[LAB] AST & ALT Flashcards
EC CODE OF AST
2.6.1.1
EC CODE OF ALT
2.6.1.2
NORMAL VALUES OF AST
30C — Up to 28
37C — Up to 40
QUICK:
28–30
40–37
MULTIPLIER OF AST and ALT
x 1768
SI CONVERSION FACTOR OF AST AND ALT
x 16.67
CONC. OF 2-OXOGLUTARATE IN AST REAGENT
0.12 mM
CONC. OF 2-KETOGLUTARATE IN ALT REAGENT
0.13 mM
CONC. OF L-ASPARTATE IN AST REAGENT
200 mM
CONC. OF L-ALANINE IN ALT REAGENT
400 mM
CONC. OF LDH AND MDH IN AST REAGENT
LDH — 800 mM
MDH — 600 mM
DETECTION LIMIT/ LINEARITY OF AST AND ALT
Up to 500 IU/L
TROUBLESHOOTING SAMPLES WITH VALUES HIGHER THAN THE LINEARITY
- Dilute with 1:1 Saline Solution
- Re-assay
- Multiply results by 2
STORAGE CONDITION OF DRY REAGENTS OF AST AND ALT
Store at 2-8C
STABILITY OF RECONSTITUTED REAGENTS
Stable for 8 hours at RT (15-30C)
OR
21 days when refrigerated immediately
STABILITY OF SPECIMEN
Stable at 4C for a minimum of 7 days
ALT ACTIVITY IN HEMOLYZED SERUM
7x ALT ACTIVITY
AST ACTIVITY IN HEMOLYZED SERUM
15x AST ACTIVITY
NORMAL VALUES OF ALT
30C — Up to 26 IU/L
37C — Up to 38 IU/L
QUICK
26 — 30
38 — 37
RECONSTITUTE DRY REAGENT WITH __ mL of ___
12 mL of Distilled Water
WAVELENGTH OF AST AND ALT ASSAYS
340 nm
BLANK SOLUTION COMPOSITION
1.0 mL of Distilled Water
SAMPLE SOLUTION COMPOSITION
1.0 mL Reagent
0.1 mL Patient’s Sample
AST/ALT INCUBATION TEMPERATURE AND PERIOD
37C for 3 minutes
AST/ALT INCUBATION TEMPERATURE AND PERIOD IN BETWEEN READINGS
37C for 1 minute
REACTION PRINCIPLE OF AST
L-aspartate + a-ketoglutarate —AST—> oxaloacetate + glutamate
oxaloacetate + NADH + H —MDH—> malate + NAD
REACTION PRINCIPLE OF ALT
L-alanine + a-ketoglutarate —ALT—> oxaloacetate + pyruvate
pyruvate + NADH + H —LDH—> lactate + NAD
FORMER NAME OF AST
SGOT
Serum Glutamic Oxaloacetate Transferase
FORMER NAME OF ALT
SGPT
Serum Glutamic Pyruvic Transferase
MAIN TISSUE SOURCE OF AST
HEART
LIVER
SKELETAL MUSCLES
MINOR SOURCES OF AST
KIDNEYS
SPLEEN
RBC
PANCREAS
LUNGS
TIP: Where does blood go/flow?
MAJOR TISSUE SOURCE OF ALT
LIVER**
KIDNEYS
MINOR TISSUE SOURCE OF ALT
HEART
SKELETAL MUSCLES
CLINICAL CONDITION IF AST REMAINS NORMAL BUT ALT INCREASES
ALT ↔️
AST ⬆️
MYOCARDIAL INFARCTION
CLINICAL SIGNIFICANCE OF AST
- Verify course of hepatic diseases
- Monitor patients with myocardial infarction, skeletal muscle disease
ABSORBANCE TREND OF AST/ALT
DECREASING
CLINICAL SIGNIFICANCE OF ALT
- Monitor or help diagnose hepatic diseases (ex.: hepatitis, hepatic obstructions)
- Diagnose myocardial infarction (in conjunction with AST)
ULN REACHED TO STOP MEDICATIONS
3x ULN
AMOUNT OF ABSORBANCES RECORDED
4
FACTORS THAT CAUSE FALSE ELEVATIONS
- Hemolyzed samples
- Lipemic or icteric samples
- Pyridoxal phosphate (Coenzyme of AST & ALT btw)
- Pyruvate
- Drugs
DRUGS THAT CAUSE FALSE ELEVATIONS
- Anabolic steroids
- Chloramphenicol
- Antacids
ISOENZYMES OF AST
- Cytoplasmic
- Mitochondrial
ASSAY METHODS USED IN BOTH AST AND ALT
- Reitman - Frankel
- Babson
- Chemiluminesence
HALF LIFE OF AST
24 hours
HALF LIFE OF ALT
16 Hours
AMOUNT OF Hb THAT CAUSES INTERFERENCE
Hb > 45 mg/dL
AMOUNT OF BILIRUBIN AND TRIGLYCERIDES THAT CAUSE INTERFERENCE
Bilirubin > 19 mg/dL
Triglycerides > 650 mg/dL
DO ANTICOAGULANTS AFFECT AST/ALT ACTIVITY?
NO — Which is why plasma can be used.
WHY IS LDH INCLUDED IN AST REAGENTS?
To convert pyruvate to lactate.
Pyruvate intensifies AST activity.
WHAT DOES PYRIDOXAL PHOSPHATE ACTIVATE
THE APOENZYME FORM OF TRANSAMINASE
SOURCE OF PYRIDOXAL PHOSPHATE IN ASSAYS
WATER CONTAMINATED WITH MICROBIAL GROWTH
AMOUNT OF PYRUVATE THAT CAUSES INTERFERENCES IN ALT ASSAYS
Pyruvate > 0.2 mmol/L
ELEVATION PERIODS OF AST IN ANGINA PECTORIS
Rise — 6-8 hrs
Peak — 24 hrs
Return — 3-5 days or Within 5 days
[Assay] OPTIMAL PH IS 7.3 TO 7.8
KARMEN METHOD
REITMAN - FRANKEL
[Assay] ALT ASSAY METHOD RECOMMENDED BY THE IFCC
KARMEN METHOD
[Assay] REACTS WITH 2,4 DINITROPHENYLHYDRAZONE
REITMAN-FRANKEL
[Assay] COLORED COMPOUND OF REITMAN-FRANKEL
BLUE
[Assay] WAVELENGTH OF REITMAN-FRANKEL
550 nm
[Assay] DISADVANTAGE OF REITMAN-FRANKEL
NON-SPECIFIC
Can react with any ketoacid
[Assay] USES DIAZONIUM SALT WITH OXALOACETIC ACID
BABSON
REACTION WITH DIAZONIUM SALT
[Assay] ADVANTAGE OF BABSON
SPECIFIC AND SENSITIVE
Lesser need for reagent blank
[Assay] COLORED COMPOUND OF BABSON
RED
[Assay] ADVANTAGE OF CHEMILUMINESENCE
SENSITIVE
Low background signal level
Wide angle of concentration
[Assay] DISADVANTAGE OF CHEMILUMINESENCE
INHIBITORS
1. Superoxide dismutase
2. N-nitro-Larginine-methyl ester hydrochloride
[Assay] SIDE REACTIONS MONITORED/ DIRECT DETECTIONS MADE BY CHROMATOGRAPHY
- Xanthine
- Glutamate
[Assay] CONCENTRATION OF AST AND GLUTAMATE IN CHROMATOGRAPHY
AST — 0.1 UI-L
GLUTAMATE — 5 UI-L
[Assay] METHOD THAT CAN IDENTIFY ALL COMPONENTS IN THE REACTION
CHROMATOGRAPHY
[Assay] SEPARATION OF AST ISOENZYMES
ELECTROPHORESIS
[Assay] AST ISOENZYME CHARGES
Cytoplasmic — anionic
Mitochondrial — cationic
[Assay] PRINCIPLE OF IMMUNOPRECIPITATION IN AST
Using antibodies against both mitochondrial and cytoplasmic fraction
[Assay] ALT ASSAY METHOD RECOMMENDED BY THE IFCC
WROBLEWSKI AND LA DUE
[Assay] OPTIMAL PH OF WROBLEWSKI AND LA DUE
pH 7.3 to 7.8
[Assay] WAVELENGTH OF WROBLEWSKI AND LA DUE
340 nm
[Assay] PRINCIPLE OF CHEMILUMINESENCE
2H202+ luminol + 20H -> 4H20 + N2+ 3-aminophthalate + hu
BASTA — oxidation of luminol by hydrogen peroxide creates chemiluminesence