[M1] Enzyme Methodologies Flashcards
Liver enzymes is also called as
Hepatic enzymes
Enumerate the Liver Enzymes
Alkaline Phosphatase
Aspartate Aminotransferase
Alanine Aminotransferase
Gamma-Glutamyltransferase
5’-Nucleotidase
ALP
ALKALINE PHOSPHATASE
E.C OF ALP
E.C. 3.1.3.1
Catalyze the hydrolysis of phosphomonoesters at an alkaline pH (9.0-10.0)
ALKALINE PHOSPHATASE
ALP catalyzes the hydrolysis of _________________ at an (acidic/alkaline) pH (_____________)
ALP
phosphomonoesters
alkaline
9.0-10.0
Common name of ALP
alkaline orthophosphoric monoester phosphohydrolase
ALP
Function:
remove (organic/inorganic) phosphate from an (organic/inorganic) phosphate ester with the concomitant production of ___________
inorganic
organic
alcohol
T/F: ALP is a non-specific enzyme
T
Activators of ALP
Mg2+
Co2+
Mn2+
inhibitors of ALP
phosphate
borate
oxalate
ALP is useful for evaluation of ___________ and ________ disorders
hepatobiliary
bone
RF of ALP in 20-50 MALE
53-128 U/L
RF of ALP in 20-50 FEMALE
42-98 U/L
RF of ALP in ≥60 MALE
56-119 U/L
RF of ALP in ≥60 FEMALE
53-141 U/L
RF of ALP in 4-15
54-369 U/L
What are the major ALP isoenzymes
Liver ALP
Bone ALP
Placental ALP
Intestinal ALP
Half life of Liver ALP
3 days
Half life of Bone ALP
1 day
Half life of Placental ALP
7 days
Half life of Intestinal ALP
1 day
Rank the ALP isoenzymes from most ANODAL to least ANODAL
- Liver ALP
- Bone ALP
- Placental ALP
- Intestinal ALP
Rank the ALP isoenzymes from most CATHODAL to least CATHODAL
- Intestinal ALP
- Placental ALP
- Bone ALP
- Liver ALP
Rank the ALP isoenzymes from most heat stable to least heat stable
- Placental ALP
- Intestinal ALP
- Liver ALP
- Bone ALP
Rank the ALP isoenzymes from most heat-labile to least heat-labile
- Bone ALP
- Liver ALP
- Intestinal ALP
- Placental ALP
inhibitor of Liver ALP
Levamisole
inhibitor of Bone ALP
Levamisole &
3M urea
Inhibitor of Placental ALP
Phenylalanine
Inhibitor of Intestinal ALP
Phenylalanine
BEFORE HEAT
Placental ALP
Liver ALP
Bone ALP
100%
100%
100%
AFTER HEAT
Placental ALP
Liver ALP
Bone ALP
100%
>20%
<20%
Heat Fractionation/ Stability Test is measured @ ____ for ____ minutes
56°C
10 minutes
T/F: ALP isoenzymes are NOT measured before and after heating
F; Measured before and after heating
separates liver and bone ALP
Neuraminidase
Wheat Germ Lectin
Steps in Heat Fractionation/ Stability Test
- Measure ALP
- Heat sample @ 56°C for 10 minutes
- Measure ALP
Heating sample @ 56°C for 10 minutes significance
To inactivate other isoenzymes
Abnormal ALP isoenzyme associated with neoplasms or cancer
Carcinoplacental ALPs
Carcinoplacental ALPs
Abnormal ALP isoenzyme associated with __________ or _________
neoplasms
cancer
2 types of Carcinoplacental ALPs
Regan ALP
Nagao ALP
Detected in gynecological CA (ovarian, breast), lung CA, and colon CA.
Regan ALP
Regan ALP
Detected in ________ (ovarian, breast), ___ CA, and ____ CA.
gynecological CA
lung CA
colon CA
Migrates to the same position as the Bone ALP
Regan ALP
Regal ALP migrates to the same position as ___________
Bone ALP
Most heat stable ALP isoenzyme
Regan ALP
T/F: Regan ALP is more heat stable than placental ALP
T
Regan ALP is stable @ ______for ____minutes
65°C
30
Inhibitor of Regan ALP
phenylalanine
Detected in metastatic carcinoma of pleural surfaces, and adenocarcinoma of pancreas and bile ducts
Nagao ALP
Nagao ALP
Detected in _______________ of pleural surfaces, and ____________ of pancreas and bile ducts
metastatic carcinoma
adenocarcinoma
Nagao ALP
Detected in metastatic carcinoma of _________________, and adenocarcinoma of _______ and _________
pleural surfaces
pancreas and bile ducts
Variant of Regan ALP
Nagao ALP
Inhibitor of Nagao ALP
phenylalanine
L-Leucine
Method of Analysis for ALP
Bowers and McComb
A continuous monitoring method allowing calculation of ALP activity based on the molar absorptivity of _________________
BOWERS AND MCCOMB
p-nitrophenol
The most specific method; IFCC recommended method
Bowers and McComb
Enumerate the Other Methodologies for ALP
- Bodansky
- Shinowara
- Jones
- Reinhart
- King and Armstrong
- Bessy, Lowry, and Brock
- Bowers and McComb
- Huggins and Talalay
- Moss
- Klein, Babson and Read
Substrate for
- Bodansky
- Shinowara
- Jones
- Reinhart
Beta-Glycerol Phosphate
End Product for
- Bodansky
- Shinowara
- Jones
- Reinhart
Inorganic phosphate + glycerol
Substrate for King and Armstrong
Phenylphosphate
End product for King and Armstrong
Phenol
Substrate for
- Bessy, Lowry, and Brock
- Bowers and McComb
p-nitrophenyl phosphate
End product for
- Bessy, Lowry, and Brock
- Bowers and McComb
p-nitrophenol/
yellow nitrophenoxide ion
Substrate for Huggins and Talalay
Phenolphthalein phosphate
End product for Huggins and Talalay
Phenolphthalein red
Substrate for Moss
Alpha-naphthol phosphate
End product for Moss
Alpha-naphthol
Substrate for Klein, Babson, and Read
Buffered phenolphthalein phosphate
End product for Klein, Babson, and Read
Free phenolphthalein
False Increase of ALP
Hemolysis
Diet (Fatty meals)
Stored at low temperature (4°C)
FALSE INC. IN ALP
Hemolysis - ALP in RBC is ___ more concentrated than serum or plasma
6x
FALSE INC. IN ALP
Diet (fatty meals) - ALP is ____ higher
25%
Diagnostic Significance of ALP
ALP - #1 marker in ______________
obstructive jaundice
Increased ALP
● Osteitis deformans
● Obstructive Jaundice
● Osteomalacia
● Rickets
● Osteoblastic bone tumors
● Sprue
● Hyperparathyroidism
● Hepatitis and Cirrhosis
Osteitis deformans is also known as
“Paget’s disease”
AST
ASPARTATE AMINOTRANSFERASE
E.C of ASPARTATE AMINOTRANSFERASE
E.C. 2.6.1.1
Old name of ASPARTATE AMINOTRANSFERASE
Serum Glutamic
Oxaloacetate Transaminase
ASPARTATE AMINOTRANSFERASE
Catalyze the transfer of amino groups
between _________ (substrate) and
___________
aspartate
a-ketoacids
ASPARTATE AMINOTRANSFERASE
Catalyze the transfer of amino groups
between aspartate (substrate) and
a-ketoacids with the formation of
________ and ___________
oxaloacetate
glutamate
ASPARTATE AMINOTRANSFERASE
Coeznyme
Pyridoxal phosphate
ASPARTATE AMINOTRANSFERASE
Major Tissue Source
Cardiac tissues
liver and skeletal muscle
ASPARTATE AMINO TRANSFERASE
RR
5-35 U/L
ASPARTATE AMINOTRANSFERASE
Isozenzymes
Cytoplasmic AST
Mitochondrial AST
Predominant in the circulation of a healthy
individual
Cytoplasmic AST
Method of Analysis for AST
Karmen Method
Karmen Method is what type of enzymatic method
Coupled enzymatic method
Indicator enzyme for Karmen method
Malate
dehydrogenase (MD)
KARMEN METHOD (AST)
It monitors the (increase/decrease) in
absorbance at _____ nm
decrease
340
KARMEN METHOD (AST)
Storage: stable for ____ days at __________
3-4
refrigerated temp.
Variables for Karmen Method
Hemolysis
Hemolysis in Karmen Method results in false (increase/decrease) up to ___
increase
10x
Exhibits highest level AST
Acute Hepatocellular Disorder
ALT
ALANINE AMINOTRANSFERASE
E.C OF ALANINE AMINOTRANSFERASE
● E.C 2.6.1.2
old name for Alanine Aminotransferase
Serum Glutamic Pyruvic Transaminase
ALANINE AMINOTRANSFERASE
Catalyze transfer of amino group
between _________ (substrate) and
______________
alanine
a-ketoglutarate
ALANINE AMINOTRANSFERASE is significant in the evaluation of what disorders
Hepatic disorders
ALANINE AMINOTRANSFERASE (ALT)
Coenzyme
Pyridoxal phosphate
ALANINE AMINOTRANSFERASE (ALT)
Major Tissue Source
Liver
ALANINE AMINOTRANSFERASE (ALT)
RR
7-45 U/L
Method of Analysis for ALANINE AMINOTRANSFERASE (ALT)
- Coupled Enzymatic Method
ALT
Indicator Enzyme for Coupled Enzymatic Method
Lactate Dehydrogenase (LDH)
Coupled Enzymatic Method (ALT)
The change in absorbance at ______ nm
measured ________________ is directly
proportional to ALT activity (pH
_________).
340
continuously
7.3-7.8
Coupled Enzymatic Method (ALT)
Storage
3-4 days at 4°C
T/F: Hemolysis does NOT affect ALT
True
Increase Aminotransferases (AST, ALT)
- Toxic Hepatitis
- Acute Myocardial Infarction (AST)
- Wolff-Parkinson White Syndrome
- Trichinosis (AST)
- Chronic alcoholism
- Dermatomyositis
- Hepatic cancer
- Reye’s Syndrome
- Viral Hepatitis
- Muscular dystrophy (AST)
- Acute pancreatitis (AST)
Which disorders/diseases causes Increase in AST only
Acute Myocardial Infarction (AST)
Trichinosis (AST)
Muscular dystrophy (AST)
Acute pancreatitis (AST)
Acute Myocardial Infarction (AST)
Rise
Peak
Normal
6-8 hours
24 hours
within 5 days
Trichinosis is caused by
Trichinella spiralis
unknown source in GGT is caused by pancreas
Acute pancreatitis
AST/ALT ratio
DE RITIS RATIO
Used to differentiate the cause
of hepatic disorder
DE RITIS RATIO
DE RITIS RATIO
> 1: __________
non-viral cause
DE RITIS RATIO
<1: __________
viral
GGT
GAMMA-GLUTAMYLTRANSFERASE
E.C of GAMMA-GLUTAMYLTRANSFERASE
E.C. 2.3.2.2
GAMMA-GLUTAMYLTRANSFERASE catalyzes ____________________
transpeptidation
The transfer of ɣ-glutamyl residue
from ɣ-glutamyl peptides to amino
acids, water and other peptides
transpeptidation
Used for diagnosis of hepatobiliary
disorders and alcoholism
GAMMA-GLUTAMYLTRANSFERASE
(GGT)
GAMMA-GLUTAMYLTRANSFERASE
(GGT) is sed for diagnosis of _____________
and _____________
hepatobiliary disorders
alcoholism
Useful for differentiating source of
serum ALP elevation
GAMMA-GLUTAMYLTRANSFERASE
(GGT)
LIVER DISORDER
ALP
GGT
Increased
Increased
BONE DISEASE
ALP
GGT
Increased
Normal
Critical for the intracellular
maintenance of reduced glutathione
GAMMA-GLUTAMYLTRANSFERASE
(GGT)
GAMMA-GLUTAMYLTRANSFERASE (GGT)
Tissue source
Liver (epithelial cell lining of biliary ducts and bile canalicular)
Kidneys
Brain
Pancreas,
Intestine
Prostate.
Method of Analysis for GAMMA-GLUTAMYLTRANSFERASE (GGT)
Szasz Assay
Method for Szasz Assay
Fixed-point or continuous monitoring
Szasz Assay
Substrate
γ-glutamyl-p-nitroanilide
Szasz Assay
End product
p-nitroaniline
Szasz Assay
wavelength
405-420 nm
Szasz Assay
Preferred specimen
Serum
EDTA Plasma
Szasz Assay
Storage:
4°C (1 week)
-20°C (1 month)
Other Method for GGT
Rosalki and Tarrow, Orlowski
T/F; GGT is not effected by hemolysis
T
Why is GGT not affected by hemolysis?
GGT is not found in RBC
GGT
RR: Male
6-55 U/L
GGT
RR: female
5-38 U/L
Most sensitive marker of acute
alcoholic hepatitis
GAMMA-GLUTAMYLTRANSFERASE
(GGT)
Normal levels in patients with bone
disease and during pregnancy
GAMMA-GLUTAMYLTRANSFERASE
(GGT)
GGT
Normalize: ___ weeks after consumption
2-3
5’N
5’-NUCLEOTIDASE
E.C. of 5’-NUCLEOTIDASE
E.C. 3.1.3.5
5’-NUCLEOTIDASE
Other name
5’ ribonucleotide phosphohydrolase
A phosphoric monoester hydrolase
reacting only on nucleoside-5’-phosphates (_______,
__________) releasing (inorganic/organic) phosphate
5’-NUCLEOTIDASE
AMP
adenylic acid
inorganic
5’-NUCLEOTIDASE
Major Source
Liver
Marker of hepatobiliary disease and
infiltrative lesions of the liver
5’-NUCLEOTIDASE
5’-NUCLEOTIDASE
Marker of _____________ and
____________________ of the liver
hepatobiliary disease
infiltrative lesions
Secondary Marker for Obstructive Jaundice
5’-NUCLEOTIDASE
Method for 5’-NUCLEOTIDASE
Dixon & Purdon
Campbell
Belfield & Goldberg
Method for 5’-NUCLEOTIDASE
Dixon & Purdon
Campbell
Belfield & Goldberg
5’-NUCLEOTIDASE
RR
0-1.6 U/L
5’-NUCLEOTIDASE
Storage
4°C (4 days)
-20°C (4 months)
Enumerate the Pancreatic Enzymes
- Amylase
- Lipase
AMS
Amylase
E.C for Amylase
E.C. 3.2.1.1
AMYLASE
Other name
alpha-1,4-glucan-4-glucohydrolase
AMYLASE
Catalyzes the hydrolysis of __________________ in
polysaccharides (_____, _____)
𝛂-1,4-glycosidic bonds
starch, glycogen
Earlier pancreatic marker
Amylase
Smallest enzyme (freely filtered by the
_________)
AMYLASE
glomerulus
AMYLASE
Activators
Calcium, Chloride
AMYLASE
Major Tissue Source
Pancreas (acinar cells)
Salivary Glands
AMYLASE (AMS)
RR; Serum
28-100 U/L
AMYLASE (AMS)
RR; Urine
1-15 U/h
Enumerate the isoenzymes of Amylase
- S-type/Ptyalin/Salivary
- P-type / Amylopsin / Pancreatic
↑ parotitis/mumps
- S-type/Ptyalin/Salivary
The most anodal AMY isoenzyme
- S-type/Ptyalin/Salivary
S-type/Ptyalin/Salivary is secreted by __________________
salivary glands
Inhibitor of S-type/Ptyalin/Salivary (inhibits activity of salivary glands)
Wheat Germ Lectin
↑ in acute pancreatitis
P-type / Amylopsin / Pancreatic
the most predominant subtype of P-type / Amylopsin / Pancreatic
P3
● Secreted by the pancreas and also by
the fallopian tube & lungs
P-type / Amylopsin / Pancreatic
T/F: In the laboratory, we measure the Total Amylase
TRUE
Specimen for Amylase
Serum,
Heparinized Plasma
Substrate for Amylase
Starch
Enumerate the different Method of Analysis of Amylase
- Saccharogenic
- Amyloclastic
- Chromogenic
- Coupled-Enzyme
AMYLASE
Classic reference method
Saccharogenic
It measures the amount of __________ produced by the hydrolysis of ______ by the usual glucose method
SACCHAROGENIC
reducing sugars
starch
Enumerate the glucose methods used in Saccharogenic
Hexokinase
Glucose oxidase
Glucose methodology reference method
Hexoklnase
End product for Saccharogenic
Glucose
AMYLASE
inverse method
Amyloclastic
It measures decrease in starch substrate
Amyloclastic
Amyloclastic
Substrates are coupled with_____
iodine
Iodine + Starch = _____________
Dark-blue color
Why Dark Blue Color in Amyloclastic?
e iodine is trapped
within the structure of the starch
T/F: In amyloclastic, decrease in color is proportional to AMS activity
T
Measures the formation of soluble
starch fragments coupled with
chromogenic dyes.
Chromogenic
Color intensity is proportional to AMS
activity
Chromogenic
Measured amylase activity by a continuous-monitoring/kinetic technique.
Coupled-Enzyme
Substrate for Couple-Enzyme (Amylase)
Starch
Wavelength for Couple-Enzyme (Amylase)
340 nm
pH for Couple-Enzyme (Amylase)
6.9
Storage for Couple-Enzyme (Amylase)
Room temp (1 week)
4°C (2 months)
False Decrease in AMS
● Ca2+ Chelating Anticoagulant
● Triglycerides
False Increase in AMS
● Morphine, other opiates
amylase bound to immunoglobulin
Macroamylasemia
asymptomatic amylasemia
Macroamylasemia
In Macroamylasemia, (increase/decrease) serum AMS & (increase/decrease) N Urine AMS
Increase
Decrease
↑ serum AMS & ↑ urine AMS
Hyperamylasemia
Amylase/Creatinine Ratio:
Normal
1-4% (0.01-0.04)
Amylase/Creatinine Ratio:
Acute Pancreatitis
> 4%-15%
LPS
Lipase
E.C of Lipase
E.C. 3.1.1.3
LIPASE
Other name
Triacylglycerolacylhydrolase
Catalyzes hydrolysis of glycerol esters
of complex lipids to produce ___________
and ________.
LIPASE
alcohol
fatty acid.
Catalyzes partial hydrolysis of ________ to________________,
with production of long-chain fatty
acids.
LIPASE
dietary TAG
2-monoglyceride intermediate
More pancreatic specific
LIPASE
LIPASE
Cofactor
Colipase (coenzyme)
Bile salts
LIPASE
Major Tissue Source
Pancreas
LIPASE
RR
<38 U/L
Method of Analysis for Lipase
Specimen
Serum
Method of Analysis for Lipase
Storage
room temp (1 week)
4°C (3 weeks)
Method of Analysis for Lipase
Interferences
Hemolysis
Enumerate the Methods for Lipase
- Cherry-Crandall
- Tietz
- Peroxidase Coupling
Substrate for Cherry-Crandall and Tietz
50% Olive oil (Triolein)
Titrating Agent for Cherry-Crandall and Tietz
0.4N NaOH
Indicator for Cherry-Crandall
Phenolphthalein
Indicator for Tietz
Thymolphthalein + Veronal
Endpoint for Cherry-Crandall and Tietz
Fatty Acids (Oleic acid)
End color of Cherry-Crandall
Pink
End color of Tietz
Blue
Most commonly used method (LPS)
Peroxidase Coupling
Does not use 50% olive oil (LPS)
Peroxidase Coupling
most specific pancreatic marker
Lipase
In chronic pancreatitis:
→ _____________ are destroyed
→ Loss of_____ and _____
Acinar cells
AMS and LPS
AMYLASE
Rise
Peak
Normalize
5-8 HOURS
24 HOURS
3-5 DAYS
LIPASE
Rise Peak Normalize
4-8 HOURS
24 HOURS
8-14 DAYS
Enumerate the Cardiac Markers
- Creatine Kinase
- Lactate Dehydrogenase
CK
Creatine Kinase
E.C for Creatine Kinase
● E.C. 2.7.3.2
CREATINE KINASE
Other Name
ATP-Creatine-N-phosphotransferase
Catalyzes phosphorylation of creatine
to form _____________
CREATINE KINASE
creatine phosphate.
Involved in the storage of high-energy
creatine phosphate in ___________
CREATINE KINASE
muscle cells
contains two subunits
CREATINE KINASE
Dimeric Molecule
2 subunits of CREATINE KINASE
→ M subunit (Muscle)
→ B subunit (Brain)
CREATINE KINASE
Major Tissue Source
brain
muscles (smooth, skeletal, cardiac)
CREATINE KINASE
RR: Male
46-171 U/L
CREATINE KINASE
RR: Female
35-145 U/L
CREATINE KINASE
RR: CK-MB
<5% of total CK
Enumerate the Normal Isoenzymes of Creatine Kinase
CK-BB
CK-MB
CK-MM
Rank the CK isoenzymes from most ANODAL to least ANODAL
- CK-BB
- CK-MB
- CK-MM
Rank the CK isoenzymes from most CATHODAL to least CATHODAL
- CK-MM
- CK-MB
- CK-BB
Brain type CK Isoenzyme
CK-BB
Hybrid type CK Isoenzyme
CK-MB
Muscle type CK Isoenzyme
CK-MM
Dominant in brain, intestines, and smooth muscles
CK-BB
CK-BB is dominant in:
brain
intestines
smooth muscles
Present in significant concentration in the cardiac
muscles
CK-MB
CK-MB is present in significant concentration in the ___________________
cardiac muscles
Abundantly present in striated muscles
CK-MM
CK-MM is abundantly present in _________
striated muscles
Rarely found in Serum
CK-BB
Why is CK-BB rarely found in serum?
it cannot pass through the blood-brain barrier
Serodiagnostic tool for AMI
CK-MB
Major Isoenzyme in normal
individual (______)
CK-MM
94-100%
Useful Non-Specific Tumor Marker
CK-BB
RV of CK-MB
<5% of Total CK
Myocardial Damage/AMI:
● Elevated CK-MB
● >6 of Total CK
earliest enzymatic cardiac marker
CK-MB
increased CREATINE KINASE
- Acute Myocardial Infarction
- Duchenne-type Muscular Dystrophy
- Rhabdomyolysis
- Cerebrovascular Accident
- Seizures
- Nerve Degeneration
- CNS Shock
- Hypothyroidism
- Malignant Hyperpyrexia
- Reye’s Syndrome
Highest elevation of total CK
Duchenne-type Muscular Dystrophy
Method of Analysis for CK
Specimen
Serum
Heparinized Plasma
Inhibitor of sulfhydryl group oxidation
→ N-acetylcysteine
→ Mercaptoethanol
→ Thioglycerol
→ Dithiothreitol
T/F: Serum activity in CK is stable
FALSE; VERY UNSTABLE
Why is serum in CK very unstable?
because it is easily be inactivated by sulfhydryl
group by oxidation
Variables for CK
Hemolysis
Light
Non-heparinized Anticoagulant
Physical Activity
IM Injection
VARIABLES FOR CK
Hemolysis hgb level
> 320 mg/L HGB
Total CK in Hemolysis is falsely elevated due to ______________ (enzyme) that is present in RBC
Adenylate kinase
To prevent false elevation because of adenylate kinase, add __________________
Adenosine monophosphate
Why is light a variable in CK
because CK is inactivated if exposed to light
Methods for CK
- Tanzer-Gilvarg Assay
- Oliver-Rosalki Method
Methods for CK
Forward/Direct Method
- Tanzer-Gilvarg Assay
Tanzer-Gilvarg Assay is coupled with _______________________________________________
pyruvate kinase-lactate dehydrogenase-NADH system
Tanzer-Gilvarg Assay
Optimal pH
9.0
Tanzer-Gilvarg Assay
Wavelength
340 nm
Methods for CK
Reverse/indirect Method
Oliver-Rosalki Method
Most commonly performed method for CK
Oliver-Rosalki Method
Oliver-Rosalki Method is couple with _______________________________________________________
Hexokinase-Glucose-6- Phosphate-Dehydrogenase-NADP system
Oliver-Rosalki Method
Optimal pH
6.8
Oliver-Rosalki Method
Wavelength
340 nm
LDH
Lactate Dehydrogenase
E.C of Lactate Dehydrogenase
E.C. 1.1.1.27
Catalyze the interconversion (reversible) of ________ and __________
LACTATE DEHYDROGENASE
lactic acid
pyruvic acid
LDH
Coenzyme
NAD+
LDH
Inhibitor
Ethylenediamine tetraacetic acid (EDTA)
contain A-peptide or M-peptide unit
LACTATE DEHYDROGENASE
Tetrametic Molecule
● Widely distributed
LACTATE DEHYDROGENASE (LDH)
LDH
Highest Activity
Heart
RBCs
skeletal muscles
LDH
RV
125-220 u/L
Enumerate the LDH Isoenzymes
LD 1
LD 2
LD 3
LD 4
LD 5
LD 6
Most predominant LDH isoenzyme
LD 2
increased in pulmonary involvement and various
carcinomas
LD3
LD3 is increased in ______________ and _____________________
pulmonary involvement
various carcinomas
LDH Isoenzyme
Seen in muscular injuries
LD 4
LD 6 a.k.a
Alcohol dehydrogenase
Present in Arteriosclerotic cardiovascular failure
LD 6
LD6 is present in _________________________
Arteriosclerotic cardiovascular failure
SUBUNITS
LD 1
LD 2
LD 3
LD 4
LD 5
LD 6
HHHH
HHHM
HHMM
HMMM
MMMM
none
Tissue source of LD 1
Heart, RBC
Tissue source of LD 2
Heart, RBC
Tissue source of LD 3
Liver
Spleen,
Lymphocytes
Pancreas
Tissue source of LD 4
Liver
Tissue source of LD 5
Skeletal muscle
Percentage based on Total LDH
LD 1
14-26%
Percentage based on Total LDH
LD 2
29-39%
Percentage based on Total LDH
LD 3
20-26%
Percentage based on Total LDH
LD 4
8-16%
Percentage based on Total LDH
LD 5
6-16%
used as tumor markers
LD 2
LD 3
LD 4
LD2, LD3, LD4 -used as tumor markers for:
→ acute leukemia
→ germ cell tumor
→ breast cancer
→ lung cancer
Most to least dominant cases of: (Descending Order)
Normal Pattern
LD 2>1>3>4>5
Method of Analysis LDH
Specimen
Serum
Method of Analysis LDH
Substrate
→ Lactate
→ Pyruvate
→ α-hydroxybutyrate (LD1)
Method of Analysis LDH
Storage: Total LD
25°C (48 hours)
Method of Analysis LDH
Storage: LD isoenzymes
25°C (24 hours)
Variables for LDH
→ Plasma specimen
→ Hemolysis
→ Cold storage
Plasma specimen and Hemolysis in LDH causes false (increase/decrease) because of the presence of ____________
increase
plaelets
Cold Storage of LDH causes false (increase/decrease) in _____ and _____
Decrease
LD 4 and LD 5
Enumerate the methods used in LDH
- Wacker Method
- Wrobleuski La Due
Method of Analysis LDH
Forward/Direct Method
Wacker Method
Method of Analysis (LDH)\
Most commonly used method
Wacker Method
pH in Wacker Method
8.3-8.9
Wavelength in Wacker Method
340 nm
Method of Analysis LDH
Reverse/Indirect Method
Wrobleuski La Due
Wrobleuski La Due is ____ the rate of forward method
thrice
● Preferred Method for Dry-Slide
Technology
Wrobleuski La Due
Example of Dry-Slide Techbology
Vitros Analyzer
Uses less costly cofactor and it has a
smaller specimen volume requirement
Wrobleuski La Due
Wrobleuski La Due
pH
7.1-7.4
Increased LDH
- Acute Myocardial Infarction
- Hemolytic anemia
- Pernicious anemia
- Pulmonary infarction
- Muscle dystrophy
- Hepatic carcinoma, toxic hepatitis,
cirrhosis, viral hepatitis - Blood transfusion
- Pneumocystis jirovecii infection
INCREASED LDH
Acute Myocardial Infarction exhibits what type of pattern
→ Flipped pattern
→ LD 1>2>3>4>5
INCREASED LDH
Pulmonary Infarction exhibits what pattern
LD 3>4>2>1>5
INCREASED LDH
Muscle dystrophy what pattern
LD 5>4>3>2>1
Which disorders/diseases in Increased LDH exhibits increased LD 4
Hepatic carcinoma, toxic hepatitis,
cirrhosis, viral hepatitis
__ & __ = LD is increased but normalize after ______
6 and 7
24 hours
T/F: Increase in Total LDH is insignificant
T
Why is an increase in Total LDH insignificant?
because LDH is present almost virtually in cells
Acute Myocardial infarction
______ of total CK ______
> 6%
1:20:0
Acute Myocardial infarction
Enumerate the Enzymes involved
Aspartate Aminotransferase
Creatine Kinase-MB
Lactate Dehydrogenase
ACUTE MYCOCARDIAL INFARCTION
Enzyme not liver specific
Aspartate aminotransferase
ACUTE MYOCARDIAL INFARCTION
Enzyme cardiac specific
Creatine Kinase-MB
ASPARTATE AMINOTRANSFERASE
Rise
Peak
Normalize
6-8 hours
24 hours
Within 5 days
CREATINE KINASE-MB
Rise
Peak
Normalize
4-8 hours ‘
12-24 hours
48-72 hours
LACTATE DEHYDROGENASE
Rise
Peak
Normalize
12-24 hours
48-72 hours
After 10-14 days
Enumerate the other Clinically Significant Enzymes
- Acid Phosphatase
ACP
Acid Phosphatase
E.C of ACID PHOSPHATASE
E.C. 3.1.3.2
ACID PHOSPHATASE
Other name
Acid orthophosphoric
monoester phosphohydrolase
Catalyze the hydrolysis of phosphomonoesters at an acidic pH
ACID PHOSPHATASE (ACP)
ACID PHOSPHATASE
Major Tissue Source
Prostate
Useful for evaluation of metastatic-prostatic carcinoma
ACID PHOSPHATASE (ACP)
ACID PHOSPHATASE (ACP) i useful for evaluation of _____________________
metastatic-prostatic carcinoma
ACID PHOSPHATASE
TOTAL ACP
RR: Male
2-5-11.7 U/L
ACID PHOSPHATASE
PROSTATIC ACP
RR: Male
0.2-5.0 U/L
ACID PHOSPHATASE
TOTAL ACP
RR: Female
0.3-9.2 U/L
ACID PHOSPHATASE
PROSTATIC ACP
RR: Female
0.0-0.8 U/L
T/F: Total ACP is measured in the laboratory
True
Method of Analysis (ACP)
Specimen
Serum
Method of Analysis ACP
Substrates
Thymolphthalein monophosphate
α-naphthyl phosphate
Substrate to use in ACP for end-point methods
Thymolphthalein monophosphate
Specific substrate for prostatic ACP
Thymolphthalein monophosphate
ACP substrate for continuous monitoring methods/kinetic assay
α-naphthyl phosphate
Methods of Analysis ACP
INHIBITORS
L-Tartrate
2% formaldehyde, cupric sulfate solution
Inhibitor in ACP that inhibits prostatic ACP
L-Tartrate
Inhibitor in ACP that also Inhibit lysosomal ACP
L-Tartrate
Inhibitor in ACP that inhibits RBC ACP/TRAP
2% formaldehyde, cupric sulfate solution
Method of Analysis ACP
Storage
Frozen
Acidified
ACP is acidified in what pH
< pH of 6.5
When ACP is acidified, ACP is stable for ______ @ ________
2 days
Room temp
Methods of Analysis ACP
Variables
Room temp
Bilirubin
Hemolysis
Heparin, Oxalate, Fluorides
VARIABLES (ACP)
Room temp
-ACP stable within ________ @ RT because CO2 from
blood sample is released which (increases/decreases) pH of the sample
1-2 hrs
increases
false (decrease/increase) due TRAP/RBC ACP
Bilirubin - decrease
Hemolysis - increase
Heparin, Oxalate, Fluorides -
false (decrease/increase)
decrease
in measuring prostatic ALP what is the inhibitor
(L. tartrate -
Enumerate the steps in measurign prostatic ACP
- Measure total ACP
- Add L-tartrate
- Measure total ACP (TRAP)
○ Prostatic ACP = Total ACP - TR
Enumerate the different kind of ACP methods
- Gutman and Gutan
- Shinowara
- Babson, Read & Philips
- Roy and Hillman
GUTMAN AND GUTAN
Substrate
Phenyl phosphate
GUTMAN AND GUTAN
End product
Inorganic phosphate
SHINOWARA
Substrate
P-nitrophenylphosphate
SHINOWARA
End Product
p-nitrophenol
Babson, Read & Philips
Substrate
α-naphthylphosphate
BABSON, READ & PHILIPS
End product
α-naphthol
ROY AND HILLMAN
Substrate
Thymolphthalein monophosphate
ROY AND HILLMAN
End product
Free thymolphthalein
