Clinical Biochemistry
Lipid Profile
Serum lipid profile include:
1. Total cholesterol
2. Triglyceride
3. High-density lipoprotein cholesterol (HDL-cholesterol)
4. Low-density lipoprotein cholesterol (LDL-cholesterol)
5. very low-density lipoprotein cholesterol (VLDL-cholesterol)
There are five major classes of lipoproteins :
1- Chylomicrons.
2- Very-low-density lipoproteins (VLDL).
3- Intermediate-density lipoproteins (IDL).
4- Low-density lipoproteins (LDL)
5- High-density lipoproteins (HDL).
Cholesterol levels are fairly constant, but triglyceride levels
fluctuate considerably from day to day and are highest 1- 4 hours
after meals.
Plasma lipoprotein particles contain variable proportions of four
major elements: cholesterol, triglycerides, phospholipids and
specific proteins called apoproteins.
Clinical Significance:
1- Screening and diagnosis of hyperlipidemia:
Types of hyperlipidemia
a- Primary hyperlipidemia : genetic or familial.
1- primary hypercholesterolemia
2- primary hypertriglyceridemia
3- primary combined hyperlipidemia
b- Secondary hyperlipidemia:
Certain conditions, medications or food stuffs may lead to
hypercholesterolemia or hypertriglyceridemia or both.
Most common causes of secondary hyperlipidemia are:
1) Uncontrolled diabetes mellitus.
2) Nephrotic syndrome (nephrosis).
3) Chronic obstructive liver disease.
4) Hypothyroidism.
5) Exogenous factors such as excessive intake of dietary fat,
chronic alcohol consumption, or corticosteroid drugs.
2- Prediction of coronary heart disease
increased levels of Serum total cholesterol and low-density
lipoprotein cholesterol (LDL- cholesterol) is directly related to a
greater incidence of coronary heart disease (CHD).
Elevated plasma triglycerides and very-low-density lipoproteins
(VLDL) is also directly associated with increased risk of CHD.
In contrast, high levels of high-density lipoprotein cholesterol
(HDL- cholesterol) have been found to be a protective factor for
the development of CHD, so that decreased levels constitute a
risk factor.
3- Diagnosis and monitoring treatment of atherosclerotic diseases.
Total Cholesterol:
Almost all of the endogenous cholesterol is synthesized by the liver and
gut.
Cholesterol exists in two forms:
- Esterified (75%)
- Non-esterified (Free): (25%).
• Cholesterol is an unsaturated alcohol of the steroid family of
compounds.
• It is essential for the normal function of all human and animal cells
and is a fundamental element of their cell membranes.
• It is also a precursor of various essential substances such as adrenal
and gonadal steroid hormones, vitamin D and bile acids.
Laboratory measurement of total serum cholesterol
Specimen Collection:
• Use serum or plasma.
• For plasma, use heparin or EDTA as anticoagulant.
• Collect blood from a fasting patient (at least 12 hours), non fasting blood
sample is acceptable if the purpose of the test is screening for
hyperlipidemia.
• Separate serum from blood cells within 2 hours.
• Cholesterol is stable in serum or plasma specimen for 5-7 days at 2-8 °C
Reagents:
Reagent 1: Buffer: phosphate Buffer, chloro-4-phenol ,Triton,
Sodium cholate, preservative .
Reagent 2: enzymes (Cholesterol oxidase , Cholesterol esterase,
peroxidase) and 4-amino-antipyrine .
Reagent 3: Standard Cholesterol 200 mg/ dl
Principle:
Enzymatic colourimetric method
Cholesterol esters cholesterol esterase Cholesterol + Free fatty acids
Cholesterol + O2 Cholesterol oxidase Cholesten 4 one 3 + H2O2
2H2O2 + Phenol + 4-amino-antipyrine Peroxidase quinoneimine (pink) + 4H₂O
The intensity of pink color of the complex (quinoneimine) is directly
proportional to total cholesterol concentration in the specimen. The
absorbance is measured at 500 nm.
Manual procedure:
Pipette into well identified test tubes Blank Standard Test
Working reagent (R1+R2) 1ml 1ml 1ml
Standard 10µl
Specimen 10µl
• Mix well, let stand for 5 minutes at 37 °C or 10 minutes at room
temperature.
• Record absorbance (A) of the blank (B), standard (S) and test (T) at 500
nm.
• Color is stable for 1 hour.
Calculation:
𝐀𝐛𝐬 𝐓𝐞𝐬𝐭 − 𝐀𝐛𝐬 𝐁𝐥𝐚𝐧𝐤
𝐂𝐨𝐧𝐜. 𝐨𝐟 𝐓𝐨𝐭𝐚𝐥 𝐂𝐡𝐨𝐥𝐞𝐬𝐭𝐞𝐫𝐨𝐥 = 𝐗 𝐒𝐭d 𝐂𝐨𝐧𝐜.
𝐀𝐛𝐬 𝐒𝐭𝐝 − 𝐀𝐛𝐬 𝐁𝐥𝐚𝐧𝐤
Abs = absorbance, it is measured by spectrophotometer.
Std = standard.
Cholesterol standard concentration = 200 mg/dl
Expected values:
Values for adults, estimated in relation to the risk of atherosclerotic
diseases:
• Recommended (normal) level < 200 mg/dl .
• Borderline high (low risk for CHD) 200 - 239 mg/dl
• High level (high risk for CHD) ≥ 240mg/dl
CHD = Coronary Heart Disease
