Laboratory Investigation Of Suspected Haemostatic Disorders

Patient History

  • Family history (ie. inherited deficiencies)
  • Medication (eg. aspirin)
  • Type of bleeding
  • Age of onset
  • Post-op bleeding

Physical Examination

  • Organ involvement may provide info into underlying disease processes

Screening Tests

  • Haematology profile
  • Blood smear

Collection of a blood sample to investigate the coagulation mechanism

  • Not EDTA:
      Effect on function of fibrinogen
      Need to add calcium to effect clotting
  • Use citrate, collecting this sample last to avoid as much contamination by tissue thromboplastin as possible
  • Platelet Poor Plasma (PPP) is obtained by centrifugation of the sample at high speed
  • Plate Rich Plasma (PRP) is obtained by centrifugation at low speed - useful in platelet aggregation studies
The haemostatic method is tested at three major levels:
  1. The vascular level (platelet number and function via bleeding time)
  2. Coagulation factor presence and function
  3. vWF (and VIII)

Tests Include:

Bleeding Time (BT)

  • Can be carried out using either the Modified Ivy Technique or the Standardised Template Method (most sensitive)
  • Detects abnormalitites of vessel contraction
  • Detects abnormalities of platelet number and/or function (BT is inversely proportional to the platelet number - see Fig 1.)
  • Detects vWF syndrome

Prothrombin Time (PT)

  • Tests the extrinsic coagulation pathway
  • Relies on a reference thromboplastin (eg rabbit brain emulsion, which is added to citrated PPP, then "recalcified") and is carried out at 37oC
  • The reactions are carried out in glass
  • The PT is a non-specific test potentially detecting a deficiency in VII, X, V, II and I (singly or multiply)
  • Warfarin therapy affects II, VII, IX and X by inhibiting vitamin K synthesis
  • the result is reported as a Prothrombin Ratio (PR)

Activated Partial Thromboplastin Time (aPTT)

  • Tests the intrinsic pathway ie. XII, XI, IX, VIII, X, V, II, I
  • Relies on an activator (see below) + phospholipid added to citrated PPP, then "recalcified") and is carried out at 37oC
    • Ellagic acid
    • Celite
    • Kaolin
  • Monitors heparin therapy

Factor XIII Testing

  • Uses PPP to which calcium and thrombin are added
  • The clot which forms is incubated o/n in 2M urea. If the clot remains then Factor XIII is considered to have acted to form a stable clot
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