0003333752190c501V8.

ALP2
Alkaline Phosphatase acc. to IFCC Gen.2
Order information
Analyzer(s) on which cobas c pack(s) can be used
03333752 190 Alkaline Phosphatase acc. to IFCC Gen.2 ALP2S 200 tests System‑ID 07 6761 1 Roche/Hitachi cobas c 311, cobas c 501/502
03333701 190 Alkaline Phosphatase acc. to IFCC Gen.2 ALP2L 400 tests System‑ID 07 6760 3 Roche/Hitachi cobas c 311, cobas c 501/502
10759350 190 Calibrator f.a.s. (12 x 3 mL) Code 401
10759350 360 Calibrator f.a.s. (12 x 3 mL, for USA) Code 401
12149435 122 Precinorm U plus (10 x 3 mL) Code 300
12149435 160 Precinorm U plus (10 x 3 mL, for USA) Code 300
12149443 122 Precipath U plus (10 x 3 mL) Code 301
12149443 160 Precipath U plus (10 x 3 mL, for USA) Code 301
10171743 122 Precinorm U (20 x 5 mL) Code 300
10171735 122 Precinorm U (4 x 5 mL) Code 300
10171778 122 Precipath U (20 x 5 mL) Code 301
10171760 122 Precipath U (4 x 5 mL) Code 301
05117003 190 PreciControl ClinChem Multi 1 (20 x 5 mL) Code 391
05947626 190 PreciControl ClinChem Multi 1 (4 x 5 mL) Code 391
05947626 160 PreciControl ClinChem Multi 1 (4 x 5 mL, for USA) Code 391
05117216 190 PreciControl ClinChem Multi 2 (20 x 5 mL) Code 392
05947774 190 PreciControl ClinChem Multi 2 (4 x 5 mL) Code 392
05947774 160 PreciControl ClinChem Multi 2 (4 x 5 mL, for USA) Code 392
04489357 190 Diluent NaCl 9 % (50 mL) System‑ID 07 6869 3

English p‑nitrophenyl phosphate + H2O phosphate + p‑nitrophenol

System information
For cobas c 311/501 analyzers: The p‑nitrophenol released is directly proportional to the catalytic ALP
activity. It is determined by measuring the increase in absorbance.
ALP2S: ACN 158
Reagents - working solutions
ALP2L: ACN 683
For cobas c 502 analyzer: R1 2‑amino‑2‑methyl‑1‑propanol: 1.724 mol/L, pH 10.44 (30 °C);
ALP2S: ACN 8158 magnesium acetate: 3.83 mmol/L; zinc sulfate: 0.766 mmol/L;
N‑(2‑hydroxyethyl)‑ethylenediamine triacetic acid: 3.83 mmol/L
ALP2L: ACN 8683
R2 p‑nitrophenyl phosphate: 132.8 mmol/L, pH 8.50 (25 °C);
Intended use preservatives
In vitro test for the quantitative determination of alkaline phosphatase in
human serum and plasma on Roche/Hitachi cobas c systems. R1 is in position B and R2 is in position C.
Summary1,2,3,4,5,6 Precautions and warnings
Alkaline phosphatase in serum consists of four structural genotypes: the For in vitro diagnostic use.
liver‑bone‑kidney type, the intestinal type, the placental type and the variant Exercise the normal precautions required for handling all laboratory
from the germ cells. It occurs in osteoblasts, hepatocytes, leukocytes, the reagents.
kidneys, spleen, placenta, prostate and the small intestine. The Disposal of all waste material should be in accordance with local guidelines.
liver‑bone‑kidney type is particularly important. Safety data sheet available for professional user on request.
A rise in the alkaline phosphatase occurs with all forms of cholestasis, This kit contains components classified as follows in accordance with the
particularly with obstructive jaundice. It is also elevated in diseases of the European directive 1999/45/EC:
skeletal system, such as Paget’s disease, hyperparathyroidism, rickets and R1 contains 2‑Amino‑2‑methyl‑1‑propanol.
osteomalacia, as well as with fractures and malignant tumors. A
considerable rise in the alkaline phosphatase activity is sometimes seen in
children and juveniles. It is caused by increased osteoblast activity following Xi Irritant
accelerated bone growth.
The assay method was first described by King and Armstrong, modified by
Ohmori, Bessey, Lowry and Brock and later improved by Hausamen et al. R 36/38 Irritating to eyes and skin.
In 1983 the International Federation of Clinical Chemistry (IFCC)
recommended a standardized method for the determination of alkaline S 24/25 Avoid contact with skin and eyes.
phosphatase using an optimized substrate concentration and
2‑amino‑2‑methyl‑1‑propanol as buffer plus the cations magnesium and Contact phone: all countries: +49-621-7590, USA: +1-800-428-2336
zinc. The assay described here is based on this recommendation, but was Reagent handling
optimized for performance and stability. The assay was standardized Ready for use
against the IFCC reference formulation proposed above.
Test principle6 Storage and stability
Colorimetric assay in accordance with a standardized method. ALP2S, ALP2L
In the presence of magnesium and zinc ions, p‑nitrophenyl phosphate is
cleaved by phosphatases into phosphate and p‑nitrophenol. Shelf life at 2‑8 °C: See expiration date
on cobas c pack
ALP
label.

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0003333752190c501V8.0

ALP2
Alkaline Phosphatase acc. to IFCC Gen.2

On‑board in use and refrigerated on the analyzer: 8 weeks Reagent pipetting Diluent (H2O)
Diluent NaCl 9 % R1 75 µL 25 µL
Shelf life at 2‑8 °C: See expiration date R2 17 µL 21 µL
on cobas c pack
label. Sample volumes Sample Sample dilution

On‑board in use and refrigerated on the analyzer: 12 weeks Sample Diluent (NaCl)
Normal 2.8 µL – –
Specimen collection and preparation
For specimen collection and preparation only use suitable tubes or Decreased 2.8 µL 20 µL 80 µL
collection containers. Increased 2.8 µL – –
Only the specimens listed below were tested and found acceptable.
Serum. cobas c 502 test definition
Plasma: Li‑heparin plasma.
Assay type Rate A
The sample types listed were tested with a selection of sample collection
tubes that were commercially available at the time of testing, i.e. not all Reaction time / Assay points 10 / 19‑48
available tubes of all manufacturers were tested. Sample collection systems Wavelength (sub/main) 480/450 nm
from various manufacturers may contain differing materials which could
affect the test results in some cases. When processing samples in primary Reaction direction Increase
tubes (sample collection systems), follow the instructions of the tube Units U/L (µkat/L)
manufacturer.
Centrifuge samples containing precipitates before performing the assay. Reagent pipetting Diluent (H2O)
R1 75 µL 25 µL
Stability:7 7 days at 15‑25 °C
R2 17 µL 21 µL
7 days at 2‑8 °C
Sample volumes Sample Sample dilution
2 months at (-15)‑(-25) °C
Sample Diluent (NaCl)
Materials provided
Normal 2.8 µL – –
See “Reagents – working solutions” section for reagents.
Decreased 2.8 µL 20 µL 80 µL
Materials required (but not provided)
▪ See “Order information” section Increased 5.6 µL – –
General laboratory equipment Calibration
Assay Calibrators S1: H2O
For optimum performance of the assay follow the directions given in this S2: C.f.a.s.
document for the analyzer concerned. Refer to the appropriate operator’s
manual for analyzer‑specific assay instructions. Calibration mode Linear
The performance of applications not validated by Roche is not warranted Calibration frequency 2‑point calibration
and must be defined by the user. • after reagent lot change
Applications for serum and plasma • as required following quality control
procedures
cobas c 311 test definition
Traceability: This method has been standardized against the proposed
Assay type Rate A IFCC formulation6 using calibrated pipettes together with a manual
photometer providing absolute values and the substrate‑specific
Reaction time / Assay points 10 / 13‑31 absorptivity, ε.
Wavelength (sub/main) 480/450 nm Quality control
Reaction direction Increase For quality control, use control materials as listed in the "Order information"
Units U/L (µkat/L) section.
In addition, other suitable control material can be used.
Reagent pipetting Diluent (H2O)
The control intervals and limits should be adapted to each laboratory’s
R1 75 µL 25 µL individual requirements. Values obtained should fall within the defined
R2 17 µL 21 µL limits. Each laboratory should establish corrective measures to be taken if
values fall outside the defined limits.
Sample volumes Sample Sample dilution Follow the applicable government regulations and local guidelines for
Sample Diluent (NaCl) quality control.
Normal 2.8 µL – – Calculation
Roche/Hitachi cobas c systems automatically calculate the analyte activity
Decreased 2.8 µL 20 µL 80 µL of each sample.
Increased 2.8 µL – – Conversion factor: U/L x 0.0167 = µkat/L
cobas c 501 test definition Limitations - interference
Criterion: Recovery within ± 10 % of initial value at an alkaline phophatase
Assay type Rate A activity of 100 U/L (1.67 µkat/L).
Reaction time / Assay points 10 / 19‑48 Icterus:8 No significant interference up to an I index of 60 for conjugated
Wavelength (sub/main) 480/450 nm and unconjugated bilirubin (approximate conjugated and unconjugated
bilirubin concentration: 1026 µmol/L or 60 mg/dL).
Reaction direction Increase Hemolysis:8 No significant interference up to an H index of 200
Units U/L (µkat/L) (approximate hemoglobin concentration: 124 µmol/L or 200 mg/dL).

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ALP2
Alkaline Phosphatase acc. to IFCC Gen.2

Lipemia (Intralipid):8 No significant interference up to an L index of 2000. Specific performance data

There is poor correlation between the L index (corresponds to turbidity) and Representative performance data on the analyzers are given below.
triglycerides concentration. Results obtained in individual laboratories may differ.
Drugs: No interference was found at therapeutic concentrations using Precision
common drug panels.9,10
Precision was determined using human samples and controls in an internal
In very rare cases, gammopathy, in particular type IgM (Waldenström’s protocol with repeatability (n = 21) and intermediate precision (3 aliquots
macroglobulinemia), may cause unreliable results.11 per run, 1 run per day, 21 days). The following results were obtained:
For diagnostic purposes, the results should always be assessed in
conjunction with the patient’s medical history, clinical examination and other Repeatability Mean SD CV
findings.
U/L (µkat/L) U/L (µkat/L) %
ACTION REQUIRED
Special Wash Programming: The use of special wash steps is mandatory Precinorm U 99.2 (1.65) 0.7 (0.01) 0.7
when certain test combinations are run together on Roche/Hitachi
cobas c systems. The latest version of the carry‑over evasion list can be Precipath U 241 (4.02) 1 (0.02) 0.6
found with the NaOHD/SMS/Multiclean/SCCS or the Human serum 1 54.6 (0.912) 0.5 (0.008) 0.9
NaOHD/SMS/SmpCln1+2/SCCS Method Sheets. For further instructions
refer to the operator’s manual. cobas c 502 analyzer: All special wash Human serum 2 648 (10.8) 4 (0.1) 0.7
programming necessary for avoiding carry‑over is available via the
cobas link, manual input is not required. Intermediate precision Mean SD CV
Where required, special wash/carry‑over evasion programming must U/L (µkat/L) U/L (µkat/L) %
be implemented prior to reporting results with this test.
Limits and ranges Precinorm U 92.8 (1.56) 2.2 (0.04) 2.4
Measuring range Precipath U 224 (3.74) 4 (0.06) 1.7
5‑1200 U/L (0.084‑20.0 µkat/L) Human serum 3 82.2 (1.37) 1.8 (0.03) 2.1
Determine samples having higher activities via the rerun function. Dilution
of samples via the rerun function is a 1:5 dilution. Results from samples Human serum 4 1025 (17.1) 9 (0.2) 0.9
diluted using the rerun function are automatically multiplied by a factor of 5. Method comparison
Lower limits of measurement Alkaline phosphatase values for human serum and plasma samples
Lower detection limit of the test obtained on a Roche/Hitachi cobas c 501 analyzer (y) were compared with
5 U/L (0.084 µkat/L) those determined using the corresponding reagent on a
Roche/Hitachi 917 analyzer (x).
The lower detection limit represents the lowest measurable analyte level
that can be distinguished from zero. It is calculated as the value lying 3 Sample size (n) = 203
standard deviations above that of the lowest standard (standard 1 + 3 SD, Passing/Bablok15 Linear regression
repeatability, n = 21).
y = 0.988x + 1.31 U/L y = 0.991x + 0.799 U/L
Expected values
(measured at 37 °C) τ = 0.961 r = 0.997
The sample activities were between 50.0 and 1002 U/L (0.835 and
Adults12 16.7 µkat/L).
Males (n = 221) 40‑129 U/L (0.67‑2.15 µkat/L) References
Females (n = 229) 35‑104 U/L (0.58‑1.74 µkat/L) 1 Greiling H, Gressner AM, eds. Lehrbuch der Klinischen Chemie und
Consensus values13 Pathobiochemie, 3rd ed. Stuttgart/New York: Schattauer Verlag 1995.
2 King EJ, Armstrong AR. Can Med Assoc J 1934;31:376
Males 40‑130 U/L (0.67‑2.17 µkat/L)
3 Ohmori Y. Uber die Phosphomomesterase. Enzymologia
Females 35‑105 U/L (0.58‑1.75 µkat/L) 1937;4:217-231.
Children a) 4 Bessey OA, Lowry OH, Brock MJ. A method for the rapid determination
aged 1 day < 250 U/L (< 4.17 µkat/L) of alkaline phosphatase with five cubic millimeters of serum. J Biol
Chem 1946;164:321-329.
aged 2‑5  days < 231 U/L (< 3.84 µkat/L)
5 Hausamen TU, Helger R, Rick W, et al. Optimal conditions for the
aged 6 days‑6 months < 449 U/L (< 7.49 µkat/L) determination of serum alkaline phosphatase by a new kinetic method.
Clin Chim Acta 1967;15:241-245.
aged 7 months‑1 year < 462 U/L (< 7.69 µkat/L)
6 Tietz NW, Rinker AD, Shaw LM. J Clin Chem Clin Biochem
aged 1‑3 years < 281 U/L (< 4.67 µkat/L) 1983;21:731-748.
aged 4‑6 years < 269 U/L (< 4.48 µkat/L) 7 Use of Anticoagulants in Diagnostic Laboratory Investigations. WHO
aged 7‑12 years < 300 U/L (< 5.00 µkat/L) Publication WHO/DIL/LAB/99.1 Rev.2.
aged 13‑17 years (f) < 187 U/L (< 3.11 µkat/L) 8 Glick MR, Ryder KW, Jackson SA. Graphical Comparisons of
Interferences in Clinical Chemistry Instrumentation. Clin Chem
aged 13‑17 years (m) < 390 U/L (< 6.51 µkat/L) 1986;32:470-475.
a) Calculated from published reference ranges for the ALP opt. method 9 Breuer J. Report on the Symposium “Drug effects in Clinical Chemistry
(DGKC)14 using a factor of 0.417 derived from a method comparison. Methods”. Eur J Clin Chem Clin Biochem 1996;34:385-386.
Roche has not evaluated reference ranges in a pediatric population. 10 Sonntag O, Scholer A. Drug interference in clinical chemistry:
Each laboratory should investigate the transferability of the expected values recommendation of drugs and their concentrations to be used in drug
to its own patient population and if necessary determine its own reference interference studies. Ann Clin Biochem 2001;38:376-385.
ranges. 11 Bakker AJ, Mücke M. Gammopathy interference in clinical chemistry
assays: mechanisms, detection and prevention.
Clin Chem Lab Med 2007;45(9):1240-1243.

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ALP2
Alkaline Phosphatase acc. to IFCC Gen.2

12 Abicht K, El-Samalouti V, Junge W, et al. Multicenter evaluation of new

GGT and ALP reagents with new reference standardization and
determination of 37 °C reference intervals. Clin Chem Lab Med
2001;39:Special Supplement pp S 346.
13 Thomas L, Müller M, Schumann G, et al. Consensus of DGKL and
VDGH for interim reference intervals on enzymes in serum. J Lab Med
2005;29:301-308.
14 Fischbach F, Zawta B. Age-dependent Reference Limits of Several
Enzymes in Plasma at Different Measuring Temperatures. Klin Lab
1992;38:556-561.
15 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.
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.
Symbols
Roche Diagnostics uses the following symbols and signs in addition to
those listed in the ISO 15223‑1 standard.
Contents of kit
Volume after reconstitution or mixing

FOR US CUSTOMERS ONLY: LIMITED WARRANTY

Roche Diagnostics warrants that this product will meet the specifications
stated in the labeling when used in accordance with such labeling and will
be free from defects in material and workmanship until the expiration date
printed on the label. THIS LIMITED WARRANTY IS IN LIEU OF ANY
OTHER WARRANTY, EXPRESS OR IMPLIED, INCLUDING ANY IMPLIED
WARRANTY OF MERCHANTABILITY OR FITNESS FOR PARTICULAR
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FOR INCIDENTAL, INDIRECT, SPECIAL OR CONSEQUENTIAL
DAMAGES.
COBAS, COBAS C,PRECINORM, PRECIPATH and PRECICONTROL are trademarks of Roche.
All other product names and trademarks are the property of their respective owners.
Significant additions or changes are indicated by a change bar in the margin.
© 2013, Roche Diagnostics

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www.roche.com
Distribution in USA by:
Roche Diagnostics, Indianapolis, IN
US Customer Technical Support 1-800-428-2336

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