GPT ALAT 88 36 15 - AXIOM Solutions

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GPT / ALAT Fluid 5+1
88 36 15 12 x 20 ml
88 13 35 6 x 100 ml
D I A G N O S T I C
IN VITRO TEST FOR THE QUANTITATIVE DETERMINATION OF ALANINE AMINOTRANSFERASE (ALT) IN
HUMAN SERUM AND PLASMA BY IFCC METHOD.
REF
RE
F
QUALITY CONTROL
Control Serum N 6x5ml 88 41 48B
Control Serum P 6x5ml 88 46 85B
The control intervals and limits must be adapted to the individual laboratory and country-specific requirements.
Values obtained should fall within established limits. Each laboratory should establish corrective measures to be
taken if values fall outside the limits.
SUMMARY
Alanine aminotransferase (glutamate pyruvate transaminase) belongs to the group of transaminases which
catalyze the conversion of amino acids to the corresponding !-keto acids via the transfer of amino groups; they
also catalyze the reverse process. Although higher activities exist in the liver, minor activity can also be detected
in the kidneys, heart, skeletal muscle, pancreas, spleen, and lungs. Elevated levels of transaminases are indicative
of myocardial infarction, hepatopathies, muscular dystrophy, and damage to internal organs. Increased ALT
activity in the serum, however, is a rather specific indicator of damage to the liver parenchyma, while AST is not
necessarily a liver-specific parameter.
In 1956, Wroblewski and LaDue described the first kinetic determination of ALT in serum. In 1977 and 1980, the
International Federation of Clinical Chemistry (IFCC) recommended standardized methods for the determination
of ALT with optimized substrate concentrations, use of TRIS buffer*, simultaneous preincubation of serum with
buffer (to avoid competing reactions with NADH), substrate start, and pyridoxal phosphate activation. The
method described here is derived from the IFCC Reference method.
*TRIS = Tris(hydroxymethyl)-aminomethane
TEST PRINCIPLE
UV test according to a standardized method
ALT
2 " oxoglutarate + L " alanine # # $ L " glutamate + pyruvate
ALT is the enzyme which catalyzes this equilibrium reaction. The pyruvate increase is measured in a subsequent
indicator reaction which is catalyzed by lactate dehydrogenase.
pyruvate + NADH+H + " LDH "#
L $ lactate + NAD +
In the second reaction, NADH is oxidized to NAD + . The rate of decrease in NADH (Measured photometrically) is
directly proportional to the rate of formation of pyruvate, and thus the ALT activity.
NOTES
For in vitro diagnostic use.
Exercise the normal precautions required for handling all laboratory reagents.
LIMITATIONS - INTERFERENCE
Criterion: Recovery within ±10% of initial value.
Icterus: No significant interference up to an index I of 20 (approximate conjugated and unconjugated bilirubin: 20
mg/dl)
Hemolysis: No significant interference up to an index H of 1100 (approximate haemoglobin concentration: 1100
mg/dl).
Lipemia (Intralipid): No significant interference up to an index L of 300 (approximate triglycerides concentration:
600 mg/dl). There is poor correlation between turbidity and triglycerides concentration.
Lipemia may cause absorbance flagging as a result of an absorbance increase.
IVD
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D I A G N O S T I C
MEASURING/REPORTABLE RANGE
0.160 at 340 nm or 0.080 at 365 nm
At higher activities dilute the sample with 0.9% NaCl (e.g. 1+2). Multiply the result by the appropriate dilution
factor (e.g. 3)
REFERENCE VALUES
Bases of the IFCC-method and optimised standard-method of the DGKC (1972) at +37°C:
Men
up to 41 U/l or up to 0.68 µkat/l
Women
up to 31 U/l or up to 0.52 µkat/l
Each laboratory should investigate the transferability of the expected values to its own patient population and if
necessary determine its own reference range. For diagnostic purposes, the ALT results should always be assessed
in conjunction with the patients medical history, clinical examination and other findings
ANALYTICAL SENSITIVITY (LOWER DETECTION LIMIT)
Detection limit: 4 U/l or 0.07 µkat/l
The lower detection limit represents the lowest measurable ALT concentration that can be distinguished from
zero.
IMPRECISION
Reproducibility was determined using controls in an internal protocol between day (n = 20). The following results
were obtained:
Between day
Sample Mean U/l SD U/l CV %
Sample 1 44.1 1.34 3.04
Sample 2 115 1.89 1.64
Sample 3 116 2.12 1.83
Reproducibility was determined using controls within protocol between run (n = 20). The following results were
obtained:
Within run
Sample Mean U/l SD U/l CV %
Sample 1 33.5 1.22 3.63
Sample 2 88.3 1.50 1.69
Sample 3 129 1.81 1.41
METHOD COMPARISON
A comparison of the AXIOM GPT / ALAT Fluid 5+1 (y) with a commercial obtainable assay according to the
recommendations of DGKC (1974) (x) gave with 56 samples the following result:
y = 0.974 x + 0,45; r =0.998
REAGENT CONCENTRATION:
R1:
Tris buffer pH 7.8
L-Alanine
LDH
R2:
NADH 2
2-Oxoglutarate
100 mmol/l
500 mmol/l
1200 U/l
0.18 mmol/l
15 mmol/l
PREPARATION AND STABILITY
R1: Ready for use.
R2: Ready for use.
Unopened kit components: Up to the expiration date at +2°C to +8°C.
Onboard stability: R1 28 days
R2
90 days
AXIOM Gesellschaft für Diagnostica und Biochemica mbH; Siegfriedstraße 14, 67547 Worms, Germany
Tel.: 0049-6241-50040 Fax: 0049-6241-5004499 e-mail: info@axiom-online.net webseite: www.axiom-online.net
AXIOM Gesellschaft für Diagnostica und Biochemica mbH; Siegfriedstraße 14, 67547 Worms, Germany
Tel.: 0049-6241-50040 Fax: 0049-6241-5004499 e-mail: info@axiom-online.net webseite: www.axiom-online.net

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D I A G N O S T I C
SPECIMEN
Collect serum using standard sampling tubes.
Heparin or EDTA plasma.
Stability: 24 hours at +20°C to +25°C
3 days at +2°C to +8°C
Separate serum/plasma from clot/cells within 8 hours at room temperature or 48 hours at +2°C to +8°C.
Centrifuge samples containing precipitate before performing the assay
TESTING PROCEDURE
Materials provided
• Working solutions as described above
Additional materials required
• Controls as indicated below
• 0.9% NaCl
Manual procedure serum start:
Wavelength
Hg 334 nm, 340 nm or Hg 365 nm
Temperature
+25°C / +30°C / +37°C
Cuvette
1cm light path
Zero adjustment
against air
Mix R1 and R2 5+1. This solution is stable: up to 10 days at +2°C to +8°C or
up to 1 day at +20°C to +25°C
Reagent mixture 1000 µl
Serum / plasma 100 µl
Mix and incubate 60 seconds at assay temperature and start stopwatch simultaneously. Read again after exactly
1, 2 and 3 minutes.
Calculation:
Hg 365 nm
Hg 340 nm
Hg 334 nm
3235 x !A/min
1746 x !A/min
1780 x !A/min
Manual procedure substrat start:
Wavelength
Hg 334 nm, 340 nm or Hg 365 nm
Temperature
+25°C / +30°C / +37°C
Cuvette
1cm light path
Zero adjustment
against air
R 1 1000 µl
Serum / plasma 100 µl
Mix and incubate 60 seconds at assay temperature then add
R2 200 µl
Mix, read initial absorbance and start stopwatch simultaneously. Repeat reading after exactly 1, 2 and 3 min.
Calculation:
Hg 365 nm
Hg 340 nm
Hg 334 nm
CALIBRATION FREQUENCY
Two point calibration is recommended
• after reagent lot change
• as required following quality control procedures
Calibration verification: Not necessary
3823 x !A/min
2063 x !A/min
2103 x !A/min
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DISPOSAL
Please note the legal regulations.
D I A G N O S T I C
LITERATURE
1. Bablok W et al. A General Regression Procedure for Method Transformation. J Clin Chem Clin Biochem 1988; 26:783-790.
2. Berg Meyer HU, Herder M, Red R. Approved recommendation (1985) on IFCC methods for the measurement of catalytic
concentration of enzymes. Part 3. (FCC Method for alanine aminotransferase. J Clin Chem Clin Biochem 1986; 24:481-489.
3. Glick MR, Ryder KW, Jackson SA. Graphical Comparisons of Interferences in Clinical Chemistry Instrumentation.
Clin Chem 1986;32:470-474.
4. Greiling H, Gressner AM (Hrsg.). Lehrbuch der Klinischen Chemie und Pathobiochemie, 3rd Stuttgart/New York:
Schattauer Verlag, 1995.
5. Passing H, Bablok W. A New Biometrical Procedure for Testing the Equality of Measurements from Two Different Analytical
Methods. J Clin Chem Clin Biochem 1983;21 :709-720.
6. Thefeld W et al. Dtsch med Wschr 1974;99:343.
7. Tietz NW (Hrsg.). Clinical Guide to Laboratory Tests, 3rd Philadelphia, Pa: WB Saunders, 1995:20-21.
8. Wallnöfer H, Schmidt E, Schmidt FW (Hrsg.). Synopsis der Leber- krankheiten. Stuttgart: Georg Thieme Verlag, 1974.
9. Wroblewski F LaDue JS. Ann Intern Med 1956;45:801.
10. Wroblewski F, LaDue JS. Proc Soc Exp Biol Med 1956;91 :569.
AXIOM Product range Clinical Chemistry
Enzymes Ions Other Metabolites
Acid Phosphatase Ammonium fluid Bilirubin T/D
Alkaline Phosphatase Copper fluid Creatinine fluid
!-Amylase direct Calcium fluid Glucose GOD-PAP fluid
CK-NAC actived Chloride fluid Glucose Hexokinase fluid
CK-MB (NAC- actived) Inorganic Phosphorus UV fluid Urea Enzymatic fluid
"-GT fluid Iron fluid Urea UV fluid
LDH fluid TIBC Uric Acid PAP fluid
Cholinesterase
Magnesium fluid
GOT/ASAT fluid
Potassium fluid
GPT/ALAT fluid
Sodium fluid
Controls
Lipase UV fluid
Control Serum N
Lactate PAP
Control Serum P
Proteins
!-HBDH
Lipids
Albumin
CSF-Protein fluid
Cholesterol fluid
Microprotein fluid
HDL Cholesterol
Hemoglobin
LDL Cholesterol
Protein Total fluid
Triglycerides fluid
06/09 M/kd
AXIOM Gesellschaft für Diagnostica und Biochemica mbH; Siegfriedstraße 14, 67547 Worms, Germany
Tel.: 0049-6241-50040 Fax: 0049-6241-5004499 e-mail: info@axiom-online.net webseite: www.axiom-online.net
AXIOM Gesellschaft für Diagnostica und Biochemica mbH; Siegfriedstraße 14, 67547 Worms, Germany
Tel.: 0049-6241-50040 Fax: 0049-6241-5004499 e-mail: info@axiom-online.net webseite: www.axiom-online.net