Homologous blood transfusion – Doping analysis

Doping practices involving the administration of the athlete’s own blood (autologous blood transfusion) or blood from another person (homologous blood transfusion) have been known since the early 1970s and were banned by the IOC in 1988 as doping methods.

The goal of this method is to increase the number of red blood cells (erythrocytes) and thereby improve the blood’s oxygen-carrying capacity.

Following the 2004 Olympic Games in Athens, a new method for detecting blood transfusions from third parties was implemented for the first time. Until then, homologous blood transfusions could not be reliably detected.

Detection of homologous blood transfusion

The procedure for detecting a homologous blood transfusion was developed by an Australian group and published as early as late 2003 (see References). The procedure was validated in 2004 in the WADA-accredited laboratories in Sydney, Athens, and Lausanne for use in doping controls.

To minimize transfusion risks, when using donor blood, only transfusions that are compatible with the recipient’s blood in the AB0 system and Rhesus D factor are administered. The detection method is based on the fact that additional antigens are located on the erythrocyte membrane and allow for a distinction between donor and recipient blood.

These additional antigens, for which the human body has generally not yet produced antibodies, are usually not tested for during a transfusion because the risk of complications is minimal.

The most common antigens (besides the known blood group antigens) are:

C, c, E, e (Rh); K, k (KELL); M, N, S, s (MNS); Lu^a, Lu^b (Lutheran); Le^a, Le^b (Lewis); Fy^a, Fy^b (Duffy); Jk^a, Jk^b (Kidd)

Since, due to genetic variations in blood groups and the distribution of these additional antigens, not every person possesses all antigens, the probability is very low that the surface of a foreign donor’s red blood cells will carry exactly the same antigens.

In the developed testing procedure, the blood sample is therefore tested for up to 12 different antigens (see above, bold text). To determine the antigens, a specific antiserum is used for each of these twelve antigens, which contains the (primary) antibodies to the respective antigen. A non-covalent antigen-antibody binding occurs. The subsequent labeling is performed with a non-specific antibody conjugated to a fluorescent dye.

The samples are then analyzed using a flow cytometer. Based on the principle of hydrodynamic focusing, the cells are separated and then guided through a flow cell in a bead-like chain. In the measuring cell, the cells pass through a laser that excites the fluorescently labeled cells. By detecting the fluorescence intensity, forward scatter, and side scatter, unlabeled erythrocytes can be distinguished from labeled erythrocytes as well as from leukocytes.

Possible results of a test for erythrocyte antigens, partial result for a determination, e.g., for antigen C

A blood transfusion is indicated if, during testing for a specific antigen, two populations of red blood cells are detected: unstained and stained. In such a case, the antigen profiles of the donor and recipient blood differ. However, if both have antigen C or neither has antigen C, the blood transfusion cannot be confirmed by antigen C testing. To minimize the probability that donor and recipient blood are incompatible, up to 12 different antigens are therefore tested.

Further information

References:

M. Nelson et al., Proof of homologous blood transfusion through quantification of blood group antigens, 2003, Haematologica, 88(11):1284-1295

M. Nelson et al., Detection of homologous blood transfusion by flow cytometry: a deterrent against blood doping, 2002 Haematologica, 87(8):881-882 

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Further information on the topic of blood doping can be found here: Article on blood doping