04528123190V10

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Tina-quant Hemoglobin A1c Gen.2
• Indicates cobas c systems on which reagents can be used
Order information Roche/Hitachi cobas c systems
Tina-quant Hemoglobin A1c Gen.2 cobas c 311 cobas c 501/502
150 tests Cat. No. 04528123 190 System-ID 07 6850 2 • •
C.f.a.s. HbA1c (3 × 2 mL) Cat. No. 04528417 190 Code 674
HbA1c Control N (4 × 0.5 mL) Cat. No. 20764833 322 Code 357
HbA1c Control P (4 × 0.5 mL) Cat. No. 20764841 322 Code 358
PreciControl HbA1c norm (4 × 1 mL) Cat. No. 05479207 190 Code 208
PreciControl HbA1c path (4 × 1 mL) Cat. No. 05912504 190 Code 209
Hemolyzing Reagent Gen.2 (51 mL)* Cat. No. 04528182 190 System-ID 07 6873 1
HbA1c Hemolyzing Reagent for Tina-quant HbA1c
Cat. No. 11488457 122 For Hemolysate Application only
(1000 mL)

* The value encoded in the instrument settings is 45 mL to account for the dead volume of the bottles.
English IFCC standardization (recalculated acc. to ref. 8)
System information • Estimated average glucose [mmol/L] = 0.146 × HbA1c (mmol/mol) + 0.834
or
Whole Blood Application - Standardized according to IFCC transferable • Estimated average glucose [mg/dL] = 2.64 × HbA1c (mmol/mol) + 15.03
to DCCT/NGSP
Standardization acc. to DCCT/NGSP8
HB-W2: ACN 870 Hemoglobin (Hb)
• Estimated average glucose [mmol/L] = 1.59 × HbA1c (%) - 2.59
A1-W2: ACN 880 Hemoglobin A1c (HbA1c) or
RWI2: ACN 890 Ratio (% HbA1c IFCC; not • Estimated average glucose [mg/dL] = 28.7 × HbA1c (%) - 46.7
recommended for patient reporting)
The risk of diabetic complications, such as diabetic nephropathy and
A1CD2: ACN 952 Hemolyzing reagent
retinopathy, increases with poor metabolic control. In accordance with its
Hemolysate Application - Standardized according to IFCC transferable function as an indicator for the mean blood glucose level, HbA1c predicts
to DCCT/NGSP the development of diabetic complications in diabetes patients.3,5
HB-H2: ACN 840 Hemoglobin (Hb) For monitoring of long term glycemic control, testing every 3 to
A1-H2: ACN 850 Hemoglobin A1c (HbA1c) 4 months is generally sufficient. In certain clinical situations, such as
RHI2: ACN 860 Ratio (% HbA1c IFCC; not gestational diabetes, or after a major change in therapy, it may be
recommended for patient reporting) useful to measure HbA1c in 2 to 4 week intervals.7
A1CD2: ACN 952 Hemolyzing reagent Test principle10,11,12
Intended use This method uses TTAB* as the detergent in the hemolyzing reagent to
In vitro test for the quantitative determination of mmol/mol hemoglobin A1c eliminate interference from leukocytes (TTAB does not lyse leukocytes).
(IFCC) and % hemoglobin A1c (DCCT/NGSP) in whole blood or hemolysate Sample pretreatment to remove labile HbA1c is not necessary.
on Roche/Hitachi cobas c systems. HbA1c determinations are useful for All hemoglobin variants which are glycated at the β-chain N-terminus and
monitoring of long-term blood glucose control in individuals with diabetes which have antibody-recognizable regions identical to that of HbA1c are
mellitus. Moreover, this test is to be used as an aid in diagnosis of diabetes measured by this assay. Consequently, the metabolic state of patients
and identifying patients who may be at risk for developing diabetes. having uremia or the most frequent hemoglobinopathies (HbAS, HbAC,
HbAE) can be determined using this assay.13,14
Summary1,2,3,4,5,6,7,8,9
*TTAB = Tetradecyltrimethylammonium bromide
Hemoglobin (Hb) consists of four protein subunits, each containing a heme
Hemoglobin A1c
moiety, and is the red-pigmented protein located in the erythrocytes. Its main
The HbA1c determination is based on the turbidimetric inhibition
function is to transport oxygen and carbon dioxide in blood. Each Hb molecule
immunoassay (TINIA) for hemolyzed whole blood.
is able to bind four oxygen molecules. Hb consists of a variety of subfractions
and derivatives. Among this heterogeneous group of hemoglobins HbA1c is • Sample and addition of R1 (Antibody reagent):
one of the glycated hemoglobins, a subfraction formed by the attachment Glycohemoglobin (HbA1c) in the sample reacts with anti-HbA1c
of various sugars to the Hb molecule. HbA1c is formed in two steps by the antibody to form soluble antigen-antibody complexes. Since the
nonenzymatic reaction of glucose with the N-terminal amino group of the specific HbA1c antibody site is present only once on the HbA1c
β-chain of normal adult Hb (HbA). The first step is reversible and yields labile molecule, complex formation does not take place.
HbA1c. This is rearranged to form stable HbA1c in a second reaction step. • Addition of R2 (Polyhapten reagent) and start of reaction:
In the erythrocytes, the relative amount of HbA converted to stable The polyhaptens react with excess anti-HbA1c antibodies to
HbA1c increases with the average concentration of glucose in the form an insoluble antibody-polyhapten complex which can
blood. The conversion to stable HbA1c is limited by the erythrocyte’s be measured turbidimetrically.
life span of approximately 100 to 120 days. As a result, HbA1c reflects Hemoglobin
the average blood glucose level during the preceding 2 to 3 months. Liberated hemoglobin in the hemolyzed sample is converted to a derivative
HbA1c is thus suitable to monitor long-term blood glucose control in having a characteristic absorption spectrum which is measured bichromatically
individuals with diabetes mellitus. Glucose levels closer to the time of during the preincubation phase (sample + R1) of the above immunological
the assay have a greater influence on the HbA1c level.1 reaction. A separate Hb reagent is consequently not necessary.
The approximate relationship between HbA1c and mean blood glucose Ratio definition
values during the preceding 2 to 3 months was analyzed in several The final result is expressed as mmol/mol HbA1c or % HbA1c and is
studies. A recent study obtained the following correlation: calculated from the HbA1c/Hb ratio as follows:

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A1C-2
Tina-quant Hemoglobin A1c Gen.2
Protocol 1 (% HbA1c acc. to IFCC; not recommended for patient result 3. Dilute the sample with Hemolyzing Reagent for Tina-quant HbA1c
reporting): (Cat. No. 11488457 122) in the ratio 1:101 (1+100) using one
HbA1c (%) = (HbA1c/Hb) × 100 of the following pipetting schemes.
Protocol 2 (% HbA1c acc. to DCCT/NGSP): Pipette into tubes:
HbA1c (%) = (HbA1c/Hb) × 91.5 + 2.15 HbA1c Hemolyzing Reagent 500 µL 1000 µL 2000 µL
Protocol 3 (mmol/mol HbA1c acc. to IFCC): for Tina-quant HbA1c
HbA1c (mmol/mol) = (HbA1c/Hb) × 1000 Specimen (patient or control) 5 µL 10 µL 20 µL

Reagents – working solutions 4. Mix using a vibration mixer or by gentle swirling.

5. The hemolysate can be used after the solution has changed color
R1 Antibody reagent from red to brownish-green (approx. 1-2 min).
MES buffer: 0.025 mol/L; TRIS buffer: 0.015 mol/L, pH 6.2; HbA1c
antibody (bovine serum) ≥ 0.5 mg/mL; stabilizers; preservatives (liquid) Stability of the hemolysate:15 4 hours at 15-25 °C
R2 Polyhapten reagent 24 hours at 2-8 °C
MES buffer: 0.025 mol/L; TRIS buffer: 0.015 mol/L, pH 6.2; HbA1c 6 months at (-15)-(-25) °C
polyhapten: ≥ 8 µg/mL; stabilizers; preservatives (liquid)
Materials provided
Precautions and warnings See “Reagents - working solutions” section for reagents.
For in vitro diagnostic use.
Exercise the normal precautions required for handling all laboratory reagents. Materials required (but not provided)
Safety data sheet available for professional user on request. See “Order information” section.
Disposal of all waste material should be in accordance with local guidelines. General laboratory equipment

Reagent handling Assay

Ready for use. For optimum performance of the assay, follow the directions given in
this document for the analyzer concerned. Refer to the appropriate
Storage and stability operator’s manual for analyzer-specific assay instructions.
The performance of applications not validated by Roche is not
A1C-2
warranted and must be defined by the user.
Shelf life at 2-8 °C: See expiration date on
cobas c pack label. Whole Blood application for Hb (HB-W2) and HbA1c (A1-W2)
On-board in use and refrigerated on the analyzer: 4 weeks
cobas c 311 test definition Hb (HB-W2)
Assay type 1 Point
Hemolyzing reagent Reaction time / Assay points 10 / 23
Shelf life at 2-8 °C: See expiration date on Wavelength (sub/main) 660/376 nm
pack label Reaction direction Increase
When storing at temperatures below 3 °C, the reagent may become cloudy. Unit g/dL
This has no effect on the function of the reagent and is reversible at higher Reagent pipetting Diluent (H2O)
temperatures. It is therefore recommended to equilibrate the reagent at room R1 120 µL –
temperature for approximately 10 minutes and mix thoroughly before use. –
R3 24 µL
On-board in use and refrigerated on the analyzer: 4 weeks
Sample volumes Sample Sample dilution
Specimen collection and preparation Sample Diluent
For specimen collection and preparation, only use suitable (Hemolyzing
tubes or collection containers. reagent)
Only the specimens listed below were tested and found acceptable. Normal 5 µL 2 µL 180 µL
Venous or capillary blood with anticoagulant. Decreased 5 µL 2 µL 180 µL
The only acceptable anticoagulants are Li-heparin, K2-EDTA, Increased 5 µL 2 µL 180 µL
K3-EDTA and potassium fluoride/Na2-EDTA.
The sample types listed were tested with a selection of sample collection tubes cobas c 311 test definition HbA1c (A1-W2)
that were commercially available at the time of testing, i.e. not all available Assay type 2 Point End
tubes of all manufacturers were tested. Sample collection systems from Reaction time / Assay points 10 / 23-57
various manufacturers may contain differing materials which could affect Wavelength (sub/main) 660/340 nm
the test results in some cases. When processing samples in primary tubes Reaction direction Increase
(sample collection systems), follow the instructions of the tube manufacturer. Unit g/dL
Stability:15 3 days at 15-25 °C Reagent pipetting Diluent (H2O)
7 days at 2-8 °C R1 120 µL –
6 months at (-15)-(-25) °C R3 24 µL –
Freeze only once. Mix specimen thoroughly before use. Sample volumes Sample Sample dilution
Sample Diluent
Hemolysate preparation for Hemolysate Application (Hemolyzing
1. Allow blood specimen and Hemolyzing Reagent for Tina-quant HbA1c reagent)
to equilibrate at room temperature before use.
Normal 5 µL 2 µL 180 µL
2. Moderately mix the sample immediately prior to pipetting to
Decreased 5 µL 2 µL 180 µL
ensure a homogeneous mixture of erythrocytes. Take care
to avoid the formation of foam. Increased 5 µL 2 µL 180 µL

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A1C-2
Tina-quant Hemoglobin A1c Gen.2
cobas c 501/502 test definition Hb (HB-W2) It is recommended to report % HbA1c values to one decimal place
Assay type 1 Point and mmol/mol HbA1c values without decimal point, which can be
Reaction time / Assay points 10 / 34 entered in the editable field “expected values”.
Wavelength (sub/main) 660/376 nm
Hemolysate Application for Hb (HB-H2) and HbA1c (A1-H2)
Reaction direction Increase
Unit g/dL cobas c 311 test definition Hb (HB-H2)
Reagent pipetting Diluent (H2O) Assay type 1 Point
R1 120 µL – Reaction time / Assay points 10 / 23
R3 24 µL – Wavelength (sub/main) 660/376 nm
Sample volumes Sample Sample dilution Reaction direction Increase
Sample Diluent Unit g/dL
(Hemolyzing Reagent pipetting Diluent (H2O)
reagent) R1 120 µL –
Normal 5 µL 2 µL 180 µL R3 24 µL –
Decreased 5 µL 2 µL 180 µL
Increased 5 µL 2 µL 180 µL Sample volumes Sample Sample dilution
Sample Diluent
cobas c 501/502 test definition HbA1c (A1-W2)
Assay type 2 Point End Normal 5 µL – –
Reaction time / Assay points 10 / 34-70 Decreased 5 µL – –
Wavelength (sub/main) 660/340 nm Increased 5 µL – –
Reaction direction Increase
Unit g/dL cobas c 311 test definition HbA1c (A1-H2)
Assay type 2 Point End
Reagent pipetting Diluent (H2O) Reaction time / Assay points 10 / 23-57
R1 120 µL –
Wavelength (sub/main) 660/340 nm
R3 24 µL –
Reaction direction Increase
Sample volumes Sample Sample dilution Unit g/dL
Sample Diluent Reagent pipetting Diluent (H2O)
(Hemolyzing –
R1 120 µL
reagent)
R3 24 µL –
Normal 5 µL 2 µL 180 µL
Decreased 5 µL 2 µL 180 µL Sample volumes Sample Sample dilution
Increased 5 µL 2 µL 180 µL Sample Diluent
Normal 5 µL – –
Ratio definition for mmol/mol HbA1c and % HbA1c calculation
Decreased 5 µL – –
Protocol 1 (% HbA1c acc. to IFCC, not recommended for – –
Increased 5 µL
patient result reporting):
Abbreviated ratio name RWI2 (890) cobas c 501/502 test definition Hb (HB-H2)
Equation (A1-W2/HB-W2) × 100 Assay type 1 Point
Unit % Reaction time / Assay points 10 / 34
Wavelength (sub/main) 660/376 nm
Protocol 2 (% HbA1c acc. to DCCT/NGSP):
Reaction direction Increase
Abbreviated ratio name RWD2 Unit g/dL
Equation (A1-W2/HB-W2) × 91.5 + 2.15
Reagent pipetting Diluent (H2O)
Unit % –
R1 120 µL
Protocol 1 is already implemented in the application (ACN 890). However a R3 24 µL –
patient result reporting in % HbA1c (IFCC) units is not recommended. The
common % HbA1c (DCCT/NGSP) units can be achieved by modifying the Sample volumes Sample Sample dilution
application according to protocol 2. The formula (ACN 890) can be modified Sample Diluent
by using the Administrator Level/EDIT Button accordingly. Normal 5 µL – –
Protocol 3 (mmol/mol HbA1c acc. to IFCC): Decreased 5 µL – –
The additional mmol/mol HbA1c (IFCC) results can be automatically calculated Increased 5 µL – –
from the analyzer by defining an additional calculated test:
Sample type Suprnt cobas c 501/502 test definition HbA1c (A1-H2)
Unit of Measure mM/M Assay type 2 Point End
Report Name HbA1c Gen.2 IFCC Reaction time / Assay points 10 / 34-70
ITEM RWM2 Wavelength (sub/main) 660/340 nm
Formula (A1-W2/HB-W2) × 1000 Reaction direction Increase
Unit g/dL
This equation must be entered under “Utility > Calculated Test” on
Roche/Hitachi cobas c 311 and Roche/Hitachi cobas c 501/502 analyzers. Reagent pipetting Diluent (H2O)
R1 120 µL –
The ratio for HbA1c (mmol/mol HbA1c acc. to IFCC and % HbA1c acc. to –
R3 24 µL
DCCT/NGSP) will be automatically calculated after result output of both tests.
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A1C-2
Tina-quant Hemoglobin A1c Gen.2
Sample volumes Sample Sample dilution In addition, other suitable control material can be used.
Sample Diluent The control intervals and limits should be adapted to each laboratory’s
individual requirements. Values obtained should fall within the defined
Normal 5 µL – –
limits. Each laboratory should establish corrective measures to be
Decreased 5 µL – – taken if values fall outside the defined limits.
Increased 5 µL – – Follow the applicable government regulations and local guidelines
Ratio definition for mmol/mol HbA1c and % HbA1c calculation for quality control.
Protocol 1 (% HbA1c acc. to IFCC, not recommended for Calculation for Whole Blood and Hemolysate Application
patient result reporting): Instrument factor
Abbreviated ratio name RHI2 (860) To improve the fit of the nonlinear HbA1c calibration curve, a constant and
Equation (A1-H2/HB-H2) × 100 lot independent offset of 0.6 g/dL was added to all calibrator values. This
Unit % offset is already included in the C.f.a.s. HbA1c calibrator target values for
cobas c analyzers and finally needs to be subtracted from the analyzer’s
Protocol 2 (% HbA1c acc. to DCCT/NGSP): results. Enter the instrument factor of the absolute HbA1c (A1-W2 or
Abbreviated ratio name RHD2 A1-H2) assay on Roche cobas c analyzers as follows:
Equation (A1-H2/HB-H2) × 91.5 + 2.15 Instrument factor for Whole Blood Application
Unit % Calibration => Status Screen => Instrument Factor => Instrument Factor
Protocol 1 is already implemented in the application (ACN 860). However a Window => HbA1c (A1-W2) => a = 1.0; b = – 0.6 => update => okay
patient result reporting in % HbA1c (IFCC) units is not recommended. The Instrument factor for Hemolysate Application
common % HbA1c (DCCT/NGSP) units can be achieved by modifying the Calibration => Status Screen => Instrument Factor => Instrument Factor
application according to protocol 2. The formula (ACN 860) can be modified Window => HbA1c (A1-H2) => a = 1.0; b = – 0.6 => update => okay
by using the Administrator Level/EDIT Button accordingly.
Protocol 3 (mmol/mol HbA1c acc. to IFCC): Hb, HbA1c
The additional mmol/mol HbA1c (IFCC) results can be automatically calculated Roche/Hitachi cobas c systems automatically calculate the analyte
from the analyzer by defining an additional calculated test: concentration of each sample.

Sample type Suprnt HbA1c ratio calculation

For calculation of the mmol/mol HbA1c value (IFCC) and the % HbA1c
Unit of Measure mM/M
value (DCCT/NGSP), refer to the Test principle and Ratio definition for
Report Name HbA1c Gen.2 IFCC mmol/mol HbA1c and % HbA1c calculation sections in this method sheet.
ITEM RHM2
Formula (A1-H2/HB-H2) × 1000 Limitations - interference for Whole Blood and Hemolysate
This equation must be entered under “Utility > Calculated Test” on Application13,14,18,19,20,21,22,23,24
Roche/Hitachi cobas c 311 and Roche/Hitachi cobas c 501/502 analyzers. 1. For diagnostic purposes, mmol/mol HbA1c values (IFCC) and % HbA1c
The ratio for HbA1c (mmol/mol HbA1c acc. to IFCC and % HbA1c acc. to values (DCCT/NGSP) should be used in conjunction with information
DCCT/NGSP) will be automatically calculated after result output of both tests. from other diagnostic procedures and clinical evaluations.
It is recommended to report % HbA1c values to one decimal place 2. The test is designed only for accurate and precise measurement of
and mmol/mol HbA1c values without decimal point, which can be mmol/mol HbA1c (IFCC) and % HbA1c (DCCT/NGSP). The individual
entered in the editable field “expected values”. results for total Hb and HbA1c concentration should not be reported.
3. As a matter of principle, care must be taken when interpreting any HbA1c
Calibration for Whole Blood and Hemolysate Application
result from patients with Hb variants. Abnormal hemoglobins might affect
Enter the assigned lot-specific and application-specific value of the calibrator.
the half life of the red cells or the in vivo glycation rates. In these cases even
Use the appropriate C.f.a.s. HbA1c calibrator only.
analytically correct results do not reflect the same level of glycemic control
The cobas c Hemolyzing Reagent Gen.2 (51 mL; Cat. No. 04528182 190)
that would be expected in patients with normal hemoglobin.23 Whenever
needs to be available on the analyzer.
it is suspected that the presence of an Hb variant (e.g. HbSS, HbCC or
Otherwise the calibration cannot be performed.
HbSC), affects the correlation between the HbA1c value and glycemic
Hb control, HbA1c must not be used for the diagnosis of diabetes mellitus.
Calibrators S1-S2: C.f.a.s. HbA1c 4. Any cause of shortened erythrocyte survival or decrease in mean
Calibration mode Linear erythrocyte age will reduce exposure of erythrocytes to glucose with
HbA1c a consequent decrease in mmol/mol HbA1c values (IFCC) and %
Calibrators S1-S6: C.f.a.s. HbA1c HbA1c values (DCCT/NGSP), even though the time-averaged blood
Calibration mode RCM glucose level may be elevated. Causes of shortened erythrocyte lifetime
might be hemolytic anemia or other hemolytic diseases, homozygous
Calibration frequency Hb and HbA1c: full calibration is recommended
sickle cell trait, pregnancy, recent significant or chronic blood loss,
- after 29 days during shelf life etc. Similarly, recent blood transfusions can alter the mmol/mol
- after reagent lot change HbA1c values (IFCC) and % HbA1c values (DCCT/NGSP). Caution
- as required following quality control should be used when interpreting the HbA1c results from patients
procedures with these conditions. HbA1c must not be used for the diagnosis of
Always calibrate both assays (Hb and HbA1c) diabetes mellitus in the presence of such conditions.
in parallel. Automatic calibration at QC failure 5. Glycated HbF is not detected by the assay as it does not contain the
should be deactivated. glycated β-chain that characterizes HbA1c. However, HbF is measured
in the Total Hb assay and as a consequence, specimens containing high
Traceability: This method has been standardized against the approved IFCC amounts of HbF (> 10 %) may result in lower than expected mmol/mol
reference method for the measurement of HbA1c in human blood16,17 and HbA1c values (IFCC) and % HbA1c values (DCCT/NGSP).14,24
can be transferred to results traceable to DCCT/NGSP by calculation. 6. mmol/mol HbA1c values (IFCC) and % HbA1c values (DCCT/NGSP)
Quality control for Whole Blood and Hemolysate Application are not suitable for diagnosis of gestational diabetes.25
For quality control, use control materials as listed in the 7. In very rare cases of rapidly evolving type 1 diabetes the increase
“Order information” section. of HbA1c values might be delayed compared to the acute increase
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A1C-2
Tina-quant Hemoglobin A1c Gen.2
in glucose concentrations. In these conditions diabetes mellitus 2 to 3 months or longer. According to the recommendations of the American
must be diagnosed based on plasma glucose concentrations Diabetes Association values above 48 mmol/mol HbA1c (IFCC) or 6.5 %
and/or the typical clinical symptoms.25 HbA1c (DCCT/NGSP) are suitable for the diagnosis of diabetes mellitus.25,29
Interference Criterion: Recovery within ± 10 % of initial value at a Patients with HbA1c values in the range of 39 - 46 mmol/mol HbA1c (IFCC) or
decision level of 42 mmol/mol HbA1c (IFCC). 5.7 % - 6.4 % HbA1c (DCCT/NGSP) may be at risk of developing diabetes.25,29
HbA1c levels may reach 195 mmol/mol HbA1c (IFCC) or 20 % HbA1c
Icterus: No significant interference up to an I index of 60 for conjugated
(DCCT/NGSP) and more in poorly controlled diabetes. Therapeutic
bilirubin and 60 for unconjugated bilirubin (approximate conjugated bilirubin
action is suggested at levels above 64 mmol/mol HbA1c (IFCC) or
concentration: 60 mg/dL or 1000 µmol/L, approximate unconjugated
8 % HbA1c (DCCT/NGSP). Diabetes patients with HbA1c levels below
bilirubin concentration: 60 mg/dL or 1000 µmol/L).
53 mmol/mol HbA1c (IFCC) or 7 % HbA1c (DCCT/NGSP) meet the
Lipemia (Intralipid): No significant interference up to an Intralipid goal of the American Diabetes Association.21,30
concentration of 500 mg/dL (cobas c 501/502 analyzers) and 400 HbA1c levels below the established reference range may indicate
mg/dL (cobas c 311 analyzer). There is poor correlation between recent episodes of hypoglycemia, the presence of Hb variants,
triglyceride concentration and turbidity. or shortened lifetime of erythrocytes.
Glycemia: No significant interference up to a glucose level of 55.5 mmol/L Each laboratory should investigate the transferability of the expected values to
(1000 mg/dL). A fasting sample is not required. its own patient population and if necessary determine its own reference ranges.
Rheumatoid factors: No significant interference up to a rheumatoid Specific performance data
factor level of 750 IU/mL. Representative performance data on the analyzers are given below.
Drugs: No interference was found at therapeutic concentrations Results obtained in individual laboratories may differ.
using common drug panels.26,27
Other: No cross reactions with HbA0, HbA1a, HbA1b, acetylated hemoglobin, Precision
carbamylated hemoglobin, glycated albumin and labile HbA1c were Precision was determined using human samples and controls in an
found for the anti-HbA1c antibodies used in this kit. internal protocol with repeatability* (n = 21) and intermediate precision**
For diagnostic purposes, the results should always be assessed in conjunction (3 aliquots per run, 1 run per day, 21 days). The following results were
with the patient’s medical history, clinical examination and other findings. obtained (data based on DCCT/NGSP values):
ACTION REQUIRED Whole Blood Application:
Special Wash Programming: The use of special wash steps is mandatory Repeatability* Mean SD CV
when certain test combinations are run together on Roche/Hitachi cobas c % HbA1c % HbA1c %
systems. The latest version of the carry-over evasion list can be found with Control Level 1 5.9 0.09 1.5
the NaOHD/SMS/Multiclean/SCCS or the NaOHD/SMS/SmpCln1 + 2/SCCS Control Level 2 10.5 0.11 1.1
Method Sheets. For further instructions refer to the operator’s manual. Human sample 1 5.2 0.04 0.8
cobas c 502 analyzer: All special wash programming necessary for avoiding
Human sample 2 9.0 0.11 1.2
carry-over is available via the cobas link, manual input is not required.
Where required, special wash/carry-over evasion programming must
Intermediate Mean SD CV
be implemented prior to reporting results with this test.
precision** % HbA1c % HbA1c %
Limits and ranges Control Level 1 6.0 0.12 1.9
Measuring range Control Level 2 10.5 0.21 2.0
Hemoglobin: 4-35 g/dL Human sample 3 5.6 0.07 1.3
HbA1c: 0.3-2.5 g/dL Human sample 4 11.8 0.20 1.7
The technical limit in the instrument setting is defined as 0.9-3.1 g/dL
due to the instrument factor for HbA1c (b = - 0.6; see above chapter
Hemolysate Application:
Calculation for Whole Blood and Hemolysate Application).
Repeatability* Mean SD CV
This corresponds to a measuring range of 23-189 mmol/mol HbA1c at a
% HbA1c % HbA1c %
typical hemoglobin concentration of 13.2 g/dL (IFCC values; corresponding
values for DCCT/NGSP: 4.3-18.9 % HbA1c). Control Level 1 5.9 0.13 2.2
In rare cases of “>Test” flags which might occur with the use of Control Level 2 10.4 0.11 1.0
the whole blood application, remix the whole blood sample and Human sample 1 5.2 0.08 1.5
repeat the analysis with the same settings. Human sample 2 8.8 0.07 0.8
Lower limits of measurement
Lower detection limit of the test Intermediate Mean SD CV
Hemoglobin: 0.5 g/dL precision** % HbA1c % HbA1c %
HbA1c: 0.1 g/dL Control Level 1 6.0 0.12 2.0
A typical lower detection limit for the HbA1c ratio may be calculated, Control Level 2 10.1 0.17 1.6
based on a given Hb concentration. Assuming a typical Hb concentration Human sample 3 5.6 0.09 1.6
of 13.2 g/dL, the lower detection limit for the HbA1c ratio is 8 mmol/mol Human sample 4 11.7 0.18 1.5
HbA1c (IFCC) or 2.9 % HbA1c (DCCT/NGSP). * repeatability = within-run precision
The lower detection limit represents the lowest measurable analyte ** intermediate precision = total precision / between run precision / between day precision
level that can be distinguished from 0. It is calculated as the value Method comparison
lying 3 standard deviations above that of the lowest standard % HbA1c values (DCCT/NGSP) for human blood samples obtained on
(standard 1 + 3 SD, repeatability, n = 21). a Roche/Hitachi cobas c 501 analyzer (y) were compared with those
Expected values determined using the same reagent on a COBAS INTEGRA 800 analyzer
(x) and on a Roche/Hitachi MODULAR P analyzer (x).
Protocol 1 (mmol/mol HbA1c acc. to IFCC): 29-42 mmol/mol HbA1c28
Protocol 2 (% HbA1c acc. to DCCT/NGSP): 4.8-5.9 % HbA1c28
This reference range was obtained by measuring 474 well-characterized healthy
individuals without diabetes mellitus. HbA1c levels higher than the upper end
of this reference range are an indication of hyperglycemia during the preceding
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A1C-2
Tina-quant Hemoglobin A1c Gen.2
Whole Blood Application: International Federation of Clinical Chemistry and Laboratory Medicine (IFCC)
x = COBAS INTEGRA 800 analyzer, y = cobas c 501 analyzer and the International Diabetes Federation (IDF) HbA1c results should be
Sample size (n) = 109 reported parallel both in mmol/mol HbA1c (IFCC) and % HbA1c (DCCT/NGSP)
Passing/Bablok31 Linear regression values.32 In addition an HbA1c derived mean blood glucose concentration can
y = 0.990x + 0.110 y = 1.003x + 0.023 be reported which can be calculated according to the equations given in the
Summary section of this method sheet. Former % HbA1c values (IFCC) must
τ = 0.958 r = 0.996
not be used due to the risk of mixup/misinterpretation with the % HbA1c values
The sample concentrations were between 5.04 and 12.6 % (DCCT/NGSP (DCCT/NGSP). The use of these % HbA1c values (IFCC) in this application
values). is only an internal operand. The customer is asked to modify the application
according to the consensus statement and/or local requirements.
x = Roche/Hitachi MODULAR P analyzer, y = cobas c 501 analyzer
Sample size (n) = 93
Passing/Bablok31 Linear regression
y = 0.984x + 0.136 y = 0.978x + 0.196
τ = 0.940 r = 0.995
The sample concentrations were between 5.09 and 13.1 % (DCCT/NGSP
values).

In addition, a comparison to a commercially available HPLC method was

performed. The HPLC method was standardized in conformance with
DCCT (Diabetes Control and Complications Trial).3,4
HPLC method
Sample size (n) = 40
Passing/Bablok31 Linear regression
y = 0.935x + 0.450 y = 0.924x + 0.567
τ = 0.950 r = 0.993
The sample concentrations were between 5.25 and 11.9 % (DCCT/NGSP
values).
Hemolysate Application:
x = COBAS INTEGRA 800 analyzer, y = cobas c 501 analyzer
Sample size (n) = 109
Passing/Bablok31 Linear regression
y = 1.032x - 0.145 y = 1.027x - 0.108
τ = 0.962 r = 0.998
The sample concentrations were between 5.15 and 13.1 % (DCCT/NGSP
values).

x = Roche/Hitachi MODULAR P analyzer, y = cobas c 501 analyzer

Sample size (n) = 94
Passing/Bablok31 Linear regression
y = 1.000x + 0.074 y = 1.000x + 0.105
τ = 0.950 r = 0.997
The sample concentrations were between 5.09 and 13.1 % (DCCT/NGSP
values).

In addition, a comparison to a commercially available HPLC method was

performed. The HPLC method was standardized in conformance with
DCCT (Diabetes Control and Complications Trial).3,4
HPLC method
Sample size (n) = 40
Passing/Bablok31 Linear regression
y = 0.949x + 0.407 y = 0.964x + 0.328
τ = 0.954 r = 0.994
The sample concentrations were between 5.25 and 11.9 % (DCCT/NGSP
values).

Analytical specificity
Hb derivatives Labile HbA1c (pre-HbA1c), acetylated Hb, and
carbamylated Hb do not affect the assay results.
Hb variants Specimens containing high amounts of HbF (> 10 %) may
yield lower than expected HbA1c results.
Please note
According to the consensus statement of the American Diabetes Association
(ADA), the European Association for the Study of Diabetes (EASD), the
cobas c systems 6/7 2011-09, V 10 English
04528123190V10

A1C-2
Tina-quant Hemoglobin A1c Gen.2
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A point (period/stop) is always used in this Method Sheet as the decimal
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relevance to diabetes mellitus. Science 1978;200:21-27. stated in the labeling when used in accordance with such labeling and
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method. I. Description of the method. Clin Chim Acta 1984;136:83-93. OTHER WARRANTY, EXPRESS OR IMPLIED, INCLUDING ANY IMPLIED
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tool for the determination of haemoglobin as an alternative to the PURPOSE. IN NO EVENT SHALL ROCHE DIAGNOSTICS BE LIABLE FOR
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on seven glycated hemoglobin methods. Clin Chem 2000;46 (6):864-867. Other brand or product names are trademarks of their respective holders.
Significant additions or changes are indicated by a change bar in the margin.
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Roche Diagnostics GmbH, Sandhofer Strasse 116, D-68305 Mannheim
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17. Jeppsson JO, Kobold U, Finke A, et al. Approved IFCC reference Roche Diagnostics, Indianapolis, IN
US Customer Technical Support 1-800-428-2336
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