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Horse blood, defibrinated SR0050
Horse blood, oxalated SR0049
Horse blood, haemolysed SR0048
Sheep blood, defibrinated SR0051
Horse serum SR0035

Horse and sheep blood are the most widely used animal blood products in culture media. The choice of animal is largely traditional, with the USA and much of continental Europe preferring sheep blood, whilst the UK and Commonwealth partners prefer horse blood.

The haemolytic reactions of horse and sheep blood are not identical and blood agar media designed for horse blood may not be satisfactory with sheep blood and vice versa. See Blood Agar Base (Sheep) CM0854.

It is not possible to sterilise whole blood products and therefore they must be collected aseptically. Freshly drawn blood from healthy animals can destroy small numbers of bacteria which may transiently infect it during collection but considerable expertise and skill are required to maintain high quality, sterile blood. It is difficult to obtain sterile blood from abattoirs therefore bleeding is carried out in suitable premises from animals which are specially maintained on high protein diets.

Defibrination is now accepted as the best method of preventing blood clotting. It must be carried out immediately after the drawing the blood and the agitation must be sufficient to denature the fibrinogen but not to cause rupture of the erythrocytes and haemolysis. Some lysis is inevitable in this process but the release of NAD in horse blood from the ruptured erythrocytes stimulates the growth of Haemophilus influenzae. This effect is not seen with sheep blood.

The use of potassium oxalate as an anticoagulant increases the yield of whole blood but it remains as a potential chelate of divalent metals. This chelate effect, which also applies to sodium citrate-treated blood, can be seen in the overall smaller colonies which grow on anticoagulant-containing blood agar.

To prolong the shelf life of blood products in the user's hands, blood is collected, bottled and dispatched on the same day. The physical characteristics of appearance, haemoglobin content and packed cell volume are checked before release but sterility tests are carried out beyond the time of dispatch. Should samples of individual animal blood collections show infection then all recipients of the suspect blood are notified by telephone, fax or telex.

Haemolysed or laked horse blood is used for special purposes in culture media. It has been used for many years in Corynebacterium diphtheriae media, where better growth was observed after lysis of the horse blood by the tellurite in the medium.

Lysed blood is stimulatory for many Clostridia and Haemophilus species but Fildes Peptic Digest of Blood SR0046 can also be used for this purpose.

In antibiotic susceptibility testing, lysed horse blood is added to the medium to improve the reactions with trimethoprim and sulphonamides. Most culture media, unless specially processed for susceptibility testing, contain amounts of thymidine which can antagonise the inhibitory effects of these antimicrobials. When horse blood is lysed the erythrocytes release an enzyme thymidine phosphorylase which converts thymidine into the much less antagonistic compound thymine. This enzyme does not exist in sheep erythrocytes. The lytic agent used for lysed horse blood is white saponin which appears not to affect the growth of bacteria at the level required for lysis.

Serum for Culture Media
Horse serum SR0035

Totally aseptic processing is not required for blood serum production. Blood is collected in clean vessels in a clean environment. After clotting has taken place the blood is stored at 2-8°C to allow the clot to retract and the serum separate. The separated serum is then filter-sterilised and filled out into sterile bottles. All sera should be stored at 2-8°C but for longer term storage it is best frozen at -20°C.

Serum can become cloudy on storage caused by lipase enzymes and lipid changes. Heating the serum to 56°C for 30 minutes (inactivation) will usually overcome this problem.

Sheep Erythrocytes for Immunological Reactions
Sheep blood (in Alsever solution) SR0053

After washing the sheep erythrocytes, by centrifuging with physiological saline until the supernatant saline is free from haemolysis, the washed cells may be used for any immunological/serological test where sheep cell antigens are required e. g. Rose-Waaler, Paul Bunnell and various virological complement fixation tests.

The sheep blood should be stored at 2-8°C and used before the expiry date on the label.

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