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Plasma chemical composition
It consists of 90 % water and 10%
1. Proteins ≈ 7 %
2. Inorganic salts ≈ 0.9%
3. The reminder consisting
compounds other than
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1. Organic (Nonprotein)
Constituents of Human Blood
Amino acids
Organic acids
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2. Inorganic Constituents of
Human Blood Plasma
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3. Composition of plasma
Fibrinogen Transthyrein
α1 Globulins
α2 Globulins
β Globulins
Serum proteins
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γ Globulins
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Serum is blood plasma without fibrinogen
and other clotting factors.
The serum proteins can be separated by:
1. Salting out.
2. Tiselius electrophoresis.
3. Zone electrophoresis.
4. Immunoelectrophoresis.
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1. Salting out
 Salting out is a method of separating proteins
based on the principle that proteins are less
soluble at high salt concentrations.
 The salt concentration needed for the protein to
precipitate out of the solution differs from
protein to protein.
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 Precipitation, separation, or coagulation of a
protein from its solution by saturation or partial
saturation with a neutral salt such as sodium
chloride or ammonium sulfate.
 The separation of protein fractions in the serum
or plasma by precipitation in increasing
concentrations of neutral salts.
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 Proteins are large molecules composed of covalently
linked amino acids.
 Depending on electron distributions resulting from
covalent or ionic bonding of structural subgroups,
proteins have different electrical charges at a given pH.
 serum proteins have been fractionated on the basis of
their electrical charge to separate the serum protein
components into five classifications by size and
electrical charge.
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Tiselius electrophoresis
 Moving
electrophoresis is
electrophoresis in a free
 The principle is the motion
through a stationary liquid
under the influence of an
electric field.
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59% 8% 5%12%
 The apparatus includes a U-shaped cell filled
with buffer solution and electrodes immersed at
its ends.
 The sample applied could be any mixture of
charged components like a serum. On applying
voltage, the compounds will migrate to the anode
or cathode depending on their charges.
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Zone electrophoresis.
The Serum Protein Electrophoresis procedure is
intended for the separation and quantitation of serum
proteins using cellulose acetate electrophoresis.
A drop of serum is applied in a band to a thin sheet of
supporting material, like paper, that has been soaked in
a slightly-alkaline salt solution .
At pH 8.6, which is commonly used, all the proteins
are negatively charged, but some more strongly than
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 A direct current can flow through the paper
because of the conductivity of the buffer with
which it is moistened .
 The serum proteins move toward the positive
electrode .
 The stronger the negative charge on a protein,
the faster it migrates .
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 After a time (typically 20 min),
the current is turned off and the
proteins stained to make them
 The separated proteins appear
as distinct bands .
 The most prominent of these
and the one that moves closest
to the positive electrode is
serum albumin .
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 Serum immunoelectrophoresis is a
immunoglobulins in the blood
using agar gel.
 Immunoglobulins are proteins
that function as antibodies.
 There are various types of
immunoglobulins. Some can be
 If you do have these abnormal
proteins, this test can also help
identify their specific type.
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 γ Globulins (IgA)
Multiple Myloma
 Albumine + α1,2 Globulins
 Albumin + γ Globulins
 β Globulins + γ Globulins
Cirrhosis of liver
Chronic rheumatoid arthritis
 α1 Globulins + Albumin+ γ Globulins Hodgkin’s
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 The amount in normal plasma is10-40 mg/dl
 Function: Binding and transport of thyroxin
and retinol binding protein
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 Serum albumin is the most abundant blood
plasma protein.
 The amount in normal plasma is 35004500mg/dl
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Structure and properties of albumin
 Human serum albumin is a single peptide chain
of 585 amino acids, held in three homologous
domains by 17 disulfide bonds.
 The S-S bonds provide stability while the
intervening peptide strands allow for flexibility.
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 It is characterized by its solubility in water
 Albumin is one of the few plasma proteins that is
not a glycoprotein.
 It has the lowest molecular weight of almost of
plasma proteins
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Synthesized of albumin
 Albumin is synthesized in the liver as
prealbumin which has an N-terminal peptide that
is removed before the nascent protein is released
from the rough endoplasmic reticulum.
 The product, proalbumin, is in turn cleaved in
the Golgi vesicles to produce the secreted
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Function of Albumin
1.Role of albumin in osmotic pressure
A major function of albumin is its role in
osmotic regulation.
It gives a much greater osmotic effect at
the Ph 7.4 of blood
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 Albumin is responsible for about 75- 80 % of
the osmotic effect of plasma because:
1.It constitutes slightly> half the plasma proteins
by weight
1.It has the lower molecular weight of the major
plasma proteins.
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Mechanism of osmotic pressure
 At pH 7.4 albumin
 These
concentration of plasma
the osmotic
movement of water between
plasma and extra vascular fluid.
 Changes in plasma protein concentration and
impaired water balance
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2. Transport function of albumin
 About 40% of albumin is contained in the
 The reminder being in the extra vascular space of
tissue ( skin, muscle & intestine)
 Albumin transports
 Many drugs like sulfonamides, penicillin G and
aspirin also bind tightly to albumin.
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α Globulins
α1 globulins
• α1 Acid glycoprotein
• Retinol binding
• α1 Fetoglbuline
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α2 globulins
• Ceruloplasmin
• Haptoglobin
Type 2-1
Type 2-2
α1 globulins
α1 Acid glycoprotein
 Unknown function.
 It has extraordinarily high carbohydrate
 There are intrachain disulfide bond.
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α1 globulins
α1 Fetoglobulin
 A glycoprotein, is the major protein in the human fetal plasma and
amniotic fluid.( the same function of albumin in adult)
 It is very low amounts in adults.
 After birth its conc. decrease with the increase in albumin.
 The sequences of fetoglobulin and albumin are homologous.
 Fetoglobulin levels are elevated in patients with hepatoma and
during pregnancy.
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α1 globulins
Retinol binding protein
 It serves in retinol transport.
 Its presence in such a complex could prevent
renal excretion of the small retinol binding
protein in the urine.
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α2 globulins
 It is MW about 151000
 It is an enzyme synthesized in the liver containing 8 atoms
of copper in its structure.
 Ceruloplasmin carries 90% of the copper in our plasma.
The other 10% is carried by albumin.
 Ceruloplasmin exhibits a copper-dependent oxidase
activity, which is associated with possible oxidation of Fe2+
(ferrous iron) into Fe3+ (ferric iron), therefore assisting in
its transport in the plasma in association with transferrin,
which can only carry iron in the ferric state.
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 Wilson’s disease:
1. It is rare inherited disease
2. Plasma ceruloplasmin is markedly reduced
and Cu levels increase in brain and liver
with resultant neurological changes and
liver damage.
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α2 globulins
 Hp constitute about ¼ of the α2 globulins
 It is a glycoprotein, produced by the liver
 They form specific, stable 1:1 molecular complexes with
hemoglobin, Such complexes form in vivo as the result of
intravascular hemolysis of erythrocytes
 Because of their high molecular , the complexes can’t be
excreted by the kidney; this prevent excretion of iron in the
urine and at the same time protects the kidney from
damage by hemoglobin .
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 The haptoglobin- hemoglobin complexes are
degraded by reticuloendothleial cells, and iron is
reutilizes for heme synthesis after degradation of
globins and excretion of degraded heme as bile
 Patients with a
have haptoglobin
 There three genetic types of haptoglobulin designated
Hp 1-1(MW about 100000), Hp 2-1 (MW about 9000),
and Hp 2-2 (MW about 42600), differs in the structure
according to disulfide bonds (They are distinguished
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β Globulins
1- Hemopexin
 It binds heme and prevents its urinary excretion,
thereby retaining heme iron for further use.
 Neither hemoglobin, cytochrome c, nor bilirubin binds
to hemopexin.
 The hemopexin isolated from normal individuals does
not have a full complement of heme, but when isolated
from patients with hemolytic anemia it is almost fully
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 2- Transferrin
1. It is the major component of β Globulins.
2. It makes up 3% of total plasma proteins
3. The main function to transport Fe ion to tissue where
it is required like bone marrow
4. Transferrin may also transport Cu & Zn
5. Also, regulates the concentration of free iron in
6. The concentration of transferrin increases during
pregnancy and iron deficiency
7. Free iron is toxic, but binding of Fe to transferrin
reduces iron toxicity and transports iron to tissues
where it is required
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 3- C reactive protein
 It is normally present at a concentration of < 1mg/dl
in adults but increases markedly after acute
 The function of these protein is unknown, but it has
been suggested that it promotes phagocytosis.
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 4- β2 microglobulin
 It is present in very small amount in plasma
because of it is low molecular weight .
 It is excreted in the urine where it is found
normally in a concentration of about 0.1mg/l
 It is the smaller subunit of the HLA
histocompatipility antigen complex, which regulates
rejection of transplanted tissue.
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γ Globulins
 This fraction of
immunoglobulins .
 They are often found during inflammatory
illness e.g. rheumatoid arthritis and multiple
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 Classes of conjugated proteins consisting of a
protein combined with a lipid.
 The normal functioning of higher organisms
requires movement of insoluble lipids, such as
triglycerides, between tissues.
 To accomplish this movement, lipids are
incorporated into macromolecular complexes
called lipoproteins.
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Function of lipoprotein
 Lipoproteins serve as transport vehicles for fatty
acids and cholesterol in the blood and lymph.
A combination of a lipid and protein
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Classification of lipoprotein
 They are classified according to their density as highdensity lipoproteins, low-density lipoproteins, and verylow-density lipoproteins.
 Health care workers are interested in the concentration
of the different types of lipoproteins in the blood
because it has implications for health:
– A high concentration of low-density lipoproteins appears to
present a health risk and is associated with a high incidence of
heart disease.
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Classification by density
 General categories of lipoproteins, listed in order
from larger and less dense (more fat than protein)
to smaller and denser (more protein, less fat):
1. Chylomicrons - carry triacylglycerol (fat) from the
intestines to the liver, skeletal muscle, and to
adipose tissue.
2. Very low density lipoproteins (VLDL) - carry (newly
synthesized) triacylglycerol from the liver to
adipose tissue.
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3. Intermediate density lipoproteins (IDL) - are
intermediate between VLDL and LDL. They are
not usually detectable in the blood.
4. Low density lipoproteins (LDL) - carry cholesterol
from the liver to cells of the body. Sometimes
referred to as the "bad cholesterol" lipoprotein.
5. High density lipoproteins (HDL) - collects
cholesterol from the body's tissues, and brings it
back to the liver. Sometimes referred to as the
"good cholesterol" lipoprotein.
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Classification according to
alpha and beta
 It is also possible to classify lipoproteins as
"alpha" and "beta", akin to the classification
of proteins in serum protein electrophoresis .
Moving to cathode (lipoprotein have more
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Serum lipids (plasma lipoproteins)
 Because of their relationship to cardiovascular disease, the
analysis of serum lipids has become an important health
Typical values mg/dl
Desirable mg/dl
Cholesterol (total)
LDL cholesterol
< 100
HDL cholesterol
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Total cholesterol is the sum of
 HDL cholesterol
 LDL cholesterol and
 20% of the triglyceride value
 Note that
 high LDL values are bad, but
 high HDL values are good.
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Plasma Enzymes
 Enzymes are biological catalysts responsible
for supporting almost all of the chemical
reactions in the body.
 Enzymes are found in all tissues and fluids
of the body.
 Intracellular enzymes catalyze the reactions
of metabolic pathways.
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 Plasma membrane enzymes regulate
catalysis within cells in response to
extracellular signals, and enzymes of the
circulatory system are responsible for
regulating the clotting of blood.
 Most plasma enzymes don’t have metabolic
roles in plasma, except for the enzymes
concerned in blood coagulation.
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1- Lipoprotein lipase
 It
an enzyme that hydrolyzes lipids in
lipoproteins ,like those found in chylomicrons and
very low-density lipoproteins (VLDL), into three
free fatty acids and one glycerol molecule.
 Lipoprotein
endothelial cells lining the capillaries
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Lipoprotein lipase deficiency
 Severe mutations that cause LPL deficiency result
in type I hyperlipoproteinemia, while less extreme
mutations in LPL are linked to many disorders of
lipoprotein metabolism.
 High-fat diets have been shown to cause tissuespecific over expression of LPL: This has been
implicated in tissue-specific insulin resistance and
consequent development of type 2 diabetes
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 It is an enzyme that catalyzes the hydrolysis
of the neurotransmitter acetylcholine into
choline and acetic acid,
 A reaction necessary to allow a cholinergic
neuron to return to its resting state after
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There are two types of
 1- Acetyl cholinesterase, also known as RBC
cholinesterase, found primarily in the blood and
neural synapses
 2- Pseudocholinesterase, also known as plasma
cholinesterase, found primarily in the liver.
The difference between the two types of
cholinesterase has to do with their respective
preferences for substrates: RBC cholinesterase
hydrolyses acetylcholine more quickly
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3. Acid phosphatase
 Acid phosphatase is a phosphatase, a type
of enzyme, used to free attached phosphate
groups from other molecules during
 Different forms of acid phosphatase are
found in different organs, and their serum
levels are used as a diagnostic for disease
in the corresponding organs.
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 For example, elevated prostatic acid
phosphatase levels may indicate the
presence of prostate cancer.
 The optimal pH for the enzyme activity is
below 7
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4. Alkaline phosphatase
 (ALP) is a hydrolase enzyme responsible for
removing phosphate groups from many types of
molecules, including nucleotides, proteins, and
 The process of removing the phosphate group is
called dephosphorylation.
 As the name suggests, alkaline phosphatases are
most effective in an alkaline environment.
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Physiology of alkaline phosphatase
 In humans, alkaline phosphatase is present
in all tissues throughout the entire body, but
is particularly concentrated in liver, bile duct,
kidney, bone, and the placenta.
 The optimal pH for the enzyme activity is10
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Diagnostic use of alkaline phosphatase
 Concentrations blood serum levels of ALP
are typically 39-117 Units per liter in adults
 Levels are significantly higher in children
and pregnant women.
 Lowered levels of ALP are less common
than elevated levels.
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 The following conditions
abnormal levels of ALP:
 Elevated levels (hyperphosphatasemia)
 Lowered levels (hypophosphatasemia)
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 Isoenzymes )are enzymes that differ in
amino acid sequence but catalyze the same
chemical reaction).
 In many cases, they are coded for by
homologous genes that have diverged over
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5. Lactate dehydrogenase
 LDH is found in many cells, but especially in
muscle cells.
 It catalyses interconversions of pyruvic acid and
lactic acid.
 The level of LDH is commonly used in exercise
physiology as a measure of the capacity of
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There are different forms of LDH
 One form called heart-specific LDH (H-LDH)
preferentially catalyses lactate oxidation to
pyruvate and predominates in slow-twitch
muscle fibers.
 Another form, muscle-specific LDH (MLDH), preferentially catalyses the reduction
of pyruvate to lactate and predominates in
fast-twitch muscle fibers.
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 LDH-1 (4H) - in the heart
 LDH-2 (3H1M) - in the reticuloendothelial
 LDH-3 (2H2M) - in the lungs
 LDH-4 (1H3M) - in the kidneys
 LDH-5 (4M) - in the liver and striated muscle
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6. Transaminase or
an aminotransferase
 It is an enzyme that catalyzes a type of
reaction between an amino acid and an αketo acid .
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I. Aspartate transaminase
Aspartate transaminase
called serum glutamic
transaminase (SGOT)
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(AST) also
Function of AST(SGOT)
 It facilitates the conversion of
Aspartate + α –ketoglutarate
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 Isozymes
 Two isoenzymes are present in humans. They
have high similarity.
 GOT1 ,the cytosolic isoenzyme derives mainly
from red blood cells and heart
 GOT2 ,the mitochondrial
predominantly present in liver .
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Clinical significance of AST
It is raised in acute liver damage.
 It is also present in
1. red blood cells,
2. cardiac muscle,
3. skeletal muscle,
4. Kidney,
5. and brain tissue
It may be elevated due to damage to those sources
as well.
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II.ALT or Alanine transaminase
 ALT is found in serum and in various bodily
 but is most commonly associated with the
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Function of ALT
 It catalyzes the transfer of an amino group from
alanine to a-ketoglutarate ,the products of this
reversible transamination reaction
pyruvate and glutamate.
Alanine + α –ketoglutarate
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glutamate +pyruvate
Clinical significance of ALT
 It is commonly measured clinically as a part
of a diagnostic liver function test ,to
determine liver health.
 It is also called serum glutamate pyruvate
transaminase( SGPT
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Elevated levels of ALT
Elevated levels of ALT often suggest the existence
of other medical problems such as
1. alcoholic or viral hepatitis ,
2. congestive heart failure,
3. liver damage ,
4. biliary duct problems,
5. infectious mononucleosis ,
6. or myopathy .
 For this reason ,ALT is commonly used as a way
of screening for liver problems. However,
elevated levels of ALT do not automatically mean
that medical problems exist.
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7. Creatine kinase (CK),
 Creatine kinase (CK), also known as
phosphocreatine kinase,
 It is an enzyme expressed by various tissue
 It catalyses the conversion of
Adenosine diphosphate (ADP)
triphosphate (ATP)
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 Creatine kinase is an
enzyme in such tissues.
 Clinically, creatine kinase is assayed
in blood tests as a marker of
1. (heart attack)
2. (severe muscle breakdown),
3. acute renal failure.
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Types of CK
 In most of the cell, the CK enzyme consists of two
subunits, which can be either B (brain type) or M (muscle
 There are, therefore, three different isoenzymes: CK-MM,
CK-BB and CK-MB.
 The genes for these subunits are located on different
chromosomes: B on 14q32 and M on 19q13.
 In addition to those, there are two mitochondrial creatine
kinases, the ubiquitous and sarcomeric form.
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Laboratory testing of CK
 CK is often
 In addition, it is determined
specifically in patients with
chest pain and acute renal
failure is suspected.
 Normal values are usually
between 25 and 200 U/L.
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Crystals of creatine kinase
 Elevation of CK is an indication of damage to
 Lowered CK can be an indication of
alcoholic liver disease and rheumatoid
 Isoenzyme determination has been used
extensively as an indication for myocardial
damage in heart attacks.
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