Preparation of the patient

Before taking a blood sample, the patient should normally fast for 10 – 12 hours, provided the physiology of the species concerned permits this. Otherwise, faulty results are to be expected, especially for cholesterol, glucose and TLI. In addition, parameters such as á-amylase, ALT, AST, bilirubin, total protein, triglycerides, serum bile acids, urea, leukocytes and calcium can be affected.

In horses, ruminants, New World camels and small mammals, prolonged periods of fasting are not recommended and fasting blood samples are not common. For special tests (e.g. insulin and glucose for the diagnosis of Equine Metabolic Syndrome), horses should not be given any concentrated feed, oats or access to pasture 4 – 6 hours before the blood sample is taken but may continue to eat hay. Ferrets should fast for a maximum of 2 to 4 hours prior to insulinoma diagnosis.

It is advisable to inform the owner about the influence of physical activity or stress on the results of a blood examination. Particularly muscular enzymes such as CK, LDH and AST can show increased levels in serum after physical exertion. Additionally, glucose and lactate can also show elevated serum levels.

Before doing any allergy tests, including feedstuff tests, any administration of corticosteroids should be stopped. To do so, the following withdrawal times are recommended:

– local/topical corticosteroids: 2 – 4 weeks

– oral corticosteroids (e.g. Prednisolone): up to 8 weeks

– depot cortisone preparations (e.g. Voren®): up to 3 months

If these times cannot be observed, false negative results are possible. If there is a positive result, the reaction class must be assessed taking into account the previous administration of cortisone.

Please note that other itch-suppressing medication may also have a negative impact on the allergy test. Our allergy team will be pleased to advise you.

Allergy tests should be performed during the season or at the end of the season and not earlier than one month after the onset of the clinical signs, as the test may be false negative if performed out of season.

Which sample?

Details on the recommended material (blood, serum, plasma) for the requested test can be taken from our test descriptions or the submission form. For labelling the sample, it is also necessary to indicate the sample type.

 

Whole blood samples

EDTA blood (EB)

– For doing a blood count, EDTA blood is the most suitable material in mammals (however,for birds and reptiles it is heparin blood, see below).
– For the serological examination of the blood type, EDTA whole blood is needed as well.
– As the cells in the sample are not stable, EDTA samples for haematological tests should not be older than 48 hours.
– For most PCR analyses and genetic tests, EDTA blood is required.
– To determine certain parameters such as ACTH, CPSE (prostate), normetanephrine/metanephrine ratio, parathyroid hormone-related protein*, taurine or pro-BNP, only EDTA plasma which was immediately centrifuged and cooled can be used to obtain reliable results.

Heparin blood (HB)

– To collect heparin samples, lithium heparin (LiHep) tubes are available.
– For doing a blood count in reptiles and birds, lithium heparin blood should generally be used.
– Since the amount of blood is often very low in small mammals, lithium heparin tubes are particularly suitable, as heparin whole blood can be used to do the blood count and heparin plasma to determine clinical chemistry parameters as well as T4.
– For the PCR, lithium heparin whole blood should only be used under exceptional circumstances, as lithium heparin can inhibit the PCR and might thus lead to false negative results.

Citrate blood (CB)

– To determine the coagulation parameters, only the appropriate citrate tubes should be used. For getting a correct evaluation, their shelf life may not be exceeded. It is also necessary to have an exact mix ratio of 1:10 (1 part citrate + 9 parts blood).
– For correctly performing platelet function tests, citrate whole blood is required.

Sodium fluoride blood (NaFB)

– Sodium fluoride inhibits enzyme activities which lead to a reduction of some parameters. It should be used for the correct determination of glucose and lactate.

– Make sure to observe the fill level when using sodium flouride tubes. Because of the smaller sample volume that is expected in small mammals, it is also possible to submit either serum which was centrifuged soon after sampling and separated (after 30 minutes) or plasma (immediately).

 

Plasma

– Samples are drawn into tubes with anticoagulants (heparin, EDTA, citrate).                                                                                                                                                                                                                                                           – Fill volume: Fill the sample tubes exactly up to the mark. If the quantity is too small or too large, results may be incorrect.
– Can be centrifuged immediately after collection (10 min, 2000 g).
– Remove the supernatant by pipette and transfer it into an uncoated test tube, then indicate the sample materials on the test tube or use the appropriate bar code label (see Labelling).
– Please note: The additives limit the number of analyses!
– Heparin plasma (HP) is needed for many clinical-chemical examinations. HP cannot be used for agglutination tests.
– The collection of EDTA plasma (EP) for clinical-chemical and/or serological parameters should only take place in exceptional cases, as EDTA disturbs through various mechanisms the measurement of individual parameters such as calcium, magnesium and AP. Likewise, potassium cannot be determined when using EDTA plasma, since EDTA is added as K-EDTA.
– Some coagulation parameters can only be analysed using citrate plasma (CP). Performing platelet function tests using centrifuged citrate plasma is not possible.

 

Serum

– Samples are drawn into tubes without anticoagulants.
– Allow to stand for 30 – 60 min.
– Centrifuge for 10 min at 2000 g.
– Remove the supernatant by pipette and transfer it into an uncoated test tube, then label the test tube.
– For the correct determination of individual parameters, only serum should be used (see detailed descriptions provided for the individual parameters).
– Sending non-centrifuged samples should only be done exceptionally (e.g. in case of a very low sample quantity), as the transport might result in cell damage and thus lead to haemolytic serum.

 

An overview of the different tubes can be found in here.

Factors interfering with analysis

Haemolysis

Haemolysis is caused by leakage of intracellular components of the erythrocytes such as phosphate, iron, potassium and especially haemoglobin due to a damage of the cell membrane. Haemoglobin causes a red colouration of serum/plasma which primarily interferes with the photometric testing done in clinical chemistry.

Lipaemia

Lipaemia refers to the milky/turbid discolouration of serum/plasma due to triglycerides. It is mostly caused by diet- or stress-related factors. Lipaemia can also occur as a result of endocrinological diseases like Cushing’s syndrome or hypothyroidism. Lipaemic samples often complicate the measurement of certain clinical parameters, e.g. bilirubin.

Icterus

Icterus is a yellowish discolouration of serum/plasma. Excess amounts of bilirubin, which is the reason for the yellow colouring, are normally caused by a medical condition
and cannot be influenced. Very severe icterus may sporadically affect certain parameters.
The yellow colouration is physiological in horses.

 

Interfering factor Parameter Level
Haemolysis LDH, HBDH, CK, AST, bilirubin, creatinine, PO4, K, Fe, fructosamines elevated
Haemolysis Ca, glucose, Mg lowered
Lipaemia ALT, AST, GLDH, γ-GT, AP, bilirubin, creatinine, haemoglobin elevated
Lipaemia amylase, Na, CI, K lowered

 

Medicine Parameter Level
Penicillin G K elevated
Tetracyclines PO4 elevated
Tetracyclines K lowered
Salicylates CK, AP, glucose, Na, total protein elevated
Salicylates K, Ca lowered
Corticosteroids CK, AP, glucose, Na, total protein elevated
Corticosteroids K, Ca lowered
Phenylbutazone Ca, Na elevated
Barbiturates CK elevated
Halothane
anaesthesia		 CK, PO4 elevated
Glucose infusion glucose elevated
Glucose infusion PO4 lowered

Specific features

Blood counts

–  EDTA or lithium heparin blood
– When collecting the sample, discard the first 0.5 ml of blood, if possible, as they contain an increased amount of coagulation factors, or first obtain a serum sample.
– Let the blood run down slowly on the side of the sample tube.                                                                                                                                                                                                                                                                                    – Pay attention to the fill volume! Preferably fill up to the mark, since an insufficient volume can result in changes in cell morphology. Do not overfill the tube in any case, as the sample might clot.
– After drawing the sample, tilt the test tube carefully several times. Do not shake it.                                                                                                                                                                                                                                                   – When requesting haematological analysis, a blood smear should always be supplied in addition to whole blood.
– Do not store blood smears in the refrigerator and not close to formalin.
– Pack the samples frost-proof in winter; possibly cool them in summer.
– Reliable results can only be obtained from samples not older than 48 hours.

Clinical chemistry of serum or heparin plasma

Prompt centrifugation of the samples will lead to better test results, as it reduces the risk of haemolysis caused by transportation. However, serum should be allowed to stand for a minimum of 30 min to ensure a complete clotting of the sample.
Serum samples can also be shipped frozen; they will then reach the laboratory cooled. Repeated freezing/thawing, though, should absolutely be avoided.

Determination of glucose and lactate

– Requires sodium fluoride blood or sodium oxalate blood or serum which was centrifuged soon after sampling.
– Fill volume: Fill the sample tubes exactly up to the mark. If the quantity is too small or too large, results may be incorrect.

Coagulation parameters

– Determination is carried out using sodium citrate plasma which is obtained from citrate blood with the mix ratio being 1:10 (1 part citrate + 9 parts blood). Centrifugation should take place promptly (< 1 hour) at the practice. When testing for the von Willebrand antigen, it is imperative to do this immediately after collection. For further information, see also section “Which sample?” on this page.
– If commercial citrate-treated tubes are used, the expiry date needs to be checked before collecting the sample. Expired tubes may no longer be used, as skewed results are to be expected. When drawing the sample, special attention must be paid to the exact fill level (marking on the tube).
– If no commercial tubes are available, sodium citrate 3.13% can be drawn into a syringe.
– No heparinised needles or catheters may be used.

Sample collection for bone marrow cytology

– Sample material from the first aspirate should be used to prepare the smears fo cytological examination (avoid contamination with peripheral blood).
– The syringe used for aspiration should be preloaded with an anticoagulant. The aspirate should be placed into an EDTA, lithium heparin or citrate tube at the latest immediately after the puncture and then inverted well to avoid clots.
– To prepare a smear, the aspirate is put in a Petri dish and gently inverted in order to find the bone marrow spicules.
– The spicules are each placed on a slide and carefully spread to form a monolayer.
– The remaining aspirate is then put back into the tube (wetted with the same anticoagulant) and sent in as well.
– In addition, peripheral blood is collected in a tube, a blood smear is prepared and both are also sent to the laboratory.

Sample material

– When requesting several services, it is necessary to submit two samples of the same material if freezing or exclusion of air is required for one of the services and it is also a service provided by a partner laboratory (marked with *). For example, when requesting ionised calcium* (exclusion of air required and also a service by a partner laboratory) and another parameter in serum, two serum samples must always be sent in, as one serum sample is forwarded to the partner laboratory without being opened.