0003333752190COINV5.

ALP2
ALP IFCC Gen.2 Enzymes
Order information
Analyzers on which cobas c pack can be used
COBAS INTEGRA 400 plus
03333752 190 ALP IFCC Gen.2 Small (200 tests) System-ID 07 6761 1
COBAS INTEGRA 800
COBAS INTEGRA 400 plus
03333701 190 ALP IFCC Gen.2 Large (400 tests) System-ID 07 6760 3
COBAS INTEGRA 800
10759350 190 Calibrator f.a.s. (12 × 3 mL) System-ID 07 3718 6
10759350 360 Calibrator f.a.s. (12 × 3 mL, for USA) System-ID 07 3718 6
12149435 122 Precinorm U plus (10 × 3 mL) System-ID 07 7999 7
12149435 160 Precinorm U plus (10 × 3 mL, for USA) System-ID 07 7999 7
12149443 122 Precipath U plus (10 × 3 mL) System-ID 07 8000 6
12149443 160 Precipath U plus (10 × 3 mL, for USA) System-ID 07 8000 6
10171743 122 Precinorm U (20 × 5 mL) System-ID 07 7997 0
10171735 122 Precinorm U (4 × 5 mL) System-ID 07 7997 0
10171778 122 Precipath U (20 × 5 mL) System-ID 07 7998 9
10171760 122 Precipath U (4 × 5 mL) System-ID 07 7998 9
05117003 190 PreciControl ClinChem Multi 1 (20 × 5 mL) System-ID 07 7469 3
05947626 190 PreciControl ClinChem Multi 1 (4 × 5 mL) System-ID 07 7469 3
05947626 160 PreciControl ClinChem Multi 1 (4 × 5 mL, for USA) System-ID 07 7469 3
05117216 190 PreciControl ClinChem Multi 2 (20 × 5 mL) System-ID 07 7470 7
05947774 190 PreciControl ClinChem Multi 2 (4 × 5 mL) System-ID 07 7470 7
05947774 160 PreciControl ClinChem Multi 2 (4 × 5 mL, for USA) System-ID 07 7470 7

English The p‑nitrophenol released is directly proportional to the catalytic ALP

activity. It is determined by measuring the increase in absorbance at
System information 409 nm.
cobas c pack ALP2S, Cat. No. 03333752 190:
Test ALP2S, test-ID 0-551 Reagents - working solutions
cobas c pack ALP2L, Cat. No. 03333701 190: R1 2‑amino-2-methyl-1-propanol: 1.724 mol/L, pH 10.44 (30 °C);
Test ALP2L, test-ID 0-550 magnesium acetate: 3.83 mmol/L; zinc sulfate: 0.766 mmol/L;
Intended use N-(2-hydroxyethyl)-ethylenediamine triacetic acid: 3.83 mmol/L
In vitro test for the quantitative determination of the catalytic activity of SR p-nitrophenyl phosphate: 132.8 mmol/L, pH 8.5 (25 °C);
alkaline phosphatase (EC 3.1.3.1; ortho-phosphoric monoester
phosphohydrolase, alkaline optimum) in human serum and plasma on preservatives
COBAS INTEGRA systems. R1 is in position B and SR is in position C.
Summary1,2,3,4,5,6 Precautions and warnings
Alkaline phosphatase in serum consists of four structural genotypes: the Pay attention to all precautions and warnings listed in
liver-bone-kidney type, the intestinal type, the placental type and the variant Section 1 / Introduction of this Method Manual.
from the germ cells. It occurs in osteoblasts, hepatocytes, leukocytes, the
kidneys, spleen, placenta, prostate and the small intestine. The liver-bone- For USA: For prescription use only.
kidney type is particularly important. This kit contains components classified as follows in accordance with the
A rise in the alkaline phosphatase occurs with all forms of cholestasis, Regulation (EC) No. 1272/2008:
particularly with obstructive jaundice. It is also elevated in diseases of the
skeletal system, such as Paget’s disease, hyperparathyroidism, rickets and
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
accelerated bone growth.
Warning
The assay method was first described by King and Armstrong, modified by
Ohmori, Bessey, Lowry and Brock and later improved by Hausamen et al. H315 Causes skin irritation.
In 1983 the International Federation of Clinical Chemistry (IFCC)
recommended a standardized method for the determination of alkaline H319 Causes serious eye irritation.
phosphatase using an optimized substrate concentration and
2‑amino-2-methyl-1-propanol as buffer plus the cations magnesium and H412 Harmful to aquatic life with long lasting effects.
zinc. The assay described here meets the recommendations of the IFCC.
Test principle6 Prevention:
Colorimetric assay in accordance with a standardized method P273 Avoid release to the environment.
In the presence of magnesium and zinc ions, p‑nitrophenyl phosphate is
cleaved by phosphatases into phosphate and p‑nitrophenol. P280 Wear eye protection/ face protection.
ALP P280 Wear protective gloves.
p‑nitrophenyl phosphate + H2O phosphate + p‑nitrophenol Response:

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ALP2
ALP IFCC Gen.2 Enzymes

P362 + P364 Take off contaminated clothing and wash it before reuse. Reaction mode R1-S-SR
Product safety labeling primarily follows EU GHS guidance. Reaction direction Increase
Contact phone: all countries: +49-621-7590, USA: 1-800-428-2336 Wavelength A/B 409/659 nm
Reagent handling Calc. first/last 58/98
Ready for use
Unit U/L
Storage and stability
Pipetting parameters
Shelf life at 2‑8 °C See expiration date on
cobas c pack label Diluent (H2O)
COBAS INTEGRA 400 plus system R1 75 µL 16 µL
On-board in use at 10‑15 °C 4 weeks Sample 2.75 µL 20 µL
COBAS INTEGRA 800 system SR 17 µL 10 µL
On-board in use at 8 °C 8 weeks Total volume 140.75 µL

Specimen collection and preparation Calibration

For specimen collection and preparation only use suitable tubes or Calibrator Calibrator f.a.s.
collection containers.
Only the specimens listed below were tested and found acceptable. Use deionized water as zero
Serum: Collect serum using standard sampling tubes. calibrator.
Plasma: Heparin (Li-, Na-, NH4+-) plasma. Calibration mode Linear regression
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 Calibration replicate Duplicate recommended
available tubes of all manufacturers were tested. Sample collection systems Calibration interval Each lot
from various manufacturers may contain differing materials which could
affect the test results in some cases. When processing samples in primary Traceability: This method has been standardized manually against the
tubes (sample collection systems), follow the instructions of the tube original IFCC formulation.
manufacturer. Quality control
Centrifuge samples containing precipitates before performing the assay.
Reference range Precinorm U, Precinorm U plus or
Stability:7 7 days at 15‑25 °C PreciControl ClinChem Multi 1
7 days at 2‑8 °C Pathological range Precipath U, Precipath U plus or
2 months at (‑15)‑(‑25) °C PreciControl ClinChem Multi 2
Control interval 24 hours recommended
Materials provided
See “Reagents – working solutions” section for reagents. Control sequence User defined
Assay Control after calibration Recommended
For optimum performance of the assay follow the directions given in this For quality control, use control materials as listed in the "Order information"
document for the analyzer concerned. Refer to the appropriate operator’s section.
manual for analyzer‑specific assay instructions. In addition, other suitable control material can be used.
Application for serum and plasma The control intervals and limits should be adapted to each laboratory’s
COBAS INTEGRA 400 plus test definition individual requirements. Values obtained should fall within the defined
limits. Each laboratory should establish corrective measures to be taken if
Measuring mode Absorbance values fall outside the defined limits.
Abs. calculation mode Kinetic Follow the applicable government regulations and local guidelines for
quality control.
Reaction mode R1-S-SR
Calculation
Reaction direction Increase COBAS INTEGRA analyzers automatically calculate the analyte activity of
Wavelength A/B 409/659 nm each sample. For more details, please refer to Data Analysis in the Online
Help (COBAS INTEGRA 400 plus/800 analyzers).
Calc. first/last 41/64
Conversion factor: U/L × 0.0167 = µkat/L
Unit U/L
Limitations - interference
Pipetting parameters Criterion: Recovery within ± 10 % of initial value.
Diluent (H2O) Icterus:8 No significant interference up to an I index of 42 for conjugated
and unconjugated bilirubin (approximate conjugated and unconjugated
R1 75 µL 16 µL bilirubin concentration: 718 µmol/L or 42 mg/dL).
Sample 2.75 µL 20 µL Hemolysis:8 No significant interference up to an H index of 250
(approximate hemoglobin concentration: 155 µmol/L or 250 mg/dL).
SR 17 µL 10 µL
Lipemia (Intralipid):8 No significant interference up to an L index of 2000.
Total volume 140.75 µL There is poor correlation between the L index (corresponds to turbidity) and
triglycerides concentration.
COBAS INTEGRA 800 test definition Drugs: No interference was found at therapeutic concentrations using
Measuring mode Absorbance common drug panels.9,10
In very rare cases, gammopathy, in particular type IgM (Waldenström’s
Abs. calculation mode Kinetic macroglobulinemia), may cause unreliable results.11

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ALP2
ALP IFCC Gen.2 Enzymes

For diagnostic purposes, the results should always be assessed in Repeatability Mean SD CV
conjunction with the patient’s medical history, clinical examination and other U/L (µkat/L) U/L (µkat/L) %
findings.
Precinorm U 80.1 (1.34) 1.3 (0.02) 1.6
ACTION REQUIRED
Special Wash Programming: The use of special wash steps is mandatory Pecipath U 228 (3.81) 4 (0.07) 1.8
when certain test combinations are run together on COBAS INTEGRA
analyzers. Refer to the CLEAN Method Sheet for further instructions and for Human serum 1 72.7 (1.21) 1.5 (0.03) 2.0
the latest version of the Extra wash cycle list. Human serum 2 225 (3.76) 4 (0.07) 1.8
Where required, special wash/carry-over evasion programming must
be implemented prior to reporting results with this test. Intermediate precision Mean SD CV
Limits and ranges U/L (µkat/L) U/L (µkat/L) %
Measuring range Precinorm U 81.8 (1.37) 2.3 (0.04) 2.8
3.0‑1200 U/L (0.05‑20 µkat/L)
Pecipath U 230 (3.84) 6 (0.10) 2.8
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 1 70.0 (1.17) 1.9 (0.03) 2.7
diluted using the rerun function are automatically multiplied by a factor of 5. Human serum 2 220 (3.67) 6 (0.10) 2.7
Lower limits of measurement
Lower detection limit of the test: Method comparison
3.0 U/L (0.05 µkat/L) ALP values for human serum and plasma samples obtained on a
The lower detection limit represents the lowest measurable analyte level COBAS INTEGRA 700 analyzer with the COBAS INTEGRA
that can be distinguished from zero. It is calculated as the value lying ALP IFCC Gen.2 (ALP2L) reagent (y) were compared to those determined
3 standard deviations above that of a zero sample (zero sample + 3 SD, using the same reagent on a Roche/Hitachi 917 analyzer (x), and to those
repeatability, n = 21). determined using the previous reagent (ALP6) on a COBAS INTEGRA 700
analyzer (x).
Expected values
(measured at 37 °C) Roche/Hitachi 917 analyzer Sample size (n) = 97
Passing/Bablok16 Linear regression
Adults12
y = 1.021x - 2.95 U/L y = 1.036x - 5.78 U/L
Males (n = 221) 40-129 U/L (0.67-2.15 µkat/L)
τ = 0.978 r = 0.999
Females (n = 229) 35-104 U/L (0.58-1.74 µkat/L)
SD (md 95) = 10.9 Sy.x = 4.20
Consensus values13 The sample activities were between 37 and 866 U/L (0.618 and
Males 40-130 U/L (0.67-2.17 µkat/L) 14.5 µkat/L).
Females 35-105 U/L (0.58-1.75 µkat/L)
COBAS INTEGRA 700 analyzer Sample size (n) = 93
Children*
Passing/Bablok16 Linear regression
aged 1 day < 250 U/L (< 4.17 µkat/L)
y = 1.006x + 0.034 U/L y = 1.004x + 0.351 U/L
aged 2‑5 days < 231 U/L (< 3.84 µkat/L)
τ = 0.982 r = 1.00
aged 6 days‑6 months < 449 U/L (< 7.49 µkat/L)
SD (md 95) = 7.00 Sy.x = 3.17
aged 7 months‑1 year < 462 U/L (< 7.69 µkat/L)
The sample activities were between 38 and 924 U/L (0.635 and
aged 1‑3 years < 281 U/L (< 4.67 µkat/L) 15.4 µkat/L).
aged 4‑6 years < 269 U/L (< 4.48 µkat/L) References
aged 7‑12 years < 300 U/L (< 5.00 µkat/L) 1 Greiling H, Gressner AM, eds. Lehrbuch der Klinischen Chemie und
Pathobiochemie, 3rd ed. Stuttgart/New York: Schattauer Verlag 1995.
aged 13‑17 years (f) < 187 U/L (< 3.11 µkat/L)
2 King EJ, Armstrong AR. A convenient method for determining serum
aged 13‑17 years (m) < 390 U/L (< 6.51 µkat/L) and bile phosphatase activity. Can Med Assoc J 1934;31(4):376-381.
*calculated from published reference ranges for the ALP opt. method 3 Ohmori Y. Uber die Phosphomonoesterase. Enzymologia
(DGKC)14 using a factor of 0.417 derived from a method comparison. 1937;4:217-231.
Conversion factors to other temperatures have been published,15 but not 4 Bessey OA, Lowry OH, Brock MJ. A method for the rapid determination
checked by Roche for the present reagent. of alkaline phosphatase with five cubic millimeters of serum. J Biol
Chem 1946;164:321-329.
Roche has not evaluated reference ranges in a pediatric population.
5 Hausamen TU, Helger R, Rick W, et al. Optimal conditions for the
Each laboratory should investigate the transferability of the expected values determination of serum alkaline phosphatase by a new kinetic method.
to its own patient population and if necessary determine its own reference Clin Chim Acta 1967;15:241-245.
ranges.
6 Tietz NW, Rinker AD, Shaw LM. International Federation of Clinical
Specific performance data Chemistry. IFCC methods for the measurement of catalytic
Representative performance data on the analyzers are given below. concentration of enzymes, Part 5. IFCC method for alkaline
Results obtained in individual laboratories may differ. phosphatase (orthophosphoric-monoester phosphohydrolase, alkaline
optimum, EC 3.1.3.1). J Clin Chem Clin Biochem 1983;21:731-748.
Precision
Precision was determined using human samples and controls in an internal 7 Guder WG, Narayanan S, Wisser H, et al. List of Analytes;
protocol with repeatability (n = 21) and intermediate precision (1 aliquot per Preanalytical Variables. Brochure in: Samples: From the Patient to the
run, 1 run per day, 21 days). The following results were obtained: Laboratory. Darmstadt: GIT-Verlag 1996.

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ALP IFCC Gen.2 Enzymes

8 Glick MR, Ryder KW, Jackson SA. Graphical Comparisons of

Interferences in Clinical Chemistry Instrumentation.
Clin Chem 1986;32:470-475.
9 Breuer J. Report on the Symposium “Drug effects in Clinical Chemistry
Methods”. Eur J Clin Chem Clin Biochem 1996;34:385-386.
10 Sonntag O, Scholer A. Drug interference in clinical chemistry:
recommendation of drugs and their concentrations to be used in drug
interference studies. Ann Clin Biochem 2001;38:376-385.
11 Bakker AJ, Mücke M. Gammopathy interference in clinical chemistry
assays: mechanisms, detection and prevention.
Clin Chem Lab Med 2007;45(9):1240-1243.
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 Zawta B, Klein G, Bablok W. Temperature Conversion in Clinical
Enzymology? Klin Lab 1994;40:33-42.
16 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
GTIN Global Trade Item Number

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
PURPOSE. IN NO EVENT SHALL ROCHE DIAGNOSTICS BE LIABLE
FOR INCIDENTAL, INDIRECT, SPECIAL OR CONSEQUENTIAL
DAMAGES.
COBAS, COBAS C, COBAS INTEGRA, PRECINORM, PRECIPATH and PRECICONTROL are trademarks of
Roche.
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.
© 2015, Roche Diagnostics

Roche Diagnostics GmbH, Sandhofer Strasse 116, D-68305 Mannheim

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

ALP2 4/4 2015-08, V 5.0 English