regulations, please see Device Advice (https://www.fda.gov/MedicalDevices/DeviceRegulationandGuidance/) and CDRH Learn (http://www.fda.gov/Training/CDRHLearn). 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 (http://www.fda.gov/DICE) for more information or contact DICE by email (DICE@fda.hhs.gov) or phone (1-800-638-2041 or 301-796-7100).
Sincerely.
Kellie B. Kelm -S
for Courtney H. Lias, Ph.D. Director Division of Chemistry and Toxicology Devices Office of In Vitro Diagnostics and Radiological Health Center for Devices and Radiological Health
Enclosure
2
Indications for Use
510(k) Number (if known) K173829
Device Name
NeoLSD MSMS kit
Indications for Use (Describe)
The NeoLSD MSMS Kit is intended for the quantitative measurement of the activity of the enzymes acid-pglucocerebrosidase (ABG), acid-sphingomyelinase (ASM), acid-a-glucosidase (GAA), B-galactocerebrosidase (GALC), α-galactosidase A (GLA) and α-L-iduronidase (IDUA) in dried blood spots (DBS) from newborn babies. The analysis of the enzymatic activity is intended as an aid in screening newborns for the following lysosomal storage disorders (LSD) respectively; Gaucher Disease, Nieman-Pick A/B Disease, Pompe Disease, Fabry Disease, and MPS I Disease.
| 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) |
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3
510(K) SUMMARY
This 510k Summary information is supplied in accordance with the requirements of SMDA 1990 and 21 CFR 807.92.
The assigned 510 (k) number is K173829
Date: July 18, 2018
Submitted by: PerkinElmer Inc. 940 Winter Street Waltham, MA 02451
Wallac Oy, a subsidiary of PerkinElmer Inc. Mustionkatu 6 Turku, Finland 20750
Kay A. Taylor Contact Person: Tel: 317 418-1735
Trade Name: NeoLSD MSMS Kit
Common Name: NeoLSD MSMS Kit
Regulation: 21 CFR 862.1488
Classification Name: Lysosomal storage disorder newborn screening test system
Classification: 75 Chemistry
Product Code: PQW, PQT, PQU, PQV, QCL, QCM
Predicate device: Baebies SEEKER 4-Plex Assay (IDUA/GAA/GBA/GLA) (DENT50035)
4
Device Description:
The NeoLSD MSMS test system uses mass spectrometry to quantitatively measure the activity of six lysosomal enzymes simultaneously from a dried blood spot sample. The NeoLSD MSMS test system is comprised of:
-
- NeoLSD MSMS kit, including substrates, internal standards, solutions and controls
The NeoLSD MSMS Kit will contain sufficient reagents and consumables to perform 960 assays (10 x 96-well plates) as listed in the following table.
- NeoLSD MSMS kit, including substrates, internal standards, solutions and controls
| Component | Description |
|---|---|
| Kit insert | Instructions for use |
| QC certificate | QC certificate showing the kit lot specific Kit Control results determined by |
| the manufacturer, and the 1 SD limits. | |
| Internal Standards Substrate Mix | 1 vial or several vials of stable-isotope standards and designed substrates, |
| The dried substrates and internal standards are a mixture of the 6 synthetic | |
| substrates, the corresponding 6 stable-isotope labeled internal standards, | |
| and sodium oleate | |
| DBS controls | C1, C2, C3 control levels on DBS cassettes, manufactured from human blood |
| with a hematocrit value of 45-50%. | |
| Assay buffer | 1 bottle of 40 mL buffer, ready-for-use succinate buffered (pH 4.7) salt |
| solution | |
| Extraction Solution | Ethyl acetate |
| Flow Solvent reconstitution | |
| solvent | The ready-for-use Flow Solvent contains acetonitrile, water, and formic |
| acid. | |
| Incubation/Sampling plate | 20 x 96-well microplate, U-bottomed |
| Extraction plate | 10 x 96-deep well microplate |
| Aluminum foil microplate covers | 20 adhesive aluminum foil microplate covers |
| Microplate covers | 10 x adhesive microplate covers |
| Plate barcode labels | 30 x plate barcodes |
-
- Waters TQD MSMS instrument comprised of,
- a. Waters 1525 sample pump
- b. Waters 2777c autosampler
- Waters MassLynx v4.1 firmware C.
- d. Power cables, tubing, syringes, connection cables
5
-
- Waters NeoLynx v4.1 software and computer with monitor
-
- PerkinElmer MSMS Workstation Software
The NeoLSD MSMS kit evaluates enzyme activities by measuring the product generated when an enzyme reacts with a synthesized substrate to create a specific end product. The activities of the six lysosomal enzymes present in a 3.2 mm punch from a dried blood spot (DBS) are simultaneously measured by the NeoLSD MSMS kit. The punches are incubated with the assay reagent mixture which contains;
- . six substrates, one corresponding to each lysosomal enzyme
- . six stable-isotope mass-labeled internal standards (IS) each designed to chemically resemble each product generated
- . a buffer to maintain the reaction pH, and to carry inhibitors to limit activity from competing enzymes if present and additives to enhance the targeted enzyme reactions.
6
Comparison Chart:
Comparison of the NeoLSD MSMS Kit with the predicate.
| NeoLSD MSMS Kit | ||
|---|---|---|
| Characteristics | Proposed Device | Predicate (DEN150035) |
| Intended | ||
| Use/Indications for | ||
| Use | The NeoLSD MSMS Kit is intended for the quantitative | |
| measurement of the activity of the enzymes acid-β- | ||
| glucocerebrosidase (ABG), acid-sphingomyelinase (ASM), | ||
| acid-α-glucosidase (GAA), β-galactocerebrosidase (GALC), α- | ||
| galactosidase A (GLA) and α-L-iduronidase (IDUA) in dried | ||
| blood spots (DBS) from newborn babies. The analysis of the | ||
| enzymatic activity is intended as an aid in screening newborns | ||
| for the following lysosomal storage disorders (LSD) | ||
| respectively; Gaucher Disease, Niemann-Pick A/B Disease, | ||
| Pompe Disease, Krabbe Disease, Fabry Disease, and MPS I | ||
| Disease. | The SEEKER System, including the SEEKER Instrument and the | |
| SEEKER LSD Reagent Kit-IDUA GAA GBA GLA for use on the SEEKER | ||
| Instrument, is intended for quantitative measurement of the | ||
| activity of α-L-iduronidase, α-D-glucosidase, β-glucocerebrosidase | ||
| and α-D-galactosidase A from newborn dried blood spot specimens | ||
| as an aid in screening newborns for Mucopolysaccharidosis Type I, | ||
| Pompe, Gaucher and Fabry diseases. Reduced activity of these | ||
| enzymes may be indicative of these lysosomal storage diseases. The | ||
| enzymes measured using the SEEKER LSD Reagent Kit- | ||
| IDUA GAA GBA GLA and their associated lysosomal storage | ||
| diseases are listed below.. | ||
| Enzyme (abbreviation) Disease | ||
| α-L-iduronidase (IDUA) Mucopolysaccharidosis Type I (MPS I) | ||
| α-D-glucosidase (GAA) Pompe | ||
| β-glucocerebrosidase (GBA) Gaucher | ||
| α-D-galactosidase A (GLA) Fabry | ||
| Test Methodology | Quantitative mass spectrometric enzymatic activity assay | Quantitative fluorimetric enzymatic activity assay |
| Instrument / | ||
| Software Platform | Waters TQD instrument with MassLynx v4.1 firmware, with | |
| Waters 1525 sample pump, with Waters 2777c autosampler, | ||
| with Waters NeoLynx v4.1 software and with the PerkinElmer | ||
| MSMS Workstation Software | Seeker Instrument with Spot Logic Software | |
| Sample Type | Same | Punch from dried blood spot specimen |
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| Characteristics | Proposed Device | Predicate (K071649) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Reportable Range | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| (µmol/L/h) | IDUA: 0.34 – 17.2 | IDUA: 2.77 – 50.75 µmol/L/h | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| GAA: 0.44 – 24.2 | GAA: 2.18 – 94.66 µmol/L/h | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| ABG: 0.69 – 20.1 (identified as GBA in predicate) | GBA: 2.14 – 73.24 µmol/L/h | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| GLA: 0.97 – 20.9 | GLA: 4.88 – 153.74 µmol/L/h | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| ASM: 0.90 – 20.5 | ASM: not detected by predicate | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| GALC: 0.63 – 6.3 | GALC: not detected by predicate | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Lower Limits of | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Measure (µmol/L/h) | IDUA: LoB=0.059, LoD=0.24, LoQ=0.34 | IDUA (µmol/L/h): LoB (1.78), LoD (2.77), LoQ (2.77) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| GAA: LoB=0.073, LoD=0.39, LoQ=0.44 | GAA (µmol/L/h): LoB (0.50), LoD (2.18), LoQ (2.18) | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| ABG: LoB=0.165, LoD=0.63, LoQ=0.69 | GBA (µmol/L/h): LoB (0.72), LoD (1.07), LoQ (1.85) | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| GLA: LoB=0.476, LoD=0.97, LoQ=0.97 | GLA (µmol/L/h): LoB (1.96), LoD (3.18), LoQ (4.88) | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| ASM: LoB=0.110, LoD=0.27, LoQ=0.90 | ASM: not detected by predicate | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| GALC: LoB=0.106, LoD=0.34, LoQ=0.63 | GALC: not detected by predicate | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Calibrators / | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Standards | Molecular Weights & Concentrations of Internal Standards: | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| IDUA: 430.26 / 15.0 μΜ | Calibrant A: (0.0375 μΜ) | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| GAA: 502.32 / 24.0 μΜ | Calibrant B: (0.0750 μΜ) | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| ABG: 390.38 / 20.0 μΜ | Calibrant C: (0.1500 μΜ) | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| GLA: 488.31 / 24.0 μΜ | Calibrant D: (0.3000 μΜ) | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| ASM: 404.40 / 15.0 μΜ | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| GALC: 416.40 / 10.0 μΜ | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Controls | 3 levels of control material, human blood based | 4 levels of control material, human blood based | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Expected Values | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| (µmol/L/h) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| N 0.1% 0.2% 0.3% 2.5% 97.5% IDUA 5041 2.06 2.55 2.62 3.82 13.2 GAA 5041 2.33 2.69 2.92 4.28 17.5 ABG 5041 2.85 3.16 3.33 4.74 20.1 GLA 5041 3.04 3.37 3.62 4.81 25.8 ASM 5041 2.12 2.21 2.37 3.15 12.9 GALC 5041 0.43 0.52 0.56 0.95 9.34 | N 0.1% 0.5% 50% 99.5% 99.9% IDUA 102,399 3.68 5.63 18.37 46.23 >50.75 GAA 102,392 7.61 10.07 25.98 64.56 77.41 GBA 102,397 6.69 8.20 20.24 51.03 64.53 GLA 102,371 7.56 9.44 26.73 123.36 >153.74 |
8
| Subjects age 7-31 days | |||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|
| N | 0.1% | 0.5% | 50% | 99.5% | 99.9% | ||||||
| IDUA | 7,177 | 3.15 | 4.82 | 18.82 | >50.75 | >50.75 | |||||
| GAA | 7,177 | 5.04 | 7.75 | 21.53 | 73.01 | >94.66 | |||||
| GBA | 7,177 | 5.16 | 6.60 | 16:56 | 54.83 | >73.24 | |||||
| GLA | 7,176 | 5.14 | 6.39 | 19.70 | 96.19 | 134.27 | |||||
| Subjects age >14 days | |||||||||||
| N | 0.1% | 0.5% | ર૦% | 99.5% | 99.9% | ||||||
| IDUA | 7,447 | 2.83 | 3.93 | 19.41 | >50.75 | >50.75 | |||||
| GAA | 7,447 | 4.40 | 6.43 | 17.76 | 54.68 | 72.21 | |||||
| GBA | 7,447 | 4.31 | 5.83 | 14.61 | 45.21 | 64.54 | |||||
| GLA | 7,447 | 4.88 | 5.36 | 14.64 | 67.34 | 113.61 | |||||
| ASM: not detected by predicate | |||||||||||
| GALC: not detected by predicate | |||||||||||
| Reproducibility | |||||||||||
| Sample Activity | %CV Range | Sample Activity | %CV Range | ||||||||
| Range (µmol/L/h) | Range (µmol/L/h) | ||||||||||
| IDUA | 0.59 – 9.11 | 6.9 – 10.0 | IDUA | 3.53 - 24.06 | 14.2 — 28.5 | ||||||
| GAA | 1.29 — 10.77 | 5.6 - 9.4 | GAA | 4.29 – 27.37 | 12.0 – 17.0 | ||||||
| ABG | 1.03 – 8.08 | 13.0 – 21.0 | ABG | 2.84 – 15.00 | 15.7 — 38.0 | ||||||
| GLA | 1.04 — 12.49 | 8.6 - 15.7 | GLA | 6.94 – 52.66 | 9.4 — 16.3 | ||||||
| ASM | 2.47 – 9.11 | 7.6 - 11.4 | ASM: not detected by predicate | ||||||||
| GALC | 0.37 – 3.97 | 9.3 — 20.7 | GALC: not detected by predicate |
9
Summary of Studies:
Reproducibility:
Reproducibility of the NeoLSD MSMS assay was determined on three different TQD MSMS instrument at three sites, two external and one internal. The reproducibility is based on 75 determinations for each level of a six member panel for each enzyme: in each laboratory 5 plates were measured over 5 working days using one kit lot and each plate having 5 replicates per sample per enzyme. The results of reproducibility, between- and within-laboratory precisions are presented below.
| Enzyme | Sample | N | Mean | Within Laboratory | Between Laboratory | Reproducibility | |||
|---|---|---|---|---|---|---|---|---|---|
| SD | CV% | SD | CV% | SD | CV% | ||||
| ABG* | 2 | 75 | 1.03 | 0.14 | 13.8 | 0.16 | 15.8 | 0.22 | 21.0 |
| ABG* | 3 | 75 | 2.78 | 0.37 | 13.3 | 0.26 | 9.3 | 0.45 | 16.2 |
| ABG* | 4 | 75 | 7.46 | 0.89 | 12.0 | 0.77 | 10.3 | 1.18 | 15.8 |
| ABG* | 5 | 75 | 7.93 | 0.92 | 11.6 | 0.46 | 5.9 | 1.03 | 13.0 |
| ABG* | 6 | 75 | 8.08 | 1.03 | 12.8 | 0.38 | 4.7 | 1.10 | 13.6 |
| ASM* | 2 | 75 | 2.47 | 0.21 | 8.7 | 0.12 | 4.8 | 0.24 | 9.9 |
| ASM* | 3 | 75 | 3.93 | 0.43 | 11.0 | 0.11 | 2.9 | 0.45 | 11.4 |
| ASM* | 4 | 75 | 5.54 | 0.44 | 7.9 | 0.37 | 6.6 | 0.57 | 10.3 |
| ASM* | 5 | 75 | 7.29 | 0.56 | 7.6 | 0.47 | 6.4 | 0.73 | 10.0 |
| ASM* | 6 | 75 | 9.11 | 0.67 | 7.3 | 0.17 | 1.8 | 0.69 | 7.6 |
| GALC | 1 | 75 | 0.37 | 0.07 | 19.5 | 0.03 | 7.0 | 0.08 | 20.7 |
| GALC | 2 | 75 | 0.89 | 0.11 | 12.4 | 0.06 | 6.9 | 0.13 | 14.2 |
| GALC | 3 | 75 | 1.30 | 0.20 | 15.7 | 0.04 | 3.2 | 0.21 | 16.0 |
| GALC | 4 | 75 | 1.76 | 0.25 | 14.3 | 0.04 | 2.1 | 0.25 | 14.4 |
| GALC | 5 | 75 | 3.76 | 0.34 | 9.1 | 0.22 | 5.9 | 0.41 | 10.8 |
| GALC | 6 | 75 | 3.97 | 0.31 | 7.9 | 0.20 | 5.0 | 0.37 | 9.3 |
| IDUA* | 2 | 75 | 0.59 | 0.03 | 5.8 | 0.05 | 8.1 | 0.06 | 10.0 |
| IDUA* | 3 | 75 | 2.25 | 0.15 | 6.9 | 0.10 | 4.4 | 0.18 | 8.2 |
| IDUA* | 4 | 75 | 5.88 | 0.35 | 5.9 | 0.27 | 4.6 | 0.44 | 7.5 |
| IDUA* | 5 | 75 | 8.89 | 0.43 | 4.8 | 0.45 | 5.0 | 0.62 | 6.9 |
| IDUA* | 6 | 75 | 9.11 | 0.43 | 4.7 | 0.54 | 5.9 | 0.69 | 7.5 |
| GLA | 1 | 75 | 1.04 | 0.10 | 9.5 | 0.06 | 5.6 | 0.12 | 11.1 |
| GLA | 2 | 75 | 1.23 | 0.16 | 13.3 | 0.10 | 8.4 | 0.19 | 15.7 |
| GLA | 3 | 75 | 3.42 | 0.17 | 5.0 | 0.24 | 7.0 | 0.30 | 8.6 |
| GLA | 4 | 75 | 5.10 | 0.32 | 6.2 | 0.30 | 6.0 | 0.44 | 8.6 |
| GLA | 5 | 75 | 10.31 | 0.52 | 5.1 | 0.82 | 8.0 | 0.97 | 9.4 |
10
| 6 | 75 | 12.49 | 0.90 | 7.2 | 0.86 | 6.9 | 1.25 | 10.0 | |
|---|---|---|---|---|---|---|---|---|---|
| 2 | 75 | 1.29 | 0.07 | 5.5 | 0.10 | 7.6 | 0.12 | 9.4 | |
| 3 | 75 | 2.32 | 0.10 | 4.2 | 0.14 | 6.1 | 0.17 | 7.4 | |
| GAA* | 4 | 75 | 5.36 | 0.26 | 4.8 | 0.24 | 4.5 | 0.35 | 6.6 |
| 5 | 75 | 7.57 | 0.36 | 4.8 | 0.53 | 7.0 | 0.64 | 8.5 | |
| 6 | 75 | 10.77 | 0.46 | 4.3 | 0.38 | 3.5 | 0.60 | 5.6 |
(*) The results of sample 1 for ABG, ASM, IDUA and GAA were outside the measuring range and therefore not included.
Limit of Blank, Detection and Quantification:
The Limits of Blank, Detection and Quantitation were determined in accordance with CLSI document EP17-A2. The Limit of Blank (LoB) for the NeoLSD MSMS kit is defined as the 95th percentile of a distribution of blank samples determined with 120 determinations for the TQD MSMS instrument. The Limit of Detection (LoD) is based on 150 determinations of low level samples using the Waters TQD MSMS instrument.
| Enzyme | LoB
(µmol/L/h) | LoD
(µmol/L/h) |
|--------|-------------------|-------------------|
| ABG | 0.165 | 0.63 |
| ASM | 0.110 | 0.27 |
| GALC | 0.106 | 0.34 |
| IDUA | 0.059 | 0.24 |
| GLA | 0.476 | 0.97 |
| GAA | 0.073 | 0.39 |
The Limit of Quantitation (LoQ) is defined as the lowest activity fulfilling the total CV% requirement of the assay. For ABG, GLA and IDUA the CV% requirement is 0.18 g/dL) due to glucose infusion.
Hematocrit interference on ABG and GALC: Hematocrit at level of 35% decreases ABG activity levels of 1.10, 2.92 and 8.48 µM/h by 0.49 µM/h, 25% and 18%, respectively. This may cause a false positive screening result for a specimen with measured ABG activity close to the cut-off value. Also, hematocrit at level of 65% increases ABG activity levels of 2.92 and 8.48 µM/h by 29% and 28%, respectively. At hematocrit levels ≥65%, the measured ABG activity levels could be increased as much as by 35%. Therefore, at hematocrit levels of ≥65% the interference could result in misclassification of a patient with an ABG result near the cut-off value as 'normal' when in fact patient should be classified as having presumed ABG deficiency. For samples up to 30% above the ABG cut-off value, with known or suspected high hematocrit levels (≥ 65%); testing by an alternate method that is not subject to hematocrit interference is recommended.
13
Hematocrit at level of 35% increases GALC activity level of 3.38 µM/h by 22%; while at levels of 65% hematocrit decreases GALC activity levels of 0.75 and 1.02 µM/h by 24% and 20%, respectively. The decrease in GALC activity may cause a false positive screening result for a specimen with measured GALC activity close to the cut-off value. The inhibitory effect of high hematocrit values to GALC activity has been reported.
Hemoglobin interference on ABG, ASM and IDUA:
Hemoglobin was found to interfere with the assay by increasing the measured ABG, ASM and IDUA activity.
ABG: Hemoglobin at level of 16.6 g/dL increases ABG activity level of 2.36 µM/h by 27% when compared to a sample having 15.3 g/dL of hemoglobin at level of 18.0 g/dL hemoglobin increases ABG activity level of 0.65 µM/h by 0.34 µM/h when compared to a sample having 15.5 g/dL of hemoglobin at level of 18.9 g/dL increases ABG activity level of 7.68 µM/h by 30% when compared to a sample having 15.1 g/dL of hemoglobin.
ASM: Hemoglobin at level of 17.8 g/dL hemoglobin increases ASM activity level of 1.57 µM/h by 26% when compared to a sample having 15.3 g/dL of hemoglobin at level of 18.0 g/dL increases ASM activity level of 0.41 µM/h by 0.20 µM/h when compared to a sample having 15.5 g/dL of hemoglobin at level of 20.1 g/dL increases ASM activity level of 4.78 µM/h by 16% when compared to a sample having 15.1 g/dL of hemoglobin.
IDUA: Hemoglobin at level of 18.0 g/dL increases IDUA activity level of 1.18 µM/h by 19% when compared to a sample having 15.5 g/dL of hemoglobin.
Although interference by hemoglobin was observed with concentrations within the newborn reference ranges (12.0 – 22.0 g/dL for hemoglobin), it is concluded based on the external study results that the interferences are not pronounced enough to impair the separation of the affected and unaffected cases.
Triglyceride interference on GAA, GLA and IDUA: Intralipid (Triglyceride) was found to interfere with the assay increasing the measured GAA, GLA and IDUA activity.
GAA: Intralipid at level of 0.23 g/dL increases GAA activity of 2.98 and 7.10 µM/h by 27% and 18%, respectively. Intralipid at level of 0.75 g/dL increases GAA activity level of 1.05 µM/h by 0.52 µM/h.
14
GLA: Intralipid at level of 0.30 g/dL increases GLA activity level of 7.21 µM/h by 16%; while at level 0.38 g/dL Intralipid increases GLA activity level of 3.63 µM/h by 25%. Intralipid at level of 0.75 g/dL increases GLA activity level of 1.37 µM/h by 0.65 µM/h.
IDUA: Intralipid at level of 0.38 g/dL increases IDUA activity level of 8.18 µM/h by 19% and Intralipid at level of 0.75 g/dL increases IDUA activity levels of 1.17 and 2.67 µM/h by 20%.
Note! High triglyceride concentrations (hypertriglyceridemia) in newborns due to medication effects or pathological conditions may cause a false negative screening result for a specimen with measured GAA, GLA or IDUA activity close to the cut-off value.
EDTA interference on ABG and ASM: EDTA at level 0.10 g/dL increases ABG activity level of 1.80 µM/h by 0.41 µM/h. EDTA at level of 0.04 g/dL decreases ASM activity level of 3.74 µM/h by 20%; while at level of 0.10 g/dL EDTA decreases ASM activity level of 1.18 µM/h by 16%. EDTA at a level of 0.20 g/dL decreases ASM activity level of 0.44 μM/h by 0.17 μM/h.
The concentration of EDTA found to interfere with AGB and ASM were far beyond the typical therapeutic dosage 3.4 µmol/L (0.10 mg/dL) [12]. Also, the direct application of blood from the heel-puncture to the DBS paper is intended for newborn screening eliminating the risk of contamination.
Reference Range:
A prospective clinical study using four-year-old retrospective routine newborn screening samples was conducted with the NeoLSD MSMS test system to screen for the lysosomal enzymes ABG, ASM, GAA, GALC, GLA, and IDUA.
Over the course of three months 5041 newborn samples were tested at a European (EU) newborn screening laboratory to establish cut-off values. Samples in the reference range study were from newborns ranging 0-30 days of age. The initial cut-off values were based conservatively on 0.1 - 0.3 percentile of enzyme activity distribution and converted to a percentage of population median activity. The retest cut-off values were set 5% lower from the initial cut-off percentage. The cut-off percentages were applied to daily medians established based on the initial routine sample results for the day.
The cut-off percentages were not adjusted during the study. The initial and retest cut-off values used in the clinical study are shown below.
15
| Enzyme | n | Range* | Enzyme activity (µmol/L/h) | Lower percentiles | Initial cutoff | Retest cutoff | |||
|---|---|---|---|---|---|---|---|---|---|
| Mean | Median | 0.1% | 0.2% | 0.3% | |||||
| ABG | 5041 | 2.07 - 66.3 | 10.35 | 9.60 | 2.85 | 3.16 | 3.33 | 35% | 30% |
| ASM | 5041 | 1.64 - 35.2 | 6.38 | 5.86 | 2.12 | 2.21 | 2.37 | 40% | 35% |
| GALC | 5041 | 0.22 - 54.9 | 3.63 | 3.10 | 0.43 | 0.52 | 0.56 | 20% | 15% |
| IDUA | 5041 | 0.45 - 35.4 | 7.55 | 7.25 | 2.06 | 2.55 | 2.62 | 30% | 25% |
| GLA | 5041 | 1.48 - 94.8 | 11.06 | 9.72 | 3.04 | 3.37 | 3.62 | 40% | 35% |
| GAA | 5041 | 1.55 - 34.8 | 9.28 | 8.74 | 2.33 | 2.69 | 2.92 | 30% | 25% |
Descriptive statistics for the EU site
- Some results were outside the measuring range and cannot be considered accurate
Additional newborn population distributions were determined at two different newborn screening laboratories in the United States. Site A analyzed 5251 and Site B analyzed 5053 newborn dried blood spot specimens submitted for routine testing. Descriptive statistics for the samples are shown below. Samples tested in Site A and Site B were collected from newborns of ≤ 4 days and ≤ 7 days, respectively.
Descriptive statistics for the US Site A
| Enzyme activity (µmol/L/h) | |||||||
|---|---|---|---|---|---|---|---|
| Enzyme | n | Range* | Mean | Median | Lower percentiles | ||
| 0.1% | 0.2% | 0.3% | |||||
| ABG | 5251 | 1.39 – 77.9 | 10.52 | 9.67 | 2.00 | 2.16 | 2.36 |
| ASM | 5251 | 1.10 – 24.3 | 4.70 | 4.44 | 1.49 | 1.60 | 1.64 |
| GALC | 5251 | 0.52 – 85.0 | 5.49 | 4.63 | 0.86 | 0.96 | 1.02 |
| IDUA | 5251 | 0.09 – 18.7 | 6.31 | 6.08 | 0.85 | 1.40 | 1.56 |
| GLA | 5251 | 1.69 – 121 | 16.91 | 14.39 | 3.44 | 4.42 | 4.89 |
| GAA | 5251 | 0.75 - 43.0 | 10.21 | 9.62 | 2.16 | 2.38 | 2.56 |
- Some results were outside the measuring range and cannot be considered accurate
| Enzyme | n | Range* | Mean | Median | Lower percentiles | ||
|---|---|---|---|---|---|---|---|
| 0.1% | 0.2% | 0.3% | |||||
| ABG | 5053 | 1.68 – 76.5 | 11.4 | 10.4 | 2.94 | 3.23 | 3.33 |
| ASM | 5053 | 1.02 – 28.4 | 5.44 | 5.13 | 1.71 | 1.79 | 1.98 |
| GALC | 5053 | 0.53 – 33.0 | 4.67 | 4.20 | 0.68 | 0.82 | 0.86 |
| IDUA | 5053 | 0.9 – 21.8 | 6.65 | 6.39 | 1.59 | 1.98 | 2.09 |
| GLA | 5053 | 0.96 – 59.5 | 13.3 | 12.0 | 3.99 | 4.33 | 4.43 |
| GAA | 5053 | 1.11 – 36.4 | 11.0 | 10.4 | 2.53 | 2.88 | 3.37 |
Descriptive statistics for the US Site B
16
- Some results were outside the measuring range and cannot be considered accurate
Screening Performance:
The screening performance of the NeoLSD MSMS kit was determined in a prospective clinical study of four-year-old retrospective routine samples at an EU newborn screening laboratory. Over the twomonth study duration 4041 newborn samples were tested were from newborns ≤ 29 days old. All 4011 routine samples were 4 years old enabling the follow-up of clinical status at the age 4 years. Due to the low incidence of LSDs, the study population was enriched confirmed LSD positive newborn DBS specimens obtained from the site's biobank, these samples ranged from 5.8 to 17.6 years of age. Sample inclusion criteria were the same as in the reference range study.
The 4011 routine samples were tested according to the defined algorithm to determine the enzyme activities for sample interpretation. Site specific cut-off percentages determined prior to the clinical study were applied as percent of daily median activity. The routine specimens having enzyme activities below the cut-off values in the initial testing were re-tested in duplicate. The final results (normal, presumptive positive, invalid result) were classified after the initial testing for samples above the cut-off values and after the final testing for samples below the initial cut-off level.
Confirmed LSD positive newborn specimens were tested once, and assessed using the retest cut-off values for the enzymes. Given the known status of the confirmed LSD positive newborn specimens, processing the samples through the full testing algorithm provided no additional information nor changed the sample interpretation.
Clinical outcome was used as a comparator for all samples, including the 4011 routine screening samples, as derived from the civil registry status and national hospital registry. Subject´s survival at 4 years of age without LSD diagnosis or clinical signs suggestive of an LSD was used as clinical confirmation of an unaffected newborn. Subjects who did not have follow-up information available up to 4 years were classified as lost to follow-up.
Results
A total of 4041 specimens (4011 routine newborn specimens from confirmed positive LSD cases) were tested in the clinical study. The following is a summary of the samples tested.
| Routine | ABG | ASM | GALC | IDUA | GLA | GAA | Total |
|---|---|---|---|---|---|---|---|
| Screened samples | 4011 | 4011 | 4011 | 4011 | 4011 | 4011 | 4011 |
| Below initial cutoff | 15 | 11 | 12 | 33 | 12 | 10 | 88 |
| Screen positive | 3 | 0 | 10 | 5 | 5 | 4 | 26 |
| Unaffected | 3 | 0 | 10 | 5 | 5 | 3 | 25 |
Summary of the samples tested in the pivotal study.
17
Wallac Oy, PerkinElmer Inc Confidential
| Lost to follow up | 0 | 0 | 0 | 0 | 0 | 1 | 1 |
|---|---|---|---|---|---|---|---|
| Retest rate | 0.40% | 0.27% | 0.30% | 0.17% | 0.85% | 0.30% | 2.2% |
| False positive rate | 0.07% | 0.00% | 0.10% | 0.07% | 0.27% | 0.10% | 0.6% |
| Confirmed Positive | ABG | ASM | GALC | IDUA | GLA | GAA | Total |
| Screened samples | 5 | 1 | 10 | 5 | 5 | 4 | 30 |
| Below initial cutoff | 5 | 1 | 10 | 5 | 3 | 4 | 28 |
| Screen positive | 3 | 1 | 10 | 5 | 3 | 4 | 26 |
| Invalid | 2 | 0 | 0 | 0 | 0 | 0 | 2 |
| Screen negative | 0 | 0 | 0 | 0 | 2 | 0 | 2 |
Two confirmed positive Gaucher specimens were categorized as "invalid result". In routine newborn screening, if a specimen result is categorized as invalid result, a new dried blood spot specimen should be obtained and retesting performed using age-specific cut-off values.
In total 86 subjects had emigrated before the age of 4 years and therefore their clinical status could not be confirmed. None of the subjects in the lost to follow-up cohort had LSD diagnosis or showed signs and symptoms of any of the screened LSDs at the time of being lost to follow up.
The screening performance results for the 4041 specimens, including the 30 confirmed LSD positive specimens, tested with the NeoLSD MSMS kit are shown below.
| Outcome, all 6 enzymes | |||||
|---|---|---|---|---|---|
| Screening Results | Affected | Unaffected | Lost-to- | ||
| follow-up | Total | ||||
| Positive | 26 | 25 | 1 | 52 | |
| NeoLSD | |||||
| MSMS kit | Negative | 2* | 3900 | 85 | 3987 |
| Invalid | 2 | 0 | 0 | 2 | |
| Total | 30 | 3925 | 86 | 4041 | |
| Performance estimates (invalid and lost-to-follow-up excluded) | |||||
| Sensitivity | False-negative rate | Specificity | False-positive rate | ||
| 92.9 % | 7.1%* | 99.4 % | 0.6 % | ||
| (76.5%-99.1%) | (0.9% - 23.5%) | (99.1% - 99.6%) | (0.4% - 0.9%) |
Screening performance of NeoLSD MSMS kit
- includes 2 Fabry females
| Outcome, Gaucher (ABG) | |
|---|---|
| -- | ------------------------ |
18
| Screening Results | | Affected | Unaffected | Lost-to-
follow-up | Total |
|--------------------|----------|----------|------------|-----------------------|-------|
| NeoLSD
MSMS kit | Positive | 3 | 3 | 0 | 6 |
| | Negative | 0 | 3947 | 86 | 4033 |
| | Invalid | 2 | 0 | 0 | 2 |
| Total | | 5 | 3950 | 86 | 4041 |
| Outcome, Niemann-Pick A/B (ASM) | |||||
|---|---|---|---|---|---|
| Screening Results | Affected | Unaffected | Lost-to-follow-up | Total | |
| NeoLSD | |||||
| MSMS kit | Positive | 1 | 0 | 0 | 1 |
| Negative | 0 | 3952 | 86 | 4038 | |
| Invalid | 0 | 2 | 0 | 2 | |
| Total | 1 | 3954 | 86 | 4041 |
| Outcome, Krabbe (GALC) | |||||
|---|---|---|---|---|---|
| Screening Results | Affected | Unaffected | Lost-to- | ||
| follow-up | Total | ||||
| NeoLSD | |||||
| MSMS kit | Positive | 10 | 4 | 0 | 14 |
| Negative | 0 | 3939 | 86 | 4025 | |
| Invalid | 0 | 2 | 0 | 2 | |
| Total | 10 | 3945 | 86 | 4041 |
| Outcome, MPS I (IDUA) | |||||
|---|---|---|---|---|---|
| Screening Results | Affected | Unaffected | Lost-to- | ||
| follow-up | Total | ||||
| NeoLSD | |||||
| MSMS kit | Positive | 5 | 3 | 0 | 8 |
| Negative | 0 | 3945 | 86 | 4031 | |
| Invalid | 0 | 2 | 0 | 2 | |
| Total | 5 | 3950 | 86 | 4041 |
| Outcome, Fabry (GLA) | |||||
|---|---|---|---|---|---|
| Screening Results | Affected | Unaffected | Lost-to- | ||
| follow-up | Total | ||||
| NeoLSD | |||||
| MSMS kit | Positive | 3 | 11 | 0 | 14 |
| Negative | 2 | 3937 | 86 | 4025 | |
| Invalid | 0 | 2 | 0 | 2 | |
| Total | 5 | 3950 | 86 | 4041 |
19
| Outcome, Pompe (GAA) | |||||
|---|---|---|---|---|---|
| Screening Results | Affected | Unaffected | Lost-to- | ||
| follow-up | Total | ||||
| NeoLSD | |||||
| MSMS kit | Positive | 4 | 4 | 1 | 9 |
| Negative | 0 | 3945 | 85 | 4030 | |
| Invalid | 0 | 2 | 0 | 2 | |
| Total | 4 | 3951 | 86 | 4041 |
With the female Fabry subjects excluded the NeoLSD MSMS test system has no false negative results for any of the enzymes; and a false positive range of ABG (0.07%), ASM (0%), GALC (0.10%), IDUA (0.07 %), GLA (0.27 %), GAA ( 0.10%). . The retest rate was 6.6%, of which only 2.2% was due to test failure (e.g. quality control exceeded limits). Remaining retests (4.4%) were due to the screening algorithm, which requires results below initial cut-off value for any or the six enzymes to be repeated in duplicate.