Anticoagulated blood is drawn up into a tube of standardised dimensions (these days the Westergren dimensions are recommended) and left in a vertical position for exactly one hour. After that period the point at which the red cells have separated and settled from the plasma is recorded by reading from the scale on the side of the tube. Despite the fact that the test, in its present form, has been in constant use since 1926 the phenomenon of red cell sedimentation is still only partly understood. However three definite phases have been identified in the process.
During the first, or Lag Phase, the red cells form a characteristic rouleaux pattern and sedimentation is generally slow. The rate accelerates in the second period, the Decantation Phase, and slows again in the final Packing Phase as red cell aggregates pile up at the base of the tube. The size of the rouleaux aggregates formed in the Lag Phase is the critical factor affecting the final result of the ESR. The rouleaux itself appears to be influenced mainly by certain plasma proteins including fibrinogen, IgM and alpha2-macroglobulin. Opinions vary as to the accelerating and retarding properties of glycoproteins & albumin. From the above it can be seen that the sedimentation 'rate' of the red cells is not linear. Further, the time taken for each of the three phases will differ between patients. Therefore no attempt should be made to 'estimate' the result at 1 hour by doubling the value at 30 minutes or multiplying by three after 20 minutes.
Factors influencing the ESR
Although the test itself is subject to a number of variable factors affecting the results, it is basically simple to perform and provided those variables are carefully avoided the test is equally suited to the medical laboratory or the doctors' office. The factors that influence ESR results can be listed under two headings. First, those physical changes in blood plasma and cells, which to some degree the test is measuring and secondly those technical variables that must be avoided when performing the test.
Physical changes in Blood
The ESR is a manifestation of the interaction between red cells and plasma, therefore any physical variation in either component of blood will affect the result. Plasma proteins that affect the ESR include the immunoglobulin (IgM) and a number of the acute phase proteins including fibrinogen, C-reactive protein, alpha-1-anti-trypsin, haptoglobin, etc. These acute phase proteins increase with tissue damage (e.g. lupus or rheumatoid arthritis), pregnancy, chronic inflammation and chronic infection. Changes in the red cells that affect ESR include anaemia, which usually increases the rate, and abnormal cell morphologies such as sickling (HbS), or targeting, which slow the rate.
It is vitally important that anyone setting up an ESR test is aware that a number of external factors will affect the result. Correct mixing of the blood sample at venepuncture and immediately before setting up the test is one of the most important factors influencing the ESR result. Other factors of equal importance include the verticality of the measuring tube (Dispette), the ambient temperature at which the test is performed, vibration and the dimensions of the measuring tube. By using a commercial ESR test kit such as the Dispette 2 system with a properly designed stand, the effect of most of these variables can be eliminated and the test performed with confidence following the detailed instructions supplied with the kit.
Technical advice for performing the test
Although Guest ESR systems are designed to make the test as accurate, yet as simple as possible to perform, there are some general points that need to be kept in mind when setting-up ESR tests:
- The ESR measuring tube (Dispette) must be kept perfectly vertical. The blue Guest ESR stand has a levelling bubble and screw device incorporated to assist, but the level should be checked regularly in case of movement.
- The ESR must be performed at room temperature (defined by the ICSH as 65 - 72°F). The tests must be sited away from windows, direct sunlight and sheltered from draughts. If the ambient temperature of the laboratory or room where the test is being performed is above 22°C, correction can be made using the nomogram published by Manley in 1957. Click here to go to a downloadable copy of the nomogram and instructions on its use.
- Vibration can seriously affect the ESR. Ensure that the bench top is not in contact with machinery (especially centrifuges), or subject to knocks.
- Do not pick up the stand to read results as this will affect other tests in progress. Bring the eye to the level of the top of the red cells to read accurately from the scale.
- Ensure that the result of each test is read at exactly 1 hour from when you set up the tube in the rack. It must be remembered that the cells will go on sedimenting after the first hour and results read at 75 minutes will usually be higher than those read at 60 minutes. A laboratory timer or alarm should be used as a reminder to read the results at exactly the correct time.