How to perform Erythrocyte Sedimentation Rate (ESR)

Introduction

Erythrocyte sedimentation rate (ESR) is the rate at which the red blood cells settle in a tube over a given amount of time. Erythrocytes are red blood cells. The sedimentation rate is the time it takes for these red blood cells to settle at the bottom of a test tube. An Erythrocyte sedimentation test can help diagnose inflammation in body. Inflammation is simply injury, infection, and many other conditions, including immune system disorders, certain cancers, and blood disorders.

If anti-coagulated blood is left to stand undisturbed in a vertical tube, the red cells will gradually settle to the bottom of the tube leaving a clear layer of plasma. Red blood cells have a negative charge and when suspended in normal plasma, rouleaux formation is minimal and the sedimentation of the cells is slow. Changes in the proportion and concentration of plasma proteins, particularly macro-molecules like fibrinogen and globulin, reduce the negative charge and thus increase the rate of rouleaux formation. This in turn increases the rate of sedimentation.

The ESR test is a non-specific test that indicates changes in plasma protein concentrations due to infection or injury.

Factors Affecting Erythrocyte sedimentation rate

1. Plasma Protein

Changes in plasma protein happen rapidly following tissue injury or in response to inflammation. Increased concentration of fibrinogen and immunoglobulins will increase rouleaux formation and hence the rate of sedimentation. Plasma albumin retards sedimentation of RBCs.

2. RBC Size and Number

The size and number of RBCs that show alterations in their biconcavity, such as spherocytes and sickle cells, usually do not increase the ESR rate, unless there is severe anemia. Increased cell mass will decrease the sedimentation rate (e.g. polycythemia).

Technical Factors

Tilted ESR tube. ESR tubes should always be in a perpendicular (straight up) position. Slight deviations from the absolute vertical can increase results.
A temperature higher than room temperature (18- 25 º C) accelerates sedimentation. Therefore, ESR tubes should not be exposed to direct sunlight or anything that may alter their temperature. If the test is to be carried out at a higher temperature, a normal range should be established for that temperature.
Vibration can reduce the rate of erythrocyte sedimentation.

Methods of Measuring Erythrocyte sedimentation rate

There are two methods for ESR measurement:

The Westergren method
The Wintrobe method. The

Recommended method is the Westergren Method.

Westergren Method

Principle

The test measures the sedimentation of red cells (in diluted blood) after one hour in an open-ended tube mounted vertically on a stand. The recommended tube is a straight glass or rigid plastic transparent tube 30 cm in length and not less than 2.55 mm in diameter. The bore must be uniform to within 5% throughout (0.05 mm).

Procedure

Venous blood is collected into a sodium citrate tube, or collected into an EDTA tube and diluted accurately in a proportion of 1 volume of citrate to 4 volumes of blood. The usual practice is to collect blood directly into the sodium citrate tube.
Mix the blood sample thoroughly and then draw it up into the Westegren tube to the 200 mm mark by means of a mechanical suction device.
Place the tube exactly vertical and leave undisturbed for exactly 60 minutes, free from vibrations and not exposed to direct sunlight
After the 60 min are over, read to the nearest 1 mm the height of the clear plasma above the upper limit of the column of sedimenting cells.
The result is expressed as ESR= X mm in 1 h.

Notes on procedure:

The test should be carried out within 4 hours of collecting the blood. A delay up to 6 hours is permissible provided that the blood is kept at 4◦C.
EDTA blood can be used 24 hours on a specimen that was kept in 4◦C if 1 volume of 109 mmol/l trisodium citrate is added to 4 volumes of blood immediately before the test is performed.