Chem. D. Mays L3 Measurement of Serum Lipids Profile

Measurement of serum Lipids profile
(Lipids Panel or Coronary Risk Panel)

The lipids profile tests
The lipids profile includes
1. Total cholesterol (TChol),
2. High density lipoprotein (HDL-cholesterol) ,good cholesterol),
3. Low density lipoprotein (LDL-cholesterol (bad cholesterol),
4. Triglycerides (TGs).
5. Sometimes the report will include additional calculated
values called atherogenic indexes such as
 LDL/ HDL
 TChol / HDL
 TG / HDL
 Log (TG / HDL)

Mechanisms that control lipids in body
There are complex mechanisms that control lipids on

two ways:
1. the release of lipids from the tissues into plasma,
2. the uptake of lipids by the tissues from plasma.
Abnormalities of these mechanisms may be associated
with the development of disease.
Measurement of the serum lipids profile (lipids and

lipoproteins) is useful in characterizing an
individual’s risk of developing cardiovascular
diseases (CVD) and in monitoring therapeutic
interventions.
Total Cholesterol

 Cholesterol is fat-like substances that can
synthesis in the body (mainly by the liver and
intestinal mucosa) and comes from the diet.
 Cholesterol plays an important role in making some
hormones and is a part of the cell membrane.
 too much cholesterol can lead to fatty deposits
inside the blood vessels. These deposits can further
lead to plaques and narrowing or complete
 Too much cholesterol (hypercholesterolemia )can
lead to fatty deposits inside the blood vessels, which
can cause cardiovascular problems. Fortunately,
high cholesterol can treat with both diet and
medications.
 Very low cholesterol values( hypocholesterolemia ),
as is sometimes seen in critically ill patients (e.g.
septicemia and cancer), can be as life threatening as
very high levels.
 Total serum cholesterol (TChol)measured for
many years to quantify the total amount of
lipoproteins.
Normal value
Determination Serum Total Cholesterol:
The shortly addition steps of serum TChol
estimation
Each tube shake well and allowed stand for 5 min at 37°
C or 10 min at room temperature.
The absorbencies read at 500 nm against reagent blank.
The color intensity was stable for 1 hr.

:Calculation
1. Triglycerides (TGs)
Triglycerides are a combination of three fatty acids and one glycerol molecule.
They are necessary to provide energy for various. Excess triglycerides are stored
in adipose tissue, and the fatty acids provide the raw materials needed for
conversion to glucose (gluconeogenesis) or for direct use as an energy source in
cases of starvation, insulin resistance, and hypoglycemia.
Although fatty acids originate in the diet, many had also derived from unused
glucose and amino acids that the liver converts into stored energy. Triglyceride
levels vary by age, sex, weight, and race: Levels increase with age. Levels are
higher in men than in women (among women, those who take oral contraceptives
have levels that are 20 to 40 mg/dL higher compared to those who do not). Levels
are higher in overweight and obese populations compared to those with normal
weight.
High density lipoproteins(HDL):
1. The smallest of the lipoprotein particles and the densest.
2. They contain :
* a large amount of protein,
* approximately equal amounts of cholesterol and
phospholipid,
* very little triglyceride.
3. Function of HDL is to remove cholesterol from cells to the liver
through activation of the plasma enzyme, lecithin-cholesterol
acyltransferase (LCAT), for hepatic excretion. This process is
through to be anti-atherogenic, so circulating HDL reduce the
risk of coronary heart disease (CHD) also HDL plays an
important role as an anti-inflammatory,
:Normal value
Determination of Serum HDL:
Principle:
 Before measurement serum HDL, the serum sample was treated
with precipitating agents (Phosphotungstic acid and magnesium
chloride or dextran and MgSO4) to precipitate other lipoproteins
(LDL, VLDL and CMs) from the sample.
 The remaining HDL hydrolyzed by PEG-CE (use a
working reagent of total cholesterol estimation
procedure) to make cholesterol and fatty acids.
Cholesterol reacts with PEG-CO to produce cholest-4-
en-3-one and peroxide (H2O2). The peroxidase reaction
results in the production of a purple colored product
that has an absorbance maximum at 500 nm.
HDLconcentration is directly proportional to the
absorbance maximum in this end-point reaction. A
sample blank also monitored to ensure no extraneous
reactions interfere with the calculations of HDL levels.
Procedure of Serum HDL estimation:
 Precipitation step: in clean test tube, 25µL of precipitating regent

added to 500 µL of serum sample, then it mixed vigorously and
allowed stand for 10 min at room temperature. Finally, it
centrifuged for 10 min at 4000 rpm.
 The supernatant separated and applied the next addition steps as in
next table by using the working reagent of total cholesterol (mono-
reagent) estimation procedure.
 The shortly addition steps of serum HDL estimation as in the next
table :
4. Each tube mixed well and allowed stand for 5 min at 37° C or 10
min at room temperature.
5. The absorbencies read at 500 nm against reagent blank. The color
was stable for 30 min, protected from light.
Very low density lipoprotein (VLDL):
1. This lipoprotein originates in the liver.

2. It contains
high concentration of TGs
moderate concentration of both cholesterol and phospholipids,
3. its function is to carry endogenously synthesized cholesterol and
triglycerides from the liver to other tissues.
4. The half-live of VLDL in plasma is about 1 hour or less.
5. After the loss of triglyceride converts some VLDL to VLDL
remnants (also called intermediate density lipoprotein, IDL).

Determination of Serum VLDL:
The concentration of Serum VLDL calculated according to the

following formula:
Low density lipoproteins (LDLs):
1. Low-density lipoproteins (LDLs) are proteins that are cholesterol
rich.
2. LDL derived from IDL by the removal of almost all the
triglycerides.
3. LDLs carry cholesterol into the tissues where it can deposit.
4. High LDL levels are a risk factor for cardiovascular disease
because the increased amount of LDL protein increases the body’s
ability to carry more cholesterol into the bloodstream.
Calculating the risk ratio of low-density
lipoprotein to high-density lipoprotein
(LDL/HDL ratio)
is helpful in determining the amount of “bad
cholesterol” in relation to the level of “good
cholesterol.”
•A high level of HDL may partially offset the
negative effects of a high LDL level
• Individuals with ratios of 3 or less have only half
the risk of heart disease .
Notes
The appearance of plasma or serum in the laboratory may provide the
first that a patient has hyperlipidemia as below
1. In healthy individuals:
In the fasting state, plasma or serum are clear.
Following meals, it often becomes specular due to light- scattering
properties of Chylomicrons and VLDL.
2. In cases with TGs concentration 4 mmol /L, plasma or serum
become turbid.
3. With severe hypertriglyceridaemia , the plasma or serum appear
milky(lipaemic).
4. Total cholesterol concentration reflects to LDL concentration
because 70 percent TChol is corporated in LDL and only 20 percent in
HDL concentration and the plasma TGs reflect to VLDL
concentrations.
5. Plasma from a fasting subject contains only LDL ,VLDL, and
HDL.
If plasma is left unseparated more than 2 hr, Chylomicrons float to the
surface, leaving a clear infranatant layer; VLDL remain in
suspension .LDL do not scatter light and even at high plasma
cholesterol the plasma remains clear.
6. Hypercholesterolemia, when due to an increase in LDL is an
important risk factor for coronary heart disease.
7. Hypertriglyceridaemia is a less important risk factor for coronary
heart diseases but when very sever, can cause pancreatitis.

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Measurement of serum Lipids profile

(Lipids Panel or Coronary Risk Panel)

There are complex mechanisms that control lipids on

Measurement of the serum lipids profile (lipids and

 Precipitation step: in clean test tube, 25µL of precipitating regent

1. This lipoprotein originates in the liver.

The concentration of Serum VLDL calculated according to the

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