NSE
Immunoassay Analyzer
REF C86029 2×50 Tests
INTENDED USE
The iFlash-NSE assay is a paramagnetic particle
chemiluminescent immunoassay (CLIA) for the
quantitative determination of Neuron-specific enolase
(NSE) in human serum and plasma using the iFlash
Immunoassay Analyzer.
SUMMARY AND EXPLANATION
Neuron-specific enolase (NSE) is informed from the
enolase isoforms αγ and γγ. NSE is described as the
marker of first choice in the monitoring of small cell,
bronchial carcinoma, whereas CYFRA 21-1 is superior to
NSE for non-small cell bronchial carcinoma. NSE is also
used to monitor the neuroblastoma and apudoma. Brain
tumors such as glioma, meningioma, neurofibroma and
neurinoma are only occasionally accompanied by
elevated serum NSE values.
For NSE there is good correlation to the clinical stage, i.e.
the extent of the disease. In response to chemotherapy
there is a temporary rise in the NSE level 24–72 hours
after the first therapy cycle as a result of cytolysis of the
tumor cells. During remission, 80–96% of the patients
have normal values. Rising NSE values are found in
cases of relapse.
ASSAY PRINCIPLE
The iFlash-NSE assay is a sandwich immunoassay.
 Incubation: NSE in the sample, anti-NSE coated
paramagnetic microparticles and anti-NSE
acridinium-ester-labeled conjugate react to form a
sandwich complex.
 Wash: The unbound materials are washed away from
the solid phase in a magnetic field.
 Trigger of signal: The Pre-Trigger and Trigger
Solutions are added to the reaction mixture. The
resulting chemiluminescent reaction is measured as
relative light units (RLUs).
 A direct relationship exists between the amount of NSE
in the sample and the RLUs detected by the iFlash
optical system.
 Results are determined via a calibration curve, which
is instrument-specifically generated by 3-point
calibration and a master curve provided via the
reagent QR code.
REAGENTS
Reagent kit, 100 tests, 2 packs, 50 tests/pack
Anti-NSE coated microparticles, 3.5
R1
mL/pack, 0.05% ProClin 300.
Anti-NSE acridinium-ester-labeled
R2
conjugate; 4.0 mL/pack; 0.05% ProClin 300.
1/4
iFlash
Calibrator 1, 1 bottle, 1.0 mL, Tris buffer
CAL1 with protein stabilizers, 0.05% ProClin 300,
lyophilized product.
Calibrator 2, 1 bottle, 1.0 mL, NSE in Tris
CAL2 buffer with protein stabilizers, 0.05%
ProClin 300, lyophilized product.
Calibrator 3, 1 bottle, 1.0 mL, NSE in Tris
CAL3 buffer with protein stabilizers, 0.05%
ProClin 300, lyophilized product.
MATERIALS REQUIRED (BUT NOT PROVIDED)
REF C89999/C89959/C89949, iFlash Pre-Trigger
Solution: hydrogen peroxide solution.
REF C89998/ C89958/ C89948, iFlash Trigger Solution:
sodium hydroxide solution.
REF C89997, iFlash Wash Buffer: phosphate buffered
saline solution with 0.05% ProClin 300.
REF C80001, iFlash Wash Buffer (10×): phosphate
buffered saline solution with 0.05% ProClin 300.
REF C89996, reaction vessels.
Controls: Commercial controls could be used.
WARNINGS AND PRECAUTIONS
IVD For in vitro diagnostic use
 No known test method can offer the complete
assurance that products derived from human sources
will not transmit infection. Therefore, all humanized
materials should be considered potentially infectious.
 Exercise the normal precautions required for handling
all laboratory reagents.
 Disposal of all waste material should be in accordance
with local guidelines.
 Wear gloves when handling specimens or reagents.
 Clean and disinfect all spills of specimens or reagents
using a suitable disinfectant.
 iFlash Trigger solution contains sodium hydroxide
(NaOH) and should be avoided contact with eyes.
REAGENT HANDLING
 The reagents may not be used after the stated
expiration date.
 Avoid the formation of foam with all reagents.
 The reagents in the pack are ready for use.
 Each tube of lyophilized powder, as a calibrator, needs
adding 1.0 mL deionized water, dissolving within 15
minutes and gently blending for the first time.
 After dissolving calibrators, they should be measured
within 4 hours. Then, they should be frozen to
preserve.
 Close the bottles of calibrator right after calibration and
store at 2–8°C.
 Do not pool reagents within a reagent kit or between
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NSE
reagent kits.
 Prior to loading the iFlash-NSE reagent pack on the
system for the first time, resuspend the microparticles
by inverting the reagent pack slightly.
 For further information on reagent handling
precautions during system operation, refer to the
iFlash system operating instruction.
STORAGE AND STABILITY
Storage:
 Store at 2–8°C in an upright position.
 The kit may be used immediately after removal from
2-8°C storage.
Stability:
 Unopened at 2–8°C: up to the stated expiration date.
 Opened at 2–8°C: 28 days.
 Store on-board: 28 days.
SPECIMEN COLLECTION AND PREPARATION
 Serum or plasma (lithium heparin, sodium heparin
potassium EDTA, and sodium citrate) are the
recommended samples. Other anticoagulants have
not been validated for use with the iFlash-NSE assay.
 Ensure that serum specimens to form complete clot
prior to centrifugation.
 Centrifuge the specimens.
 Store specimens at room temperature (20 to 25°C) for
no longer than 8 hours.
 If the testing will not be completed within 8 hours,
refrigerate the samples at 2 to 8°C.
 If the testing will not be completed within 3 days, or for
shipment of samples, freeze at -20°C or colder.
 Frozen specimens must be mixed thoroughly after
thawing.
 The samples may be frozen for maximum1 time.
 Centrifuge specimens with a lipid layer on the top, and
transfer only the clarified specimen without the lipemic
material.
 Ensure that residual fibrin and cellular matter have
been removed prior to analysis.
 Use with caution in handling patient specimens to
prevent cross-contamination.
 Do not use heat-inactivated samples.
 Ensure that the patient samples, calibrators and
controls are at ambient temperature (20–25°C) before
measurement.
 Due to the possible evaporation, specimens and
calibrators on the analyzers should be measured
within 2 hours.
ASSAY PROCEDURE
 Refer to the system operating instruction or the online
2/4
iFlash
Immunoassay Analyzer
help system for detailed information on preparing the
system.
 The test-specific parameters stored in barcode on the
reagent pack are read in. In case the barcode cannot
be read, enter the sequence numbers.
 Carry out calibration, if necessary.
 Place the calibrators CAL1, CAL2, and CAL3 in the
calibrator rack in the sample zone. Only keep
calibrators open during calibration.
 Test application.
 Load samples (Use 20 μL of sample for each
determination in addition to the sample container and
system dead volumes).
 Click RUN, the iFlash System performs all the
functions automatically and calculates the results.
CALIBRATION
 Traceability: This assay is traceable to a commercial
available kit.
 Every iFlash-NSE reagent kit has a QR code label
containing the specific information for calibration of the
particular reagent lot.
 To perform an iFlash-NSE calibration, test CAL1,
CAL2 and CAL3 in duplicate, and the predefined
master curve is adapted to the analyzer.
 Once an iFlash-NSE calibration is accepted and stored,
all subsequent samples may be tested without further
calibration unless:
 After 28 days when using the same reagent lot.
 A reagent kit with a new lot number is used.
 Controls are out of range.
 Required by pertinent regulations.
MEASURING RANGE
 0.05 – 370 ng/mL
QUALITY CONTROL
Quality control materials should be run as single
determinations at least once every 24 hours when the test
is in use, once per reagent kit and after every calibration.
Include commercially available quality control materials
that cover at least two levels of analyte. Follow
manufacturer’s instructions for reconstitution and storage.
Each laboratory should establish mean values and
acceptable ranges to assure proper performance. Quality
control results that do not fall within acceptable ranges
may indicate invalid test results.
RESULT
Calculation:
The iFlash system automatically calculates the analyte
concentration of each sample. The results are given in
ng/mL.
V2.0 English Ed.2018-02-01
NSE iFlash
Immunoassay Analyzer

Expected Values: median concentration of NSE were assayed.

A study of iFlash-NSE assay on samples from 368 The within run precision was determined by testing each
apparently healthy patients of various age groups yielded sample in replicates of 10 (n = 10), and calculating percent
the following result: coefficient of variation (%CV). The results of the study are
th shown below:
< 15.3 ng/mL (95 percentile)
It is recommended that each laboratory establish its own Sample Mean (ng/mL) SD %CV
expected reference range for the specific population.
1 2.02 0.11 5.45
LIMITATIONS
2 53.12 1.88 3.54
 The iFlash-NSE assay is limited to the determination
of NSE in human serum or plasma (lithium heparin, The between run precision was determined by testing
sodium heparin, potassium EDTA, and sodium citrate). each sample in duplicate, two separate runs daily for 20
It has not been validated for use with other types of days (n = 80), and calculating percent coefficient of
plasma. variation (%CV). The results of the study are shown
 The use of serum separator (gel) blood collection below:
tubes has been validated for use with this assay. Sample Mean (ng/mL) SD %CV
However, it is not possible to survey all manufacturers
or tube types. 1 2.03 0.09 4.43
 The upper limit of the measuring range of this assay is
2 51.77 2.18 4.21
370 ng/mL. Over-range samples may be diluted with
negative human serum and re-tested to obtain an
estimate of the actual concentration. Analytical Sensitivity
 If the results are inconsistent with clinical evidence, The detection limit representing the lowest measurable
additional testing is suggested to confirm the result. analyte level is 0.05 ng/mL, which can be distinguished
from zero. It is calculated as the value lying two standard
 For diagnostic purposes, the results should be
deviations above that of the lowest standard of the master
interpreted in light of the total clinical presentation of
curve (standard 1 + 2 SD, n = 20).
the patient, including symptoms, clinical history results.
 Specimens from heparinized patients may be partially Analytical Specificity
coagulated and erroneous results could occur due to The analytical specificity of iFlash-NSE assay was
the presence of fibrin. evaluated with CYFRA21-1 and neuron-non-specific
 The results from an alternative assays (i.e. EIA or RIA) enolase. The nonreactive NSE status of each specimen
may not be equivalent and cannot be used was verified using a commercially available NSE assay.
interchangeably. iFlash-NSE
Concentration
 Samples containing an apparent NSE level as high as Clinical Category
(ng/mL)
Nonreactive
100,000 ng/mL did not exhibit a hook effect in the (ng/mL)
iFlash-NSE assay.
Neuron-non-specific
 The assay is unaffected by icterus (bilirubin < 30 500 0.19
enolase
mg/dL), hemolysis (Hb < 1,500 mg/dL), lipemia
(Intralipid < 1,500 mg/dL) and total serum protein (< 10 CYFRA21-1 100 0.04
g/dL).
 No interference was observed from rheumatoid factors Method Comparison
up to a concentration of 2,000 IU/mL. A comparison of the iFlash-NSE assay (y) with a
 No interference was observed from anti-nuclear commercially available NSE assay (x) using clinical
antibodies up to a concentration of 500 U/mL. samples was performed, and the curve is fitted with Linear
 No interference was observed from HAMA up to a regression)
concentration of 600 ng/mL. y = 1.0243x -0.4693
r = 0.9975
PERFORMANCE CHARACTERISTICS
Sample concentration: 2.85 – 368 ng/mL
Below are the representative performance data, and the
results obtained in individual laboratories may differ. Number of samples measured: 90

Precision REFERENCES

The precision of iFlash-NSE was determined using NSE 1. Kintzel K, Sonntag J, Strauß E, Obladen M.
reagents and controls. Two controls, consisting low, and Neuron-Specific Enolase: Reference Values in Cord
Blood.Clin Chem Lab Med 1998; 36 (4): 245–247.
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NSE iFlash
Immunoassay Analyzer

2. Ebert W, Hoppe M, Muley TH, Drings P. Monitoring of ANNEX A:

Therapy in Inoperable Lung Cancer Patients by Explanation of abbreviation
Measurement of CYFRA 21-1, TPA-TP, CEA and NSE.
Anticancer Res 1997 (4B); 17: 2875–2878. Abbreviation Explanation
3. Fizazi K, Cojean I, Pignon JP, Rixe O, Gatineau M,
Hadef S, et al. Normal Serum Neuron Specific Enolase Product No.
(NSE) Value after the First Cycle of Chemotherapy.
Cancer 1998; 82 (6): 1049–1055. Calibrator
4. Jacobi, C Reiber H,. Clinical relevance of increased
neuron-specific enolase in cerebrospinal fluid. Clin
Reagent
Chim Acta 1988; 177 (1): 49–54.
5. Martens P. Serum neuron-specific Enolase as a
Prognostic Marker for Irreversible Brain Damage in Number of tests
Comatose Cardiac Arrest Survivors. Acad Emerg Med
1996; 3 (2): 126–131. Manufactured by
6. Rasmussen T, Grankvist K, Ljungberg B. Serum
gamma-enolase and prognosis of patients with renal EU Representative
cell carcinoma. Cancer 1993; 72:1324–1328.

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1st-4th Floor, No.5 Building, Lishan Industrial Caution

Area, Xinghai Road, Nanshan District,
Shenzhen 518054, P.R. China
Instructions for use

In vitro diagnostic medical

Wellkang Ltd (www.CE-marking.eu)
device
Suite B, 29 Harley St., London W1G 9QR, UK
Lot No.

Date of manufacture

Expiry date

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