Total PSA
Immunoassay Analyzer
REF C86000 2×50 Tests 0123
INTENDED USE
The iFlash-Total PSA assay is a paramagnetic particle
chemiluminescent immunoassay (CLIA) for the
quantitative determination of total prostate specific antigen
(TPSA) in human serum and plasma using the iFlash
Immunoassay Analyzer.
SUMMARY AND EXPLANATION
Prostate specific antigen (PSA) is a glycoprotein acting as
a serine protease. The proteolytic activity of PSA in blood
is inhibited by the irreversible formation of complexes with
proteinase inhibitors such as alpha-1-antichymotrypsin,
alpha-2-macroglobulin and other acute phase proteins. In
addition to being present in these complexes, PSA is also
present in blood in the free form, but is proteolytically
inactive. Elevated concentrations of PSA in serum are
generally indicative of a pathologic condition of the
prostate (prostatitis, benign hyperplasia or carcinoma).
PSA testing has significant value in detecting metastatic or
persistent prostate diseases in patients following surgical
or medical treatment of prostate cancer. Persistent
elevation of PSA following treatment, or an increase in a
post-treatment PSA level is indicative of recurrent or
residual disease. PSA testing is widely accepted as an
adjunctive test in the management of prostate cancer
patients.
PSA tests lack sufficient sensitivity and specificity to be
considered ideal or absolutely diagnostic for screening or
early detection because PSA is not specific for prostate
cancer. A number of studies have found that the % Free
PSA (FPSA) was significantly lower in patients having
prostate cancer than those with benign disease or normal
controls. The ratio FPSA/TPSA has been demonstrated to
improve the sensitivity and specificity in patients with
TPSA values in the “gray zone” of 4-10 ng/mL.
ASSAY PRINCIPLE
The iFlash-Total PSA assay is a sandwich immunoassay.
 Incubation: Total PSA in the sample, anti-PSA coated
paramagnetic microparticles and anti-PSA
acridinium-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 total
PSA in the sample and the RLUs detected by the
iFlash optical system.
 Results are determined via a calibration curve, which
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iFlash
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-PSA coated microparticles, 3.5
R1
mL/pack, 0.05% ProClin 300.
Anti-PSA acridinium-labeled conjugate, 4.0
R2
mL/pack, 0.05% ProClin 300.
Calibrator 1, 1 bottle, 1.0 mL, Tris buffer
CAL1
with protein stabilizers, 0.05% ProClin 300.
Calibrator 2, 1 bottle, 1.0 mL, PSA in Tris
CAL2 buffer with protein stabilizers, 0.05%
ProClin 300.
Calibrator 3, 1 bottle, 1.0 mL, PSA in Tris
CAL3 buffer with protein stabilizers, 0.05%
ProClin 300.
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 human
sourced 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 and calibrators are ready for
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Total PSA
use .Close the bottles of calibrator right after
calibration and store at 2–8°C.
 Do not pool reagents within a reagent kit or between
reagent kits.
 Prior to loading the iFlash-Total PSA reagent pack on the
system for the first time, resuspend the microparticles by
inverting the reagent pack 30 times 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-Total PSA 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 maximum 1 time.
 Centrifuged 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 caution in handling patient specimens to prevent
cross-contamination.
 Do not use heat-inactivated samples.
 Ensure 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.
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iFlash
Immunoassay Analyzer
ASSAY PROCEDURE
 Refer to the system operating instruction or the online
help system for detailed information on preparing the
system.
 The test-specific parameters stored in barcode on the
reagent pack are read in. In cases 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.
 Press RUN, the iFlash System performs all the
functions automatically and calculates the results.
 PSA in the sample concentration is lower than 17000
ng/mL, there will be no HOOK effect.
CALIBRATION
 Traceability: This assay has been standardized
against the Stanford Reference Standard/WHO 96/670
(90 % PSA-ACT + 10 % Free PSA).
 Every iFlash-Total PSA reagent kit has a QR code
label containing the specific information for calibration
of the particular reagent lot.
 To perform an iFlash-Total PSA calibration, test CAL1.
CAL2, and CAL3 in duplicate, and the predefined
master curve is adapted to the analyzer.
 Once an iFlash-Total PSA 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
 0.01 – 100 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
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Total PSA iFlash
Immunoassay Analyzer

concentration of each sample. The results are given in total PSA were assayed.
ng/mL. The within run precision was determined by testing each
sample in replicates of 10 (n = 10), and calculating percent
Expected Values:
coefficient of variation (%CV). The results of the study are
A study of iFlash Total PSA assay on samples from 281 shown below:
apparently healthy males of various age groups yielded the Sample Mean (ng/mL) SD %CV
following result:
1 4.36 0.17 3.90
th
< 4 ng/mL (95 percentile) 2 30.26 1.68 5.55
It is recommended that each laboratory establish its own
The between run precision was determined by testing
expected reference range for the population of interest.
each sample in duplicate, two separate runs daily for 20
LIMITATIONS days (n = 80), and calculating percent coefficient of
 The iFlash-Total PSA assay is limited to the variation (%CV). The results of the study are shown
determination of total PSA in human serum or plasma below:
Sample Mean (ng/mL) SD %CV
(lithium heparin, sodium heparin, potassium EDTA, and
sodium citrate). It has not been validated for use with 1 4.41 0.18 4.08
other types of plasma. 2 30.05 1.43 4.76
 The use of serum separator (gel) blood collection tubes
Analytical Sensitivity
has been validated for use with this assay; however it is
not possible to survey all manufacturers or tube types. The detection limit representing the lowest measurable
analyte level that can be distinguished from zero is 0.01
 The upper limit of the measuring range of this assay is
ng/mL. It is calculated as the value lying two standard
100 ng/mL. Over-range samples may be diluted with
deviations above that of the lowest standard of the master
negative human serum and re-tested to obtain an
curve (standard 1 + 2 SD, n = 20).
estimate of the actual concentration.
 If the results are inconsistent with clinical evidence, Method comparison
additional testing is suggested to confirm the result. A comparison of the iFlash-Total PSA assay (y) with a
 For diagnostic purposes, the results should be commercially available total PSA assay (x) using clinical
interpreted in light of the total clinical presentation of the samples was performed, and the curve is fitted with Linear
patient, including symptoms, clinical history results. regression)
 Specimens from heparinized patients may be partially y = 0.9938x -0.1092
coagulated and erroneous results could occur due to r = 0.998
the presence of fibrin.
Sample concentration: 0.23 – 99 ng/mL
 The results from an alternative assays (i.e. EIA or RIA)
Number of samples measured: 90
may not be equivalent and cannot be used
interchangeably. Equimolar reactivity
 Samples containing an apparent total PSA level as high On different proportions of PSA-ACT and free PSA, the
as 17000 ng/mL did not exhibit a hook effect in the deviations of total PSA should not exceed±15%.
iFlash Total PSA assay.
Accuracy
 The assay is unaffected by icterus (bilirubin < 30
mg/dL), hemolysis (Hb < 1500 mg/dL), lipemia The relative deviations.≤10%.
(Intralipid < 1500 mg/dL) and total serum protein (< 10 Range of linear
g/dL).
In the range of 0.01-80 ng/mL, the correlation coefficient
 No interference was observed from rheumatoid factors ＞0.99.
up to a concentration of 2000 IU/mL.
 No interference was observed from anti-nuclear REFERENCES
antibodies up to a concentration of 500 U/mL. 1. Tewari PC, Bluestein BI. Multiple forms of prostate
 No interference was observed from HAMA up to a specific antigen and the influences of immunoassay
concentration of 600 ng/mL. design on their measurement in patient serum. J Clin
Ligand Assay, 18 1995;3:186-196.
PERFORMANCE CHARACTERISTICS 2. Prestigiacomo AF, Stamey TA. Clinical usefulness of
Below are the representative performance data, and the free and complexed PSA. Clin Lab Invest Suppl
results obtained in individual laboratories may differ. 1995;221:32-34.

Precision 3. Semjonow A, De Angelis G, Oberpenning F et al. The

clinical impact of different assays for prostate specific
Two controls, consisting low, and median concentration of
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Total PSA iFlash
Immunoassay Analyzer

antigen. BJU Int. 2000; 86(5):590-7 ANNEX A:

4. Kuriyama M, Wang MC, Papsidero LD, et al. Explanation of abbreviation
Quantitation of Prostate-Specific Antigen in Serum by
Abbreviation Explanation
a Sensitive Enzyme Immunoassay. Cancer Res
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Assessment of the Most Useful Tumor Marker for
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1991;145:907-23.
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Its Use as a Tumor Marker for Prostate Cancer
Hematol Oncol Clin N Amer 1994;8:555-72.
Number of tests
7. Partin AW, Oesterling JE. The Clinical Usefulness of
Prostate Specific Antigen: Update 1994. J Urol
1994;152:1358-68. Manufactured by
8. Bunting S. A Guide to the Interpretation of Serum
Prostate Specific Antigen Levels. Clin Biochem EU Representative
1995;28:221-41.
9. Catalona WJ, Richie JP, Ahmann FR, Hudson MA, EC Declaration of Conformity
Scardino PT, Flanigan RC, et al. Comparison of digital
rectal examination and serum prostate-specific antigen
in the early detection of prostate cancer: results of Caution
a multicenter clinical trial of 6,630 men. J Urology
1994;151(5):1283-1290. Instructions for use
10. Chen YT, Luderer AA, Thiel RP, et al. Using
proportions of free to total prostate-specific antigen, In vitro diagnostic medical
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probability of prostate cancer. Urology
Lot No.
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11. Thiel RP, Oesterling JE, Wojno KJ, et al. A multicenter
comparison of the diagnostic performance of free Date of manufacture
prostate-specific antigen. Urology 1996;48(6A):45-50.
Expiry date

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