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    Acid Phosphatase: Orthophosphoric - Monoester Phospho-Hydrolase

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    Maher
    This document provides a pipetting scheme and instructions for performing an assay to measure acid phosphatase levels using a reagent kit. The assay involves hydrolyzing 1-naphthyl phosphate with acid phosphatase to form phosphate and 1-naphthol, which is then converted to an azo dye for measurement. The increase in absorbance at 405nm over time is proportional to the acid phosphatase activity in the sample. The document provides details on sample preparation, reagent preparation, controls, reference values and automation options.

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    © All Rights Reserved
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    Acid Phosphatase: Orthophosphoric - Monoester Phospho-Hydrolase

    Uploaded by

    Maher
    78 views1 page
    This document provides a pipetting scheme and instructions for performing an assay to measure acid phosphatase levels using a reagent kit. The assay involves hydrolyzing 1-naphthyl phosphate with acid phosphatase to form phosphate and 1-naphthol, which is then converted to an azo dye for measurement. The increase in absorbance at 405nm over time is proportional to the acid phosphatase activity in the sample. The document provides details on sample preparation, reagent preparation, controls, reference values and automation options.

    Original Description:

    Leaflet for acid phos for human diagnostic

    Original Title

    ACID PHOS

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    This document provides a pipetting scheme and instructions for performing an assay to measure acid phosphatase levels using a reagent kit. The assay involves hydrolyzing 1-naphthyl phosphate with acid phosphatase to form phosphate and 1-naphthol, which is then converted to an azo dye for measurement. The increase in absorbance at 405nm over time is proportional to the acid phosphatase activity in the sample. The document provides details on sample preparation, reagent preparation, controls, reference values and automation options.

    Copyright:

    © All Rights Reserved
    Download as PDF, TXT or read online from Scribd
    Download as pdf or txt
    78 views1 page

    Acid Phosphatase: Orthophosphoric - Monoester Phospho-Hydrolase

    Uploaded by

    Maher
    This document provides a pipetting scheme and instructions for performing an assay to measure acid phosphatase levels using a reagent kit. The assay involves hydrolyzing 1-naphthyl phosphate with acid phosphatase to form phosphate and 1-naphthol, which is then converted to an azo dye for measurement. The increase in absorbance at 405nm over time is proportional to the acid phosphatase activity in the sample. The document provides details on sample preparation, reagent preparation, controls, reference values and automation options.

    Copyright:

    © All Rights Reserved
    Download as PDF, TXT or read online from Scribd
    Download as pdf or txt
    of 1

    Acid Phosphatase Pipetting Scheme

    Warm working reagents and cuvettes up to the desired temperature.


    Orthophosphoric - Monoester Phospho- Temperature must be kept constant (±0.5°C) for the duration of the test.
    hydrolase Assay temperature 25°C / 30°C / 37°C
    Assay Semi micro Macro
    Package Size
    [REF] 10660 6 x 15 ml Complete test kit Sample 100 µl 200 µl
    [IVD]
    Working reagent 1000 µl 2000 µl
    Mix, read the absorbance A1 after 5 minutes and start the stop watch
    Method1 at the same time. Read the absorbance A2 exactly after 3 minutes at
    1-Naphthyl phosphate is hydrolysed by acid phosphatase (Ac. P.) to phos- 30°C and 37°C or after 5 minutes at 25°C. A2 – A1 = A
    phate and 1-naphthol, which is converted with FRTR-salt (see "Contents",
    [SUB]) to an azo dye. The increase of absorbance at 405 nm is propor- Calculation
    tional to the total acid phosphatase activity in the sample. Calculate the total acid phosphatase activity in the sample using the
    following factors:
    Reaction Principle
    Ac. P.
    U/l Semi micro / Macro assay
    1-Naphthyl phosphate + H2O phosphate + 1-naphthol Assay temperature 25°C 30°C or 37°C
    1-Naphthol + FRTR-salt azo dye Total acid phosphatase
    A x 149 248
    Contents
    Conversion factor of the traditional units (U/l) in SI-units (kat/l):
    [BUF] 1 x 100 ml Buffer Solution
    Citrate buffer (pH 5.2) 100 mmol/l 1 U/l = 16.67 x 10-3 µkat; 1 µkat/l = 60 U/l
    [SUB] 6 x 15 ml Substrate (powder) Performance Characteristics
    1-Naphthyl phosphate 143 µmol Linearity
    FRTR-salt 15 µmol If the absorbance change exceeds A = 0.3 at 30°C or 37°C or A = 0.5 at
    (4-chloro-2-methylphenyl diazonium salt) 25°C or if the activity is higher than 63 U/l dilute 0.1 ml of sample with
    0.2 ml physiological saline (0.9%) and repeat the assay using this dilution.
    [STAB] 1 x 4 ml Stabiliser Multiply the result by 3.
    Acetic acid 0.7 mmol/l Typical performance data can be found in the Verification Report, acces-
    sible via:
    Additional material recommended but not supplied with the kit
    www.human.de/data/gb/vr/en-acp.pdf or
    [REF] 13160
    [CAL] 4x AUTOCAL www.human-de.com/data/gb/vr/en-acp.pdf
    for 5 ml lyophilised calibrator for HUMAN Clinical Chemistry Reference Values2
    Reagents
    Total acid phosphatase
    [REF] 13511 13512 13951 13151
    Assay temperature 25°C 30°C 37°C
    [CONTROL] HumaTrol N HumaTrol P SERODOS SERODOS plus
    6 x for 5 ml 6 x for 5 ml 6 x for 5 ml 6 x for 5 ml Men up to (U/l) 3.6 5.0 6.5
    Control serum Control serum Control serum Control serum Women up to (U/l) 3.0 4.2 5.5
    normal abnormal normal abnormal
    lyophilised control serum for HUMAN Clinical Chemistry Quality Control
    Reagents For quality control use the control material recommended in “Additional
    material recommended but not supplied with the kit” or other suitable
    Reagent Preparation control material. The control intervals and limits must be adapted to the
    Working reagent (total acid phosphatase determination) individual laboratory requirements. Values obtained should fall within
    Dissolve the contents of one bottle [SUB] with exactly 15 ml of [BUF]. established limits. Each laboratory should establish corrective measures
    Mark label with date of preparation. to be taken if values fall outside the limits.

    Reagent Stability Automation


    The reagents are stable up to the stated expiry date when sealed and Proposals to apply the reagents on analyzers are available on request.
    stored at 2...8°C. Each laboratory has to validate the application in its own responsibility.
    After reconstitution, the working reagent is stable for 5 days at 2...8°C References
    and for 24 hours at 15...25°C protected from light. 1. Hillmann G., Z. Klin. Chem. Klin. Biochem. 9, 273 (1971)
    Specimen 2. Junge W. et al., Data presented at the 45th National Meeting of the
    Serum, no plasma! AACC, New York, July 1993
    Avoid hemolytic and icteric sera!
    Sample Preparation
    EN-ACP INF 1066001 GB 02-2011-14 |
    Stabilise samples, AUTOCAL and controls by addition of one drop of
    [STAB] to 1 ml sample immediately after separation / reconstitution.
    Thus samples remain stable for 3 days at 2...8°C and 24 hours at 15...25°C.
    Assay
    Wavelength: Hg 405 nm
    Optical path: 1 cm
    Temperature: 25°C, 30°C or 37°C
    Measurement: against air (increasing absorbance)

    Human Gesellschaft für Biochemica und Diagnostica mbH


    Max-Planck-Ring 21 · 65205 Wiesbaden · Germany
    Telefon +49 6122-9988-0 · Telefax +49 6122-9988-100 · e-Mail [email protected]

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