Practical coagulation and coagulation monitoring (Proceedings)


Hemostasis is the physiologic process whereby bleeding is halted. The primary functions of the hemostatic system are to: 1) maintain blood in fluid state while in the vessels; 2) arrest bleeding at the site of injury and 3) remove the clot once healing is complete.


  • Provide an overview of functional hemostasis

  • Review the coagulopathies commonly encountered in practice

  • Examine the coagulation tests commonly used in veterinary medicine

  • Provide a framework for in-house testing of the coagulopathic patient

Functional hemostasis

Hemostasis is the physiologic process whereby bleeding is halted. The primary functions of the hemostatic system are to: 1) maintain blood in fluid state while in the vessels; 2) arrest bleeding at the site of injury and 3) remove the clot once healing is complete. This lecture is intended to investigate part 2 of the hemostatic functions. Hemostasis is divided into two subcategories. These are primary hemostasis, which is composed of platelet adhesion, activation and aggregation; and secondary hemostasis, which is coagulation factor activation.

Primary hemostasis is initiated when disruption to the vascular endothelium exposes circulating platelets to collagen in the subendothelial matrix and von Willebrand factor released from disrupted endothelial cells. Adhered platelets degranulate to release a number of vasoactive and attractant factors (thromboxane, additional vWF, histamine, epinephrine, platelet activating factor, etc). This attracts and activates additional platelets, allowing formation of a platelet plug. This plug is unstable and short lived; secondary hemostasis is required to stabilize the plug and form a mature clot.

Secondary hemostasis occurs on the surface of both the endothelium and the platelets, and requires activation of the clotting factors, which are circulating proteins. The end result of secondary hemostasis is cross-linked fibrin strands. Think of the fibrin as a net, holding down a group of platelets. The platelets plug the holes in the net to prevent additional fluid from leaking out, and the net prevents the platelets from floating away. In vitro, the “Y” diagram is used to describe secondary hemostasis. In vivo, the system functions quite differently.

If we look at the “Y” diagram, it appears that the intrinsic and extrinsic systems contribute equally to the activation of the common pathway. In reality, factor VII is integral to activation of secondary hemostasis. Animals deficient in F VII have severe bleeding tendencies and usually die shortly after birth. In contrast, animals (especially cats) with complete F XII deficiency have a normal lifespan and are not coagulopathic. Even the names “primary” and “secondary” hemostasis are misnomers. The platelets and coagulation cascade are activated simultaneously, and each depends on the other for optimal function. So why use the “Y”? It is an easy way to explain laboratory testing for coagulation dysfunction.


Diseases of hemostasis

Disorders of Primary Hemostasis

These can be divided into two broad categories: thrombocytopenia (decreased platelet numbers) and thrombocytopathia (decreased platelet activation/aggregation). 

Thrombocytopenia is caused by decreased production, increased consumption, or destruction. Sequestration can occur (as with snakebite envenomation) but is rare. The most common cause of thrombocytopenia is destruction, whether primary immune-mediated (ITP) or secondary immune-mediated (such as secondary to tick-borne disease, neoplasia, etc.). Decreased production is caused by bone marrow aplasia or secondary to neoplastic processes affecting the bone marrow. Increased consumption is usually associated with DIC. Mild thrombocytopenia can also occur from platelet loss, as occurs with severe hemorrhage.

Thrombocytopathia can occur from inherited defects in platelet activation (such as occurs with greyhounds), decreased vWF, or from drug administration such as aspirin or other NSAIDs. Platelet counts are normal, but platelet function is altered. It is uncommon to see spontaneous bleeding in these patients (unless vWD is of more severe forms), but bleeding may be excessive during surgical procedures or with trauma.

Of these causes, vWD and ITP (either primary or secondary) are the most common. ITP is an immune-mediated destruction of the platelets. Primary ITP, where no underlying cause can be determined, is associated with platelet counts that tend to be < 20,000 cells/?l. Tick borne disease causing a secondary ITP tends to have higher platelet counts, near the 40,000-80,000 cells/?l range. The hallmark sign of severe thrombocytopenia is petechial hemorrhages that tend to occur on mucosal surfaces, including nasal, gingival, gastrointestinal and bladder mucosa. Spontaneous bleeding usually occurs with platelet counts < 20,000 cells/?l.

Multiple forms of vWD exist. In Type I disease, all sizes of vWF multimers are present, but in decreased amounts. This is the most common form, and bleeding is mild to moderate. Breeds affected include the Doberman Pinscher, Basset, Cocker, English Setter, German Shepards, Golden Retrievers, Poodles, Shetland Sheepdogs, Welsh Corgis and many others. In Type II disease, large multimers are absent (which are the most hemostatically active). Bleeding is typically severe, and this form usually affects German shorthair and wirehair pointers. In Type III disease, there is either a complete absence of all sized multimers or only trace amounts. Bleeding is severe. Predisposed breed include the Scottish terrier, Shetland sheepdog, Dutch Kooiker, Chesapeake Bay retriever, and Golden retriever. Unless significant hemorrhage is present, platelet numbers are normal. vWF is often confused with Factor VIII. Remember that Factor VIII levels are normal, and that this factor serves as a carrier to protect vWF from destruction.


Disorders of Secondary Hemostasis

Decreased production of the Vitamin K dependent clotting factors can occur with severe hepatic dysfunction or failure. This can also be seen with anti-coagulant rodenticide (ACR) toxicity. Prolongations of clotting times tend to be much more severe with ACR than with liver failure. There are also a number of inherited disorders of coagulation. Hemophilia A (lack of Factor VIII) and Hemophilia B (lack of Factor IX) occur occasionally in veterinary medicine. Hemophilia A tends to affect large breed dogs (especially German Shepards), and Hemophilia B tends to affect small breed dogs, although any breed can be affected with either disease. Factor VII deficiencies have been reported in the Beagle.

Increased consumption of clotting factors can occur in the late, or hypocoagulable phases of DIC. Loss of factors can occur in severe hemorrhage and subsequent hemodilution with non-clotting factor containing solutions, such as pRBCs, crystalloid, or synthetic colloid solutions. Additionally, synthetic colloid solutions such as hetastarch can cause decreased levels of Factor VIII and platelet dysfunction. Very rarely do these lead to clinical bleeding.

The hallmarks of disorders of secondary hemostasis are ecchymotic hemorrhages and cavitary bleeding. Hemorrhage can occur into any cavity, especially the pleural, pericardial and peritoneal spaces. ACR toxicity in particular can result in bleeding into any part of the body, including pulmonary hemorrhage, and bleeding into the eyes, joints, tracheal membranes, ventricles of the brain, GI tract and bladder.

Practical Coagulation Testing

For all coagulation testing, needle sticks should be as atraumatic as possible. While using a vacutainer has long been considered the standard for drawing blood for coagulation testing, this is not strictly necessary. Samples can be drawn from a needle and syringe, and then rapidly transferred to the appropriate collection tube. Plastic tubes are more likely to cause platelet clumping; therefore, glass tubes should be used if the platelet count is in question. Finally, a recent study published in AJVR showed no significant clinical difference in PT/aPTT drawn through a catheter versus that drawn with a needle stick.

Platelet Counts

If an automated platelet count is available, ensure that no clots are present in the sample. A blood smear should always be used to confirm platelets counts. The feathered edge of the smear should be evaluated for platelet clumping. Clumping can greatly reduce the reported platelet numbers on the CBC. If an automated platelet count is not available, platelet numbers can be estimated from the blood smear provided that clumping is not present. The number of platelets per high power field (100x) can be counted and multiplied by 10,000-15,000. This will give a rough estimate of the platelet count.

Buccal Mucosa Bleed Time (BMBT)

This test can be used to determine if a primary hemostatic disorder is the cause for bleeding. It is inexpensive to perform, although it is strongly recommended that a BMBT kit is used for the test. To perform the BMBT, the test kit should be placed against the gently everted buccal mucosa. The blade from the test kit is deployed, providing a standard depth and length incision. Blotting paper is used to remove blood from under the site, but should not touch the incision itself. Normal BMBT is < 4 minutes in the dog, and <3 minutes in the cat, depending on the test kit. Limitations of the BMBT are considerable inter- and intra-operator error, it does not determine the etiology of the abnormality, and it is not correlated to predicted risk of bleeding from surgical procedures.

Prothrombin Time (PT)

The PT reflects activation of the extrinsic (F VII) and common pathways (F X, V, II and I). Blood should be drawn into a citrated (blue top) tube and filled to the bottom of the top of the label (blood: citrate ratio of 9:1) The amount of blood drawn is important since dilution with citrate can alter clotting times. PT is the first parameter to increase with ACR toxicity, although overt bleeding does not occur until both PT and aPTT are elevated.

Activated Partial Thromboplastin Time (aPTT)

The aPTT reflects activity of the intrinsic (F XII, XI, IX, VIII) and common pathways. Blood should be drawn into a citrated tube as described above. The SCA-2000 coagulation analyzer is the most common machine used in private practice to determine clotting times. These machines cost roughly $2-3000. Cartridges cost around $4.00 each, and are stored at room temperature. Shelf life of the cartridges is 30 days at room temperature, or 60 days refrigerated. Cartridges that are kept in the refrigerator need approximately 10 minutes to warm up before use.

Activated Clotting Time (ACT)

The ACT is a rough estimate of the aPTT, although very low platelet counts can affect it as well. The ACT is performed by placing 2ml of whole blood into a pre-warmed (body temperature at 37 C) glass tube containing diatomaceous (Fullers) earth (grey top tube). After 60 seconds (keeping the tube at 37 C), the tube should be inverted every 10-15 seconds to check for clot formation. The test stops when a clot is noted in the tube. The normal range is 60-110 seconds in dog. A specialized heating block can be used to keep the tube warm. Alternately, a cup of water warmed to body temperature can be used.

Proteins Induced by Vitamin K Antagonism (PIVKA)

Inactive precursors of vitamin K dependent coagulation factors are stored in the liver. In the absence of vitamin K, which is required to activate these factors, the precursors build up and can spill over into the blood. Without vitamin K to activate them, there is depletion of activated coagulation factors, which leads to prolongation of clotting times and clinical bleeding. The human PIVKA test actually measures concentrations of these inactive PIVKA whereas the veterinary test marketed as the PIVKA is actually just a modified prothrombin time (PT). The veterinary PIVKA does not offer any advantage over simply measuring PT. It has also been shown to be elevated secondary to a number of disease conditions and is not specific for anticoagulant rodenticide toxicity.


vWF Assays

vWF antigen ELISA assays are useful for the diagnosis of Type I and Type III vWD. Dogs with Type I vWD generally have vWF antigen levels that are less than 15% while dogs with Type III vWD have no detectable vWF. The quantitative nature of these assays makes them unsuitable as stand-alone assays for the diagnosis of Type II vWD.

In academic or referral settings, thromboelastrography and platelet function analysis are available, but these are not yet readily available to the general practitioner.

What to do with the actively bleeding patient?

In private practice, a microscope for estimation of platelet counts, a BMBT kit and an ACT can be used to help diagnose most clotting disorders. While they may not be 100% diagnostic, at least using these tests will help to determine therapy. Unfortunately, red top clotting times and toenail trim tests are not accurate for diagnosis of coagulopathy. These are too dependent on inter- and intra-operator variability. Additionally, cross-contamination of EDTA from filling blood tubes (i.e., filling the EDTA tube first) can alter coagulation times.

Take for example the middle aged Golden Retriever with epistaxis. The platelet smear can help to rule in or out severe thrombocytopenia. The ACT can help to determine if a disorder of secondary hemostasis is the cause. Severe prolongations in ACT (> 4 min) are likely to be associated with ACR toxicity. A BMBT can be used to determine if thrombocytopathia is the underlying cause for hemorrhage. The below table can be used to help with diagnosis of hemostatic disorders.


Disease Process



Platelet Count




ACR Toxicity


Slight ↑?

Slight ↓












Hemophilia (A or B)










Normal to Slight ↑

Severe Hemorrhage

Normal to slight ↑

Slight ↓

Normal to slight ↑

Normal to slight ↑

Normal to slight ↑


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