Laboratory EvaluationLaboratory tests play a very important role in the diagnosis, monitoring, and treatment ofcoagulation disorders. When used in the appropriate manner, they can yield a great deal ofinformation and can often localize a disease process to a certain part of the cascade.Furthermore, these tests are extremely helpful in the monitoring of anticoagulant therapy.However, in order to realize their true utility and limitations, one must first understandexactly how they are performed. With this knowledge, a clinician can then form an adequateinterpretation. Indeed, the tests by far reachtheir highest specificity and senstivity only whenthe clinical picture is known. The most common laboratory evaluations used are the activatedprothrombin time (aPT) and activated partial thromboplastin time (aPTT). Other tests such asthrombin time and bleeding time are also extremely valuable, but often underused.Activated Prothrombin TimeActivated Partial Thromboplastic TimeThrombin TimeBleeding TimeFibrinogen ConcentrationsAssays for Lupus AnticoagulantsFactor VIII Inhibitor AssayvonWillebrand Disease Assays
Prothrombin Time**Purpose Screening test to indentify acquired or inherited deficiencies of factors VII, X, V,PT , FIB . Monitor oral anticoagulant therapy e’ warfarin, which decreases the activity of VII, IX, X , PT.Method Collect blood from the patient into a citrated tube Take the collected blood to the laboratory Centrifuge to isolate the plasma Take small sample of the plasma ~0.1 ml. Add calcium Add Thromboplastins - which are preparations of TF and phospholipids from rabbit brain . The time that elapses until clot forms is the activated Prothrombin Time (PT).Interpretation PT shorter than the reference range is not associated with any clinical condition and is usually related to improper procedure. Prolongation of thePT occurs when there is: - Increased hematocrit, leading to a false +ve in PT prolongation. - Liver disease. - Vitamin K deficiency - DIC. - Nephrotic Syndrome - Treatment e’ antibiotics, chemotherapeutics, or AT drugs. Note: The PT will increase shortly after a bolus dose of heparin. The monitor of anticoagulant therapy requires standardization of the PT between different laboratories in order to ensure that the degree of anticoagulation measured in one laboratory can be compared to that in another laboratory. Therefore, the INR was created. The INR takes into account variations in reagents and techniques between different labs and provides one value that can be compared between labs.
Partial Thromboplastin TimePurpose Screening test to indentify acquired or inherited deficiencies of Factors IX, VIII, and XI. Screening test for lupus anticoagulants Monitor heparin anticoagulation Screening test to assess reduction in the activity of FIB , V , X. However, the PT is more sensitive for these.Method Collect blood from the patient into a citrated tube Take the collected blood to the laboratory Centrifuge to isolate the plasma Take a small sample of the plasma ~0.1 ml Add calcium (catalyst for coagulation) Add partial thromboplastin -phospholipid source without tissue factor Add a negatively charged surface such as kaolin, silica, or dextran So4 to activate coagulation The time that elapses until clot forms is the activated Partial Thromboplastin Time (aPTT).Interpretation aPTT shorter than the reference range is usually the result of poor sample handling. Prolongation of the aPTT occurs when there is: - Factor Deficiency (see below) - Factor Inhibitor (see below) - Anticoagulation with heparin - Contamination of sample with heparin Note: Once heparin is started, the laboratory work-up of an abnormal aPTT is difficult. Low molecular weight heparin(LMWH) and heparinoids generally do not prolong the aPTT.
When attempting to determine the cause of a prolonged aPTT, one must first rule out the most common cause which is contamination of the blood specimen with heparin. This usually occurs when small amounts of heparin are used to keep venous or arterial catheters patent. If this is ruled out and the patient is not on intravenous heparin for anticoagulation, then one can begin to suspect either a factor deficiency or a factor inhibitor. A factor inhibitor is usually an antibody targeted against a specific coagulation factor, which results in activation of that factor. Examples of this are the lupus anticoagulant and Factor VIII inhibitors. The distinction of inhibitor versus factor deficiency can be made by performing a 1:1 mix of the patients plasma with plasma pooled from normal individuals. (Pooled plasma is plasma taken from a large number of individuals with no known coagulation defects.) In theory, it should contain normal levels of all coagulation factors. In a patient with factor deficiency, aPTT measured after mixing the patients and pooled plasma should be normal. In patients with factor inhibitor, mixing the pooled plasma and the patients plasma will still yield a prolonged aPTT because the inhibitor in the patients plasma will also inactivate the specific factor in the pooled plasma.Thrombin TimePurpose Screening test for hypofibrinogenemia, hyperfibrinogenemia, Dysfibrinogen , and inhibitors against thrombin or fibrin. Tests the conversion of fibrinogen to fibrin to cross-linked fibrin. Monitor anticoagulant therapy with fibrinolytic agents and hirudin.. The test can be used to measure fibringoen levels.Method Collect blood from the patient into a citrated tube Take the collected blood to the laboratory Centrifuge to isolate the plasma Take small sample of the plasma ~0.1 ml.
Add bovine thrombin to plasma The time that elapses until clot forms is the Thrombin Time (TT).Interpretation TT shorter than the reference range is not associated with any clinical condition is usually related to improper procedure. Prolongation of the TT occurs when there is: 1. Contamination with heparin. 2. TT can be repeated after addition of protamine sulfate and will be normal.. 3. Afibrinogenemia/Hypofibrinogenemia – 4. Acquired (DIC, liver disease) and familial 5. Interference with fibrinopeptide cleavage
Bleeding TimePurpose Screening test for congenital and acquired disorders of platelet function. Screening test for vonWillebrands disease.Method A spring loaded device with a blade is used to make a small incision in the skin. The timer is started. Filter paper discs are used to adsorb droplets of blood at regular intervals. The time required for bleeding to cease is the Bleeding Time.Interpretation Prolonged Bleeding Times are usually due to: - Aspirin and other drugs that interfere with platelet function. - Congenital or acquired disorders of platelet function. - vonWillebrand Disease - can further work-up by measuring PTT, Factor VIII activity, Factor VIII antigen, and Ristocetin cofactor activity.Fibrinogen ConcentrationPurpose The Thrombin Time is used to screen for reduction in fibrinogen concentration. The Fibrinogen Titer Test can also be used. (Discussed below.)Methods Thrombin Time (Click to go to Thrombin Time section) Fibrinogen Titer Test 1. Thrombin is added to serial dilutions of plasma in buffered saline. 2. The dilutions in which clot formation is visible are noted.Interpretation Clot should normally be visible at a dilution of 1:64, and is usually seen at 1:128. Visible clot in these dilution rules out a clinically significant quantitative abnormality in fibrinogen and estimates the concentration to be greater than 100 mg/dL.