03045838500V22.

Elecsys CA 15-3 II
cobas e 411
03045838122 03045838500 100 cobas e 601
cobas e 602

English ▪ The reaction mixture is aspirated into the measuring cell where the
microparticles are magnetically captured onto the surface of the
System information electrode. Unbound substances are then removed with
For cobas e 411 analyzer: test number 332 ProCell/ProCell M. Application of a voltage to the electrode then induces
For cobas e 601 and cobas e 602 analyzers: Application Code chemiluminescent emission which is measured by a photomultiplier.
Number 052
▪ Results are determined via a calibration curve which is instrument-
Please note specifically generated by 2‑point calibration and a master curve provided
via the reagent barcode or e‑barcode.
The measured CA 15‑3 value of a patient’s sample can vary depending a) Tris(2,2'-bipyridyl)ruthenium(II)-complex (Ru(bpy) )
on the testing procedure used. The laboratory finding must therefore
Reagents - working solutions
always contain a statement on the CA 15‑3 assay method used. CA 15‑3
The reagent rackpack is labeled as CA 15‑3 II.
values determined on patient samples by different testing procedures
cannot be directly compared with one another and could be the cause of M Streptavidin-coated microparticles (transparent cap), 1 bottle, 6.5 mL:
erroneous medical interpretations. If there is a change in the CA 15‑3
Streptavidin-coated microparticles 0.72 mg/mL; preservative.
assay procedure used while monitoring therapy, then the CA 15‑3 values
obtained upon changing over to the new procedure must be confirmed by R1 Anti-CA 15-3-Ab~biotin (gray cap), 1 bottle, 10 mL:
parallel measurements with both methods. Biotinylated monoclonal antibody (115D8; mouse) 1.75 mg/L;
phosphate buffer 20 mmol/L, pH 6.0; preservative.
Intended use
Immunological in vitro assay for quantitative determination of CA 15‑3 in R2 Anti-CA 15-3-Ab~Ru(bpy) (black cap), 1 bottle, 10 mL:
human serum and plasma to aid in the management of breast cancer
patients. In conjunction with other clinical and diagnostic procedures, serial Monoclonal anti-CA 15-3 antibody (DF3; mouse) labeled with
testing with this assay is an aid ruthenium complex 10 mg/L; phosphate buffer 100 mmol/L, pH 7.0;
preservative.
▪ in the early detection of recurrence in previously treated stage II and III
breast cancer patients Precautions and warnings
▪ for monitoring response to therapy in metastatic breast cancer patients For in vitro diagnostic use.
The electrochemiluminescence immunoassay “ECLIA” is intended for use Exercise the normal precautions required for handling all laboratory
on Elecsys and cobas e immunoassay analyzers. reagents.
Disposal of all waste material should be in accordance with local guidelines.
Summary Safety data sheet available for professional user on request.
The CA 15‑3 (Cancer Antigen 15‑3) is derived from glycoprotein Mucin‑1 This kit contains components classified as follows in accordance with the
(MUC‑1).1 The CA 15‑3 assay uses two monoclonal antibodies (MAb), Regulation (EC) No. 1272/2008:
115D8 and DF3, in a sandwich assay to detect two antigenic sites
associated with breast carcinoma cells. MAb 115D8 is directed against 2-methyl-2H-isothiazol-3-one hydrochloride
human milk fat globule membranes,1,2,3 whereas MAb DF3 is directed
against the membrane fraction from human breast cancer.4 EUH 208 May produce an allergic reaction.
The antigen is normally found in the luminal secretion of glandular cells and
does not circulate in the blood. When cells become malignant and their
basal membranes permeable, the antigen is detectable in serum. 5
Overexpression of MUC1 plays an important role in epithelial to
mesenchymal transition; an important and complex phenomenon that
determines cancer progression.6 The guideline landscape for advanced Warning
disease monitoring was mapped in a review by Duffy et al.7
The low cost and minimally invasive CA 15‑3 monitoring approach is H317 May cause an allergic skin reaction.
mentioned in ASCO and the European Group on Tumor Markers (EGTM)
guidelines, especially if there is non-measurable disease in conventional Prevention:
imaging.8 The ESMO breast cancer guidelines suggest that tumour markers P261 Avoid breathing dust/fume/gas/mist/vapours/spray.
such as CA 15‑3 may be useful to evaluate response to treatment,
particularly in patients with non-measureable metastatic disease. A change
in tumour markers alone should not be used to initiate a change in P272 Contaminated work clothing should not be allowed out of
treatment.7 the workplace.
Test principle P280 Wear protective gloves.
Sandwich principle. Total duration of assay: 18 minutes.
Response:
▪ 1st incubation: 20 µL of sample are automatically prediluted 1:10 with
Diluent Universal. The antigen (in 20 µL of prediluted sample), a P333 + P313 If skin irritation or rash occurs: Get medical
biotinylated monoclonal CA 15‑3‑specific antibody, and a monoclonal advice/attention.
CA 15‑3‑specific antibody labeled with a ruthenium complexa) react to
form a sandwich complex. P362 + P364 Take off contaminated clothing and wash it before reuse.
▪ 2nd incubation: After addition of streptavidin-coated microparticles, the Disposal:
complex becomes bound to the solid phase via interaction of biotin and
streptavidin. P501 Dispose of contents/container to an approved waste
disposal plant.
Product safety labeling follows EU GHS guidance.

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Elecsys CA 15-3 II
Contact phone: all countries: +49-621-7590 Additional materials for cobas e 601 and cobas e 602 analyzers:
Avoid foam formation in all reagents and sample types (specimens, ▪ 04880340190, ProCell M, 2 x 2 L system buffer
calibrators and controls).
▪ 04880293190, CleanCell M, 2 x 2 L measuring cell cleaning
Reagent handling solution
The reagents in the kit have been assembled into a ready‑for‑use unit that ▪ 03023141001, PC/CC‑Cups, 12 cups to prewarm ProCell M and
cannot be separated. CleanCell M before use
All information required for correct operation is read in from the respective ▪ 03005712190, ProbeWash M, 12 x 70 mL cleaning solution for run
reagent barcodes. finalization and rinsing during reagent change
Storage and stability ▪ 12102137001, AssayTip/AssayCup, 48 magazines x 84 reaction
Store at 2‑8 °C. cups or pipette tips, waste bags
Do not freeze. ▪ 03023150001, WasteLiner, waste bags
Store the Elecsys reagent kit upright in order to ensure complete ▪ 03027651001, SysClean Adapter M
availability of the microparticles during automatic mixing prior to use.
Additional materials for all analyzers:
Stability: ▪ 11298500316, ISE Cleaning Solution/Elecsys SysClean,
unopened at 2‑8 °C up to the stated expiration date 5 x 100 mL system cleaning solution
after opening at 2‑8 °C 12 weeks Assay
For optimum performance of the assay follow the directions given in this
on the analyzers 5 weeks document for the analyzer concerned. Refer to the appropriate operator’s
manual for analyzer‑specific assay instructions.
Specimen collection and preparation
Resuspension of the microparticles takes place automatically prior to use.
Only the specimens listed below were tested and found acceptable. Read in the test-specific parameters via the reagent barcode. If in
Serum collected using standard sampling tubes or tubes containing exceptional cases the barcode cannot be read, enter the 15-digit sequence
separating gel. of numbers.
Li‑heparin, K2‑EDTA and K3‑EDTA plasma. Bring the cooled reagents to approximately 20 °C and place on the reagent
Criterion: Recovery within 90‑110 % of serum value or slope disk (20 °C) of the analyzer. Avoid foam formation. The system
0.9‑1.1 + intercept within < ± 2x analytical sensitivity (LDL) + coefficient of automatically regulates the temperature of the reagents and the
correlation > 0.95. opening/closing of the bottles.
Stable for 48 hours at 20‑25 °C, 5 days at 2‑8 °C, 90 days at Calibration
‑20 °C (± 5 °C). Freeze only once. Traceability: This method has been standardized against the Elecsys
The sample types listed were tested with a selection of sample collection CA 15‑3 assay. This in turn has been standardized against the
tubes that were commercially available at the time of testing, i.e. not all Enzymun‑Test CA 15‑3 method and CA 15‑3 RIA by Fujirebio Diagnostics.
available tubes of all manufacturers were tested. Sample collection systems Every Elecsys reagent set has a barcoded label containing specific
from various manufacturers may contain differing materials which could information for calibration of the particular reagent lot. The predefined
affect the test results in some cases. When processing samples in primary master curve is adapted to the analyzer using the relevant CalSet.
tubes (sample collection systems), follow the instructions of the tube
manufacturer. Calibration frequency: Calibration must be performed once per reagent lot
using fresh reagent (i.e. not more than 24 hours since the reagent kit was
Centrifuge samples containing precipitates before performing the assay. registered on the analyzer).
Do not use heat‑inactivated samples. Calibration interval may be extended based on acceptable verification of
Do not use samples and controls stabilized with azide. calibration by the laboratory.
Ensure the samples, calibrators and controls are at 20‑25 °C prior to Renewed calibration is recommended as follows:
measurement. ▪ after 12 weeks when using the same reagent lot
Due to possible evaporation effects, samples, calibrators and controls on
the analyzers should be analyzed/measured within 2 hours. ▪ after 7 days (when using the same reagent kit on the analyzer)
▪ as required: e.g. quality control findings outside the defined limits
Materials provided
See “Reagents – working solutions” section for reagents. Quality control
For quality control, use PreciControl Tumor Marker.
Materials required (but not provided)
In addition, other suitable control material can be used.
▪ 03045846122, CA 15-3 II CalSet, 4 x 1.0 mL
Controls for the various concentration ranges should be run individually at
▪ 11776452122, PreciControl Tumor Marker, for 4 x 3.0 mL least once every 24 hours when the test is in use, once per reagent kit, and
▪ 11732277122, Diluent Universal, 2 x 16 mL sample diluent or following each calibration.
03183971122, Diluent Universal, 2 x 36 mL sample diluent The control intervals and limits should be adapted to each laboratory’s
▪ General laboratory equipment individual requirements. Values obtained should fall within the defined
limits. Each laboratory should establish corrective measures to be taken if
▪ cobas e analyzer values fall outside the defined limits.
Additional materials for the cobas e 411 analyzer: If necessary, repeat the measurement of the samples concerned.
▪ 11662988122, ProCell, 6 x 380 mL system buffer Follow the applicable government regulations and local guidelines for
▪ 11662970122, CleanCell, 6 x 380 mL measuring cell cleaning quality control.
solution Calculation
▪ 11930346122, Elecsys SysWash, 1 x 500 mL washwater additive The analyzer automatically calculates the analyte concentration of each
sample (either in U/mL or kU/L).
▪ 11933159001, Adapter for SysClean
▪ 11706802001, AssayCup, 60 x 60 reaction cups Limitations - interference
The assay is unaffected by icterus (bilirubin < 1112 µmol/L or < 65 mg/dL),
▪ 11706799001, AssayTip, 30 x 120 pipette tips hemolysis (Hb < 1.9 mmol/L or < 3.0 g/dL), lipemia (Intralipid < 1500 mg/dL)
▪ 11800507001, Clean‑Liner and biotin (< 409 nmol/L or < 100 ng/mL).

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Elecsys CA 15-3 II
Criterion: Recovery within ± 10 % of initial value. Subjects < 25 25-50 > 50-200 > 200
Samples should not be taken from patients receiving therapy with high total U/mL U/mL U/mL U/mL
biotin doses (i.e. > 5 mg/day) until at least 8 hours following the last biotin N Classification in percent (%)
administration.
No interference was observed from rheumatoid factors up to a Renal failure 37 81 19 0 0
concentration of 1500 IU/mL. Urological 34 82 18 0 0
Typically, no high‑dose hook effectb) can be observed at CA 15‑3 diseases
concentrations up to 20000 U/mL. However, due to the heterogeneous
nature of the CA 15‑3 antigen a high-dose hook effect below this value Bacterial 27 96 4 0 0
cannot be completely excluded. In case of an unexpected low result, the infection
sample should be diluted 1:10 (refer to chapter “Dilution”) and tested again. Pregnancy 34 97 0 3 0
In vitro tests were performed on 28 commonly used pharmaceuticals. No
interference with the assay was found. • Patients with malignant diseases (others than breast):
In rare cases, interference due to extremely high titers of antibodies to Relative distribution of CA 15‑3 concentrations in individuals with
analyte‑specific antibodies, streptavidin or ruthenium can occur. These malignancy other than breast
effects are minimized by suitable test design.
Subjects < 25 25-50 > 50-200 > 200
For diagnostic purposes, the results should always be assessed in total U/mL U/mL U/mL U/mL
conjunction with the patient’s medical history, clinical examination and other
findings. N Classification in percent (%)
b) High-dose hook effect: A sample with a true concentration clearly above the measuring range, Stomach-Cac) 36 75 14 8 3
but found within the measuring range.
Limits and ranges Hepatocellular-Ca 37 59 32 3 5
Measuring range Lung-Ca 38 82 13 5 0
1.00-300 U/mL (defined by the lower detection limit and the maximum of the Ovarian-Ca 34 47 21 29 3
master curve). Values below the lower detection limit are reported as
< 1.00 U/mL. Values above the measuring range are reported as Gynecological-Ca 5 40 20 40 0
> 300 U/mL (or up to 3000 U/mL for 10‑fold diluted samples). Prostate-Ca 48 79 17 4 0
Lower limits of measurement
Colorectal-Ca 40 93 8 0 0
Lower detection limit of the test
Lower detection limit: < 1.00 U/mL Pancreatic-Ca 40 65 33 3 0
The lower detection limit represents the lowest measurable analyte level c) Ca = Carcinoma
that can be distinguished from zero. It is calculated as the value lying two • Patients with breast cancer:
standard deviations above that of the lowest standard (master calibrator, Relative distribution of CA 15‑3 concentrations in patients with breast
standard 1 + 2 SD, repeatability study, n = 21). malignancy. The staging of patients according to UICC criteria was
Dilution performed at primary diagnosis before any treatment. The patients
Use Diluent Universal for automatic sample predilution. Samples with diagnosed with recurrent disease had developed metastases (M1).
CA 15‑3 concentrations above the measuring range despite predilution
must be diluted 1:10 with Diluent Universal (either manually for all analyzers Subjects < 25 25-50 > 50-200 > 200
or automatically by the cobas e 601 and cobas e 602 analyzers). The total U/mL U/mL U/mL U/mL
concentration of the diluted sample must be > 30 U/mL. N Classification in percent (%)
After manual dilution, multiply the result by the dilution factor.
UICC I 56 88 12 0 0
After dilution by the analyzers, the cobas e 601 and cobas e 602 software
automatically takes the dilution into account when calculating the sample UICC II 126 85 13 2 0
concentration. UICC III 77 53 30 14 3
Expected values UICC IV 24 25 17 37 21
• Healthy subjects:
Recurrent 75 15 25 36 24
Results of a reference range study using a panel of samples from 374 Disease
apparently healthy non‑pregnant females (Roche study No. RD000788)
Each laboratory should investigate the transferability of the expected values
Percentile (%) U/mL Confidence interval (U/mL) to its own patient population and if necessary determine its own reference
95 26.2 25.2-27.9 ranges.
97.5 28.5 26.7-34.5 Specific performance data
Representative performance data on the analyzers are given below.
99 34.5 28.7-57.8 Results obtained in individual laboratories may differ.
• Patients with benign diseases and pregnant women: Precision
Relative distribution of CA 15‑3 concentrations in patients with benign Precision was determined using Elecsys reagents, pooled human sera and
disease and pregnant women (Roche study No. B00P018) controls in accordance with a modified protocol (EP5‑A) of the CLSI
(Clinical and Laboratory Standards Institute): 6 times daily for 10 days
Subjects < 25 25-50 > 50-200 > 200 (n = 60); repeatability on MODULAR ANALYTICS E170 analyzer, n = 21.
total U/mL U/mL U/mL U/mL The following results were obtained:
N Classification in percent (%)
Gastrointestinal 109 84 16 0 0
Breast 58 88 12 0 0
Gynecological 42 83 12 5 0
diseases

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Elecsys CA 15-3 II
cobas e 411 analyzer recurrence
Repeatability Intermediate yes 21 14
precision
The corresponding results for positive predictive value (PPV) and negative
Sample Mean SD CV SD CV predictive value (NPV) with the 95 % confidence interval for a cutoff of 25 %
U/mL U/mL % U/mL % CA 15‑3 increase as derived from the table are:
Human serum 1 38.0 0.81 2.1 1.38 3.6 Positive predictive value: 40 % (24‑58 %)
Negative predictive value: 90 % (90‑99 %)
Human serum 2 85.5 2.72 3.2 3.66 4.3
y
Human serum 3 179 4.56 2.6 6.60 3.7 100
PreciControl TMd)1 24.5 0.62 2.5 0.87 3.6
PreciControl TM2 67.6 2.48 3.7 2.83 4.2
d) TM = Tumor Marker 80

cobas e 601 and cobas e 602 analyzers

Repeatability Intermediate precision
Sample Mean SD CV Mean SD CV 60

U/mL U/mL % U/mL U/mL %

AUC: 88.0% (77.1%–98.8%)
Human serum 1 18.9 0.24 1.3 20.1 0.64 3.2
Human serum 2 76.4 1.07 1.4 79.0 3.08 3.9 40

Human serum 3 148 1.72 1.2 156 7.75 5.0

PreciControl TM1 20.3 0.24 1.2 21.3 0.89 4.2
PreciControl TM2 47.6 0.70 1.5 49.6 1.82 3.7 20

Method comparison
A comparison of the Elecsys CA 15‑3 II assay (y) with the Elecsys CA 15‑3
assay (x) using clinical samples gave the following correlations: 0
Number of samples measured: 52
100 80 60 40 20 0
Passing/Bablok regression9
x
Slope: 1.06 (95 % confidence range: 1.01‑1.15)
Intercept: 2.66 (95 % confidence range: ‑0.99‑5.97)
x = Specificity (%); y = Sensitivity (%)
Coefficient of correlation: 0.965
The sample concentrations were between 6 and 280 U/mL. Figure 1: ROC curve: breast cancer recurrence by relative change CA 15‑3
to baseline
Analytical specificity
The Elecsys CA 15‑3 II tumor marker assay is based on the monoclonal The area under the curve (AUC) was 0.8796 (95 % CI: 0.7709‑0.9884)
115D8 and DF3 antibodies which are only available from Fujirebio Monitoring response to therapy
Diagnostics, its licensees and its representatives. The performance
characteristics of test procedures using these antibodies cannot be Fifteen (15) patients with metastatic breast cancer underwent treatment and
assumed for test methods using other antibodies. response to therapy was assessed by clinical criteria. A total of 72
assessments (median 4 assessments per patient) were made. Fourteen
Clinical performance in follow-up (14) patients had a response to therapy.
Patients diagnosed with breast cancer were examined in a retrospective
study (at least 4 samples/patient during follow-up study) and classified as 2 x 2 table for response to therapy
recurrence [yes/no] after no evidence of breast cancer or response to response
treatment [yes/no] after breast cancer metastasis based on the clinical
information (medical imaging and other clinical investigations). The CA 15‑3 CA15-3 decrease > 25 % no yes
concentrations were measured in parallel. The ROC (receiver-operating no 25 19
characteristics) analysis of relative CA15‑3 change to determine breast
cancer recurrence/ therapy response in metastasized breast cancer was yes 1 12
done to show clinical accuracy at various cut-offs and to summarize the
cutoff-independent clinical performance in a ROC plot and the related AUC The corresponding results for positive predictive value (PPV) and negative
(area under the curve). predictive value (NPV) with the 95 % confidence interval as derived from
the table are:
Early detection of recurrence
Positive predictive value: 92 % (64‑100 %)
Forty (40) patients treated for stage II or III breast cancer were followed for
up to 1351 days (median 105 days). A total of 172 samples (median 4 Negative predictive value: 57 % (41‑72 %)
samples per patient) were assessed for recurrence of disease over the
follow-up period. Recurrence was defined as the presence of clinical
symptoms in women with no evidence of disease at the beginning of the
follow-up period. Eighteen (18) patients experienced recurrence of disease.
2 x 2 table for early detection of recurrence:
recurrence
CA15-3 increase > 25% no yes
no 93 4

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Elecsys CA 15-3 II
y
CA 15‑3 is a registered trademark of
100
Fujirebio Diagnostics, Inc.

Symbols
80
Roche Diagnostics uses the following symbols and signs in addition to
those listed in the ISO 15223‑1 standard (for USA: see dialog.roche.com for
definition of symbols used):
Contents of kit
60
Analyzers/Instruments on which reagents can be used
Reagent
AUC: 86.1% (76.2%–96.0%)
Calibrator
40
Volume after reconstitution or mixing
GTIN Global Trade Item Number
20 COBAS, COBAS E, ELECSYS and PRECICONTROL are trademarks of Roche. INTRALIPID is a trademark of
Fresenius Kabi AB.
All other product names and trademarks are the property of their respective owners.
Additions, deletions or changes are indicated by a change bar in the margin.
© 2020, Roche Diagnostics
0

100 80 60 40 20 0
x
Roche Diagnostics GmbH, Sandhofer Strasse 116, D-68305 Mannheim
www.roche.com
x = Specificity (%); y = Sensitivity (%)
Figure 2: ROC curve: breast cancer response to therapy by relative
change CA 15‑3 to baseline
The area under the curve (AUC) was 0.8610 (95% CI: 0.7623-0.9598).
References
1 Duffy MJ. CA 15-3 and related mucins as circulating markers in breast
cancer. Ann Clin Biochem,1999;36:579-586.
2 Hilkens J, Buijs F, Hilgers J, et al. Monoclonal antibodies against
human milk-fat globule membranes detecting differentiation antigens of
the mammary gland. Prot Biol Fluids 1982;29:813-816.
3 Hilkens J, Buijs F, Hilgers J, et al. Monoclonal antibodies against
human milk-fat globule membranes detecting differentiation antigens of
the mammary gland and its tumors. Int J Cancer 1984;34:197-206.
4 Kufe D, Inghirami G, Abe M, et al. Differential reactivity of a novel
monoclonal antibody (DF3) with human malignant versus benign breast
tumor. Hybridoma 1984;(3):223-232.
5 Sekine H, Ohno T, Kufe DW. Purification and characterization of a high
molecular weight glycoprotein detectable in human milk and breast
carcinomas. J Immunol 1985;135(5):3610-3615.
6 Ponnusamy MP, Seshacharyulu P, Lakshmanan I, et al. Emerging role
of mucins in epithelial to mesenchymal transition. Curr cancer drug
targets 2013;13(9):945-56.
7 Duffy MJ, Walsh S, McDermott EW, et al. Chapter One - Biomarkers in
Breast Cancer: Where Are We and Where Are We Going?, In: Gregory
S. Makowski, Editor(s), Advances in Clinical Chemistry, Elsevier,
2015;71: 1-23, ISSN 0065-2423, ISBN 9780128022566.
8 Cardoso F, et al. ESMO Guidelines for advanced breast cancer. Annals
of Oncology,2018;29: 1634-1657.
9 Bablok W, Passing H, Bender R, et al. A general regression procedure
for method transformation. Application of linear regression procedures
for method comparison studies in clinical chemistry, Part III.
J Clin Chem Clin Biochem 1988 Nov;26(11):783-790.
For further information, please refer to the appropriate operator’s manual for
the analyzer concerned, the respective application sheets, the product
information and the Method Sheets of all necessary components (if
available in your country).
A point (period/stop) is always used in this Method Sheet as the decimal
separator to mark the border between the integral and the fractional parts of
a decimal numeral. Separators for thousands are not used.

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