ALKALINE PHOSPHATASE

7D55-20 and 7D55-30

30-3947/R7

ALKALINE PHOSPHATASE
This package insert contains information to run the Alkaline Phosphatase assay on the ARCHITECT c Systems™
and the AEROSET System.

NOTE: Changes Highlighted

NOTE: This package insert must be read carefully prior to product use. Package insert instructions must be followed
accordingly. Reliability of assay results cannot be guaranteed if there are any deviations from the instructions in
this package insert.

Customer Support
United States: 1-877-4ABBOTT
Canada: 1-800-387-8378 (English speaking customers)
1-800-465-2675 (French speaking customers)
International: Call your local Abbott representative

Symbols in Product Labeling

Concentration Catalog number/List number

Authorized Representative in the
Serial number
European Community
Ingredients Consult instructions for use

In vitro diagnostic medical device Manufacturer

Batch code/Lot number Temperature limitation

Reagent 1 Use by/Expiration date

Reagent 2

ABBOTT LABORATORIES ABBOTT

Abbott Park, IL 60064, USA Max-Planck-Ring 2
65205 Wiesbaden
Germany
+49-6122-580
February 2007
©2002, 2007 Abbott Laboratories

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NAME
ALKALINE PHOSPHATASE
INTENDED USE
The Alkaline Phosphatase assay is used for the quantitation of alkaline
phosphatase in human serum or plasma.
SUMMARY AND EXPLANATION OF TEST
Human alkaline phosphatase (AlkP, EC.3.1.3.1) consists of a group of at
least five tissue-specific isoenzymes which catalyzes the hydrolysis of
phosphate mono-esters at alkaline pH. A variety of disease processes
can result in the release of increased quantities of alkaline phosphatase
into the blood.
PRINCIPLES OF PROCEDURE
Several substrates have been used to measure alkaline phosphatase
activity such as glycerophosphate,1 phenyl phosphate,1 and
p-nitrophenyl phosphate.2 Bowers and McComb3 improved the method
of Bessey et al. to include a kinetic measurement. Tietz et al.4
optimized this method to include a chelated metal-ion buffer of zinc,
magnesium, and HEDTA. This Alkaline Phosphatase procedure is a
modification of this method.
Alkaline phosphatase in the sample catalyzes the hydrolysis of colorless
p-nitrophenyl phosphate (p-NPP) to give p-nitrophenol and inorganic
phosphate. At the pH of the assay (alkaline), the p-nitrophenol is in the
yellow phenoxide form. The rate of absorbance increase at 404 nm is
directly proportional to the alkaline phosphatase activity in the sample.
Optimized concentrations of zinc and magnesium ions are present to
activate the alkaline phosphatase in the sample.
Methodology: Para-nitrophenyl Phosphate
WARNINGS AND PRECAUTIONS
Precautions for Users
1. For in vitro diagnostic use.
2. Do not use components beyond the expiration date.
3. Do not mix materials from different kit lot numbers.
4. is classified per applicable European Community (EC) Directives
as: Irritant (Xi). The following are the appropriate Risk (R) and Safety
(S) phrases:
R36/38 Irritating to eyes and skin.
R52/53 Harmful to aquatic organisms, may cause long-
term adverse effects in the aquatic environment.
S26 In case of contact with eyes, rinse immediately
with plenty of water and seek medical advice.
S35 This material and its container must be disposed
of in a safe way.
S37 Wear suitable gloves.
S46 If swallowed, seek medical advice immediately
and show this container or label.
S61 Avoid release to the environment. Refer to special
instructions/safety data sheet.
For product not classified as dangerous per European Directive
1999/45/EC as amended, safety data sheet available for
professional user on request.
5. CAUTION: This product requires the handling of human specimens.
It is recommended that all human sourced materials be considered
potentially infectious and handled in accordance with the OSHA
Standard on Bloodborne Pathogens.5 Biosafety Level 26 or other
appropriate biosafety practices7,8 should be used for materials that
contain or are suspected of containing infectious agents.

REAGENTS SPECIMEN COLLECTION AND HANDLING

Reagent Kit Suitable Specimens
Alkaline Phosphatase is supplied as a liquid, ready-to-use, two-reagent Serum and plasma are acceptable specimens.
kit which contains: • Serum: Use serum collected by standard venipuncture techniques
into glass or plastic tubes with or without gel barriers. Ensure
7D55-20 complete clot formation has taken place prior to centrifugation.
10 x 84 mL Separate serum from red blood cells or gel as soon after collection
10 x 35 mL as possible.
Some specimens, especially those from patients receiving
Estimated tests per kit: 11,358* anticoagulant or thrombolytic therapy, may take longer to complete
7D55-30 their clotting processes. Fibrin clots may subsequently form in these
sera and the clots could cause erroneous test results.
5 x 48 mL • Plasma: Use plasma collected by standard venipuncture techniques
5 x 22 mL into glass or plastic tubes. Acceptable anticoagulants are lithium
heparin (with or without gel barrier) and sodium heparin. Ensure
Estimated tests per kit: 3,165* centrifugation is adequate to remove platelets. Separate plasma from
*Calculation is based on the minimum reagent fill volume per kit. red blood cells or gel as soon after collection as possible.
Reactive Ingredients Concentration Refer to the specimen collection tube manufacturer’s instructions for
2-Amino-2-methylpropanol > 1.2 mol/L processing and handling requirements.
Magnesium > 7.2 mmol/L For total sample volume requirements, refer to the instrument-specific
ASSAY PARAMETERS section of this package insert and Section 5 of
Zinc Sulfate > 3.6 mmol/L
the instrument-specific operations manual.
HEDTA > 7.2 mmol/L
Specimen Storage
4-Nitrophenyl Phosphate > 171.6 mmol/L Serum and plasma: Allow specimens to reach room temperature prior
to analysis.9
REAGENT HANDLING AND STORAGE Temperature Maximum Bibliographic
Reagent Handling Storage Reference
Remove air bubbles, if present in the reagent cartridge, with a new 20 to 25°C 7 days 10
applicator stick. Alternatively, allow the reagent to sit at the appropriate
storage temperature to allow the bubbles to dissipate. To minimize 2 to 8°C 7 days 10, 11
volume depletion, do not use a transfer pipette to remove the bubbles. -20°C 2 months 10
CAUTION: Reagent bubbles may interfere with proper detection of
reagent level in the cartridge, causing insufficient reagent aspiration Guder et al.10
suggest storage of frozen specimens at -20°C for
which could impact results. no longer than the time interval cited above. However, limitations
of laboratory equipment make it necessary in practice for clinical
Reagent Storage laboratories to establish a range around -20°C for specimen storage.
Unopened reagents are stable until the expiration date when stored This temperature range may be established from either the freezer
at 2 to 8°C. The reagent is light sensitive. Always store reagents manufacturer’s specifications or your laboratory standard operating
protected from light. procedure(s) for specimen storage.
Reagent stability is 10 days if the reagent is uncapped and onboard. NOTE: Stored specimens must be inspected for particulates. If present,
mix and centrifuge the specimen to remove particulates prior to testing.

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PROCEDURE RESULTS
Materials Provided Refer to the instrument-specific operations manual for information on
results calculations.
7D55-20 or 7D55-30 Alkaline Phosphatase Reagent Kit
• ARCHITECT System Operations Manual—Appendix C
Materials Required but not Provided • AEROSET System Operations Manual—Appendix A
• Control Material Representative performance data are given in the EXPECTED VALUES
• Saline (0.85% to 0.90% NaCl) for specimens that require dilution and SPECIFIC PERFORMANCE CHARACTERISTICS sections of this
Assay Procedure package insert. Results obtained in individual laboratories may vary.
For a detailed description of how to run an assay, refer to Section 5 of LIMITATIONS OF THE PROCEDURE
the instrument-specific operations manual.
Refer to the SPECIMEN COLLECTION AND HANDLING and SPECIFIC
Specimen Dilution Procedures PERFORMANCE CHARACTERISTICS sections of this package insert.
The ARCHITECT c Systems and the AEROSET System have automatic
dilution features; refer to Section 2 of the instrument-specific operations EXPECTED VALUES
manual for additional information.
Reference Range
Serum and plasma: Specimens with alkaline phosphatase values
exceeding 2,200 U/L (4,555 U/L for Flex Rate Linearity) are flagged Serum/Plasma12
and may be diluted using the Automated Dilution Protocol or the Manual
Dilution Procedure. Range (U/L)
Male
Automated Dilution Protocol
1 to 12 years < 500
If using the Automated Dilution Protocol, the system performs a dilution
of the specimen and automatically corrects the enzyme activity value 12 to 15 years < 750
by multiplying the result by the appropriate dilution factor. To set up the > 20 years 40 to 150
automatic dilution feature, refer to Section 2 of the instrument-specific Female
operations manual for additional information. 1 to 12 years < 500
Manual Dilution Procedure > 15 years 40 to 150
Manual dilutions should be performed as follows: It is recommended that each laboratory determine its own reference
• Use saline (0.85% to 0.90% NaCl) to dilute the sample. range based upon its particular locale and population characteristics.
• The operator must enter the dilution factor in the patient or control
order screen. The system uses this dilution factor to automatically SPECIFIC PERFORMANCE CHARACTERISTICS
correct the enzyme activity value by multiplying the result by the
entered factor. Linearity
Alkaline Phosphatase is linear up to 2,200 U/L.
• If the operator does not enter the dilution factor, the result must
be multiplied by the appropriate dilution factor before reporting the Flex Rate Linearity is 4,555 U/L. To use Flex Rate Linearity, the operator
result. must edit the linear high value to 4,555 on the appropriate screen.
NOTE: If a diluted sample result is flagged indicating it is less than the • ARCHITECT c Systems—Configure assay parameters screen,
linear low limit, do not report the result. Rerun using an appropriate Results view
dilution. • AEROSET System—Assay Configuration screen, Outline page
For detailed information on ordering dilutions, refer to Section 5 of the Linearity was verified using Clinical and Laboratory Standards Institute
instrument-specific operations manual. (CLSI) protocol NCCLS EP6-P.13
Limit of Detection (LOD)
CALIBRATION The LOD for Alkaline Phosphatase is 5.0 U/L. The LOD is the
Calibration is stable for approximately 10 days (240 hours) and is mean concentration of an analyte-free sample + 2 SD, where
required with each change in reagent lot number. Verify calibration SD = the pooled, within-run standard deviation of the analyte-free
with at least two levels of controls according to the established quality sample. A study performed on an ARCHITECT c System and an
control requirements for your laboratory. If control results fall outside AEROSET System produced an LOD for Alkaline Phosphatase
acceptable range, recalibration may be necessary. of 1.9 U/L.
A calibration factor (2150) must be entered.
Limit of Quantitation (LOQ)
• ARCHITECT c Systems—Configure assay parameters window,
Calibration view The LOQ for Alkaline Phosphatase is 5.0 U/L. The LOQ is the analyte
concentration at which the CV = 20%.
• AEROSET System—Assay Configuration screen, Calibration page
For a detailed description of how to calibrate an assay, refer to Interfering Substances14
Section 6 of the instrument-specific operations manual. Interference studies were conducted on the AEROSET System using
CLSI protocol NCCLS EP7-P.15 Interference effects were assessed by
QUALITY CONTROL Dose Response and Paired Difference methods, at the medical decision
level of the analyte.
The following is the recommendation of Abbott Laboratories for quality
control. As appropriate, refer to your laboratory standard operating Interfering Interferent Concentration N Target Observed
procedure(s) and/or quality assurance plan for additional quality control Substance (U/L) (% of Target)
requirements and potential corrective actions.
30 mg/dL (513 μmol/L) 4 155.6 102.4
• Two levels of controls (normal and abnormal) are to be run every Bilirubin
24 hours. 60 mg/dL (1,026 μmol/L) 4 155.6 102.7
• If more frequent control monitoring is required, follow the established 750 mg/dL (7.5 g/L) 4 150.8 95.3
quality control procedures for your laboratory. Hemoglobin
1,000 mg/dL (10.0 g/L) 4 150.8 94.3
• If quality control results do not meet the acceptance criteria 750 mg/dL (7.5 g/L) 4 144.3 100.4
defined by your laboratory, patient values may be suspect. Follow Intralipid
the established quality control procedures for your laboratory. 1,000 mg/dL (10.0 g/L) 4 144.3 100.2
Recalibration may be necessary. Bilirubin solutions at the above concentrations were prepared by addition
• Review quality control results and acceptance criteria following a of a bilirubin stock to human serum pools. Hemoglobin solutions at
change of reagent lot. the above concentrations were prepared by addition of hemolysate to
human serum pools. Intralipid solutions at the above concentrations were
prepared by addition of Intralipid to human serum pools.

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SPECIFIC PERFORMANCE CHARACTERISTICS BIBLIOGRAPHY
(Continued) 1. King EJ, Armstrong AR. A convenient method for determining serum
and bile phosphatase activity. Can Med Assoc J 1934;31:376–81.
Precision
2. Bessey OA, Lowry OH, Brock MJ. A method for the rapid
The imprecision of the Alkaline Phosphatase assay is ≤ 6.2% Total CV.
determination of alkaline phosphatase with five cubic millimeters of
Representative data from studies using CLSI protocol NCCLS EP5-A16
serum. J Biol Chem 1946;164:321–9.
are summarized below.
3. Bowers GN, McComb RB. A continuous spectrophotometric method
Control Level 1 Level 2 for measuring the activity of serum alkaline phosphatase. Clin Chem
1966;12(2):70–89.
N 80 80 4. Tietz NW, Burtis CA, Duncan P, et al. A reference method for
Mean (U/L) 84.4 437.8 measurement of alkaline phosphatase activity in human serum.
Clin Chem 1983;29(5):751–6.
SD 3.89 3.17
Within Run 5. US Department of Labor, Occupational Safety and Health
%CV 4.6 0.7 Administration. 29 CFR Part 1910.1030. Occupational Exposure to
Bloodborne Pathogens.
SD 2.86 5.53
Between Run 6. US Department of Health and Human Services. Biosafety in
%CV 3.4 1.3 Microbiological and Biomedical Laboratories. HHS Publication
SD 0.00 7.29 (CDC), 4th ed. Washington, DC: US Government Printing Office,
Between Day May 1999.
%CV 0.0 1.7 7. World Health Organization. Laboratory Biosafety Manual. Geneva:
SD 4.83 9.68 World Health Organization; 2004.
Total 8. Sewell DL, Bove KE, Callihan DR, et al. Protection of Laboratory
%CV 5.7 2.2 Workers from Occupationally Acquired Infections; Approved
Guideline—Third Edition (M29-A3). Wayne, PA: Clinical and
Method Comparison Laboratory Standards Institute, 2005.
Correlation studies were performed using CLSI protocol NCCLS 9. Tietz NW, Rinker AD, Shaw LM. IFCC methods for the measurement
EP9-A.17 of catalytic concentration of enzymes, part 5: IFCC method for
Serum results from the Alkaline Phosphatase assay on the AEROSET alkaline phosphatase. J Clin Chem Clin Biochem 1983;21(11):
System were compared with those from a commercially available 731–48.
p-nitrophenyl phosphate hydrolysis methodology. 10. Guder WG, da Fonseca-Wollheim F, Heil W, et al. The Quality of
Serum results from the Alkaline Phosphatase assay on an ARCHITECT Diagnostic Samples. Darmstadt, Germany: GIT Verlag; 2001:14–5.
c System were compared with those from the Alkaline Phosphatase 11. US Pharmacopeial Convention, Inc. General notices. In: US
assay on the AEROSET System. Pharmacopeia National Formulary, 1995 ed. (USP 23/NF18).
AEROSET ARCHITECT vs. Rockville, MD: The US Pharmacopeial Convention, Inc; 1994:11.
vs. Comparative AEROSET 12. Burtis CA, Ashwood ER, editors. Tietz Textbook of Clinical
Method Chemistry, 2nd ed. Philadelphia, PA: WB Saunders; 1994:836.
13. Passey RB, Bee DE, Caffo A, et al. Evaluation of the Linearity
N 120 109 of Quantitative Analytical Methods; Proposed Guideline (EP6-P).
Y - Intercept -0.124 1.209 Villanova, PA: The National Committee for Clinical Laboratory
Standards, 1986.
Correlation Coefficient 0.999 1.000
14. Young DS. Effects of Drugs on Clinical Laboratory Tests, 4th ed.
Slope 1.059 0.981 Washington, DC: AACC Press; 1995:3-26-3-35.
Range (U/L)* 29.6 to 957.1 31.6 to 4,527.1 15. Powers DM, Boyd JC, Glick MR, et al. Interference Testing in
Clinical Chemistry; Proposed Guideline (EP7-P). Villanova, PA:
*AEROSET Range
The National Committee for Clinical Laboratory Standards, 1986.
16. Kennedy JW, Carey RN, Coolen RB, et al. Evaluation of Precision
Performance of Clinical Chemistry Devices; Approved Guideline
(EP5-A). Wayne, PA: The National Committee for Clinical
Laboratory Standards, 1999.
17. Kennedy JW, Carey NR, Coolen RB, et al. Method Comparison and
Bias Estimation Using Patient Samples; Approved Guideline (EP9-A).
Wayne, PA: The National Committee for Clinical Laboratory
Standards, 1995.

TRADEMARKS
AEROSET and ARCHITECT are registered trademarks of Abbott
Laboratories.
c System is a trademark of Abbott Laboratories.
All other trademarks, brands, product names, and trade names are the
property of their respective companies.

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ARCHITECT c SYSTEMS ASSAY PARAMETERS

Alkaline Phosphatase Serum/Plasma—Conventional and SI Units

Configure assay parameters — General Configure assay parameters — SmartWash
● General о Calibration о SmartWash о Results о Interpretation о General о Calibration ● SmartWash о Results о Interpretation
Assay: AlkP Type: Photometric Version: 1 Assay: AlkP
Number: 1020
COMPONENT REAGENT / ASSAY WASH Volume Replicates
● Reaction definition о Reagent / Sample о Validity checks Cuvette Trig 10% Detergent B*** 345
Reaction mode: Rate up
Primary Secondary Read times *** Select “Detergent B” for software prior to Version 2.2.
Wavelength: 404 / 476 Main: 20 – 23
Last required read: 23 Flex: 17 – 20
Absorbance range: 0.0000 – 2.8000 Color correction: ___ – ___ Configure assay parameters — Results
Sample blank type: None о General о Calibration о SmartWash ● Results о Interpretation
Assay: AlkP Result units: U/L
о Reaction definition ● Reagent / Sample о Validity checks Assay defaults:
R1 R2 Low-Linearity: 5†
Reagent: ALKP0 Reagent volume: 60 20 High-Linearity: 2200
Diluent: Saline Water volume: 170 ___ Gender and age specific ranges:
Diluent dispense mode: Type 0 Dispense mode: Type 0 Type 0 GENDER AGE (UNITS) NORMAL EXTREME
Either 0 – 130 (Y) 40 – 150
Diluted Default
Dilution name Sample sample Diluent Water Dilution factor dilution
STANDARD : 7.0 ___ ___ ___ = 1:1.00 ● Configure result units
_________ : ___ ___ ___ ___ = о Assay: AlkP
_________ : ___ ___ ___ ___ = о Version: 1
Result units: U/L
о Reaction definition о Reagent / Sample ● Validity checks Decimal places: 0 [Range 0 – 4]
Reaction check: None Correlation factor: 1.0000
Intercept: 0.0000

Rate linearity %: 10

Configure assay parameters — Calibration

о General ● Calibration о SmartWash о Results о Interpretation
Assay: AlkP Calibration method: Factor
Factor: 2150.0000
● Calibrators о Volumes о Intervals о Validity checks
Calibrator set: Calibrator level: Concentration:
None Blank: Water 0

Replicates: 3 [Range 1 – 3]

о Calibrators ● Volumes о Intervals о Validity checks

Calibrator: Diluted
Calibrator level Sample sample Diluent Water
Blank: Water 7.0 ___ ___ ___

о Calibrators о Volumes ● Intervals о Validity checks

Calibration intervals:
Full interval: 240 (hours)

о Calibrators о Volumes о Intervals ● Validity checks

Blank absorbance range: _____ – _____

† The linear low value (Low-Linearity) is LOQ rounded up to the number of decimal places defined in the decimal places parameter field.

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AEROSET SYSTEM ASSAY PARAMETERS

Alkaline Phosphatase Serum/Plasma—Conventional and SI Units

Assay Configuration: Outline Page

Assay Name Assay # Line
AlkP 20 B-Line
Quantitative Ranges
Min Text Min Panic-L L-Reference-H Panic-H Max Max Text
* 0.0* 0.0 40 150 0.0 0.0* *
5** L-Linear Range-H 2200
Reference Ranges*
Age Male Female
0 Year 0.0 – 0.0 0.0 – 0.0
0.0 – 0.0 0.0 – 0.0
0 Year
0.0 – 0.0 0.0 – 0.0
0 Year 0.0 – 0.0 0.0 – 0.0
Qualitative Ranges N/A

Assay Configuration: Base Page

Reaction Mode Wavelength-Prim/Sec Read time-Main/Flex Linearity %
RATE UP 404 / 476 20 – 23 / 17 – 20 10
Sample Blank Test Blank Read Time Abs Window Abs Limits
_____ ( ___ ) 0–0 0–0 0.0 – 2.8
S.Vol DS.Vol D.Vol W.Vol
Standard 7.0 0.0 0 0 Rgt Name/Pos
Dil 1 7.0 0.0 0 0 Diluent: _____ _–__*
Dil 2 7.0 0.0 0 0 Type# 0
Rgt Name/Pos R.Vol W.Vol Type#
Reagent 1 ALKP061‡‡ – ___* 60 170 0
Reagent 2 ALKP052‡‡ – ___* 20 0 0
Reaction Check Read Time – A/B Range Minimum
___________ 1–1/1–1 0.0 – 0.0 0.0
Factor/Intercept Decimal Places Units
1.0 / 0.0 0 U/L

Assay Configuration: Calibration Page

Calib Mode Factor Interval (H)
Factor 2150 240
Blank/Calib Replicates Extrapolation % Span Span Abs Range
3/3 0 BLK – 1 0.0 – 0.0
Sample S.Vol DS.Vol D.Vol W.Vol Blk Abs Range
BLK Water 7.0 0.0 0 0 0.0 – 0.0
C1 2.0 0.0 0 0 Cal Deviation
C2 2.0 0.0 0 0 0.0

Assay Configuration: SmartWash Page

Rgt Probe
Reagent Wash Vol
— — —
Cuvette
Assay Name Wash Vol
— — —
Sample Probe
Wash
—

Refer to Assay Configuration in Section 2 of the AEROSET System Operations Manual for information regarding assay parameters.
* User defined or instrument defined.
** The linear low value (L-Linear Range) is LOQ rounded up to the number of decimal places defined in the decimal places parameter field.
‡‡ Rgt Name listed is for 7D55-20. For 7D55-30, change Reagent 1 name to ALKP051; Reagent 2 name remains ALKP052

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