ERYTHROCYTE SEDIMENTATION RATE (ESR)

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ERYTHROCYTE SEDIMENTATION RATE (ESR)
ESR is used as an index for presence of an active disease which could be due to many causes.

Principle
When well mixed anticoagulated blood is placed in a vertical tube, the erythrocytes tend to fall towards the bottom of the tube/pipette till they form a packed column in the lower part of the tube in a given time.

Mechanism of ESR
Fall of RBCs depends upon following factors:
i. Rouleaux formation
ii. Concentration of fibrinogen in plasma
iii. Concentration of α and β globulins
iv. Length of the tube
v. Ratio of red cells to plasma
vi. Bore of the tube
vii. Position of the tube

i) Rouleaux formation The erythrocytes sediment in the tube/pipette because their density is greater than that of plasma. When a number of erythrocytes aggregate in the form of rouleaux and settle down, their area is much less than that of the sum of the area of constituent corpuscles. The rouleaux formation is very important factor which increases the ESR.

ii) Concentration of fibrinogen It leads to colloidal changes in plasma which cause increased viscosity of plasma. Concentration of fibrinogen parallels ESR. If concentration of fibrinogen is raised, ESR is increased. In defibrinated blood, ESR is very low.

iii) Concentration of α and β globulins These protein molecules have a greater effect than other proteins in decreasing the negative charge of the RBCs that tends to keep them apart thus promoting rouleaux formation. Albumin retards the ESR; thus conditions where albumin is low ESR is more.

iv) Ratio of red cells to plasma The change in the ratio of RBCs to plasma affects ESR. When plasma is more, ESR will be increased, and vice versa.

v) Length of the tube If length of the tube/pipette is more, RBCs will have to travel a longer distance and thus ESR is low than when length of the tube is short, and vice versa.

vi) Bore of the tube If bore of the tube is more the negative charge which keeps the RBCs apart will be
less and ESR will be more, and vice versa.

vii) Position of the tube If the tube/pipette is not vertical, the RBCs will have to travel less distance and ESR will be more.

Phases in ESR
ESR takes place in the following 3 phases which are carried out in one hour:
  • Phase of rouleaux formation: In the initial period of 10 minutes, the process of rouleaux formation occurs and there is little sedimentation.
  •  Phase of settling: In the next 40 minutes, settling of RBCs occurs at a constant rate.
  •  Phase of packing: In the last 10 minutes sedimentation slows and packing of the RBCs to the bottom occurs. That is why ESR by all methods is expressed as mm first hour rather than per hour.
Methods of ESR
1. Westergren’s method
2. Wintrobe’s method
3. Micro ESR method
4. Automated methods

1. Westergren’s Method

Owing to its simplicity this method used to be the most commonly employed standard method prior to the AIDSera. Westergren’s pipette is a straight pipette 30 cm long open at both ends with internal bore diameter of 2.5 mm and is calibrated from 0-200 mm from top to bottom.
Anticoagulant Trisodium citrate (Na3C6H5O7.2H2O) as 3.8 g/dl liquid anticoagulant is used. It is used in the concentration of 1:4 i.e. four parts of blood are added to one part of anticoagulant.
Procedure
  1. The patient is advised to come in the morning fasting (as heavy protein diet affects concentraton of plasma proteins).
  2. Take 1.6 ml of patient’s blood and mix it with 0.4 ml of citrate anticoagulant already put in a tube. The test should be done within two hours of taking blood.
  3.  Fill the pipette upto mark O with citrated blood with the help of rubber teat by vacuum filling and fix it in anrack vertically away from sun light or vibrations.
  4.  Let it stand for one hour after which reading is taken at the upper meniscus of the RBCs.
Normal values
Males 3-5 mm 1st hour
Females 4-7 mm 1st hour

Advantages
i. It is a more sensitive method.
ii. It is easy to fill and clean the Westergren’s pipette.

Disadvantages
i. Requires more amount of blood.
ii. Dilution of blood in anticoagulant affects ESR.
iii. Filling of blood by mouth pipetting should be strictly discouraged.

2. Wintrobe’s Method
The Wintrobe tube is a glass tube closed at one end. The tube is 110 mm long and has an internal bore
diameter of 2.5 mm. The tube is graduated on both sides : from 0 to 10 on one side and 10 to 0 cm on the other.
Anticoagulants Either of the following 2 anticoagulants can be used:
    i. Ethylene diamine tetraacetic acid (EDTA) solid crystals 1-2 mg/ml.
    ii. Double oxalate (solid) 2-3 mg/ml(ammonium oxalate and sodium or potassium oxalate in the ratio of 3:2; the former causes swelling and the latter causes shrinkage of RBCs and hence RBC shape is
retained).


Procedure
  1.  The patient is called in the morning fasting.
  2.  Draw 1 ml of blood into the anticoagulant.
  3.  Fill the Wintrobe tube upto mark 0 with anticoagulated blood with the help of a Pasteur pipette having a long stem (15 cm) so as to fill the tube free of air bubbles.
  4.  Place the tube vertically in a stand and note the ESR after one hour.
Normal values
Males 0-7 mm 1st hour
Females 0-15 mm 1st hour

Advantages
i. It is simple method and requires small amount of blood.
ii. There is no dilution with anticoagulant.
iii. Packed cell volume (PCV) can also be done by the same tube.
iv. Filling of tube with Pasteur pipette eliminates chance of any infection due to handling of blood.

Disadvantages
i. Because of short column and choice of anticoagulant,
it is not as sensitive index of diseases.
ii. Addition of more anticoagulant can lower ESR.
iii. ESR of more than 100 mm can not be measured.

3. Micro ESR Method

This method is used in pediatric patients or in patients where venepuncture is not possible. In this method a capillary 160 mm long with an internal bore diameter of 1 mm is used. The capillary is graduated 1 mm apart for 50 mm, with two red lines on it. Alternatively, nongraduated
heparinised capillary may be used and the reading is taken by measurement of length of column.
Anticoagulant Mixture of sodium citrate and EDTA is used.
Procedure
  1. Fill the microsedimentation pipette upto first red mark with anticoagulant.
  2.  Fill the pipette with free flowing capillary blood upto second red mark.
  3.  Invert it several times and allow it to stand for one hour in the sedimentation rack. Take the reading and results are given as that for Westergren’s method.
4. Automated ESR Method
Automated closed systems use either blood collected in special evacuated tubes containing citrate or EDTA. It is taken up through a piercable cap and then automatically diluted in the system.

Clinical Significance of ESR
ESR is a non-specific method of evaluating diseases. It is seldom used for diagnostic purpose but its use is limited to monitoring the prognosis of disease process.

Diagnostic Uses
i. Rheumatoid arthritis
ii. Chronic infections
iii. Collagen diseases
iv. Multiple myeloma
v. Macroglobulinaemia

Monitoring Prognosis of Diseases To see the response to treatment in:
i. Tuberculosis
ii. Temporal arteritis
iii. Polymyalgia rheumatica
iv. In patients of Hodgkin’s disease, ESR of < 10 mm 1st hour indicates good prognosis while ESR of
> 60 mm 1st hour indicates poor prognosis. 
sums up the list of conditions causing raised and lowered ESR.

Diseases causing raised ESR 
i. Tuberculosis 
ii. SABE 
iii. Acute myocardial 
iv. Rheumatoid arthritis 
v. Shock 
vi. Anaemias
vii. Liver disease
viii. Multiple myeloma
ix. Pregnancy
x. Ankylosing spondylitis

Diseases causing raised ESR low ESR
i. Polycythaemia
ii. Spherocytosis
iii.Sickle cell anaemia infarction 
iv. Congestive heart failure
v. Newborn infant
vi. Hypofibrinogenaemia








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