D-Dimer Assays - Pathology

D-Dimer Assays 1
D-Dimer Assays
Feature
Roger S. Riley, M.D., Ph.D.
Ann Tidwell, M.T. (ASCP) SH
April, 2005
Synonyms
XDP, Fragment D-dimer, Fibrin degradation fragment.
Test Description
The D-dimer assay is specific for fibrin derivatives. In this assay, the presence of
cross-linked D-dimer domain is diagnostic for lysis of a fibrin clot, and confirm that
thrombin was formed and Factor XIII was activated with reactive fibrinolysis. Since
fibrinogen derivatives do not contain the cross-linked D-dimer domain, they are not
recognized by the D-dimer assay, even when present in high concentration. In other
words, fibrin derivatives in plasma containing D-Dimer (XDP) are specific
markers for fibrinolysis, as opposed to fibrinogenolysis. D-dimers are detected
by immunoassays using monoclonal antibodies specific for the cross-linked D-dimer
domain in fibrinogen. Commercially available assays include latex agglutination, immunoturbidimetry,
and ELISA.
Patient
Preparation
Specimen
No specific patient preparation is required for the measurement of D-dimers.
Citrated, platelet-poor plasma is used for the D-dimer assay.
Specimen
Collection
and
Preparation
Citrated, platelet-poor plasma is prepared from venous blood collected by venipuncture
or from an indwelling catheter. The blood is collected into 3.2%) trisodium citrate
at a ratio of 9:1. The blue-top tube automatically fills to the correct volume; spurious
results may occur if this ratio is not maintained. The citrate concentration must be adjusted
in patients with a HCT >55%. Plasma should be separated from the cells as
soon as possible after the specimen is obtained. D-dimers are stable for 8 hours in citrated
plasma maintained at room temperature, for seven days if stored at 2-8 o C, and
up to two months at -20 o C.
Test
Methodology
D-dimers are detected by immunoassays using monoclonal antibodies specific for the
cross-linked D-dimer domain in fibrinogen. Present commercially available assays are
based on particle agglutination, immunoturbidimetry, and ELISA.
ELISA assays are the reference standard for D-dimer quantitation. These assays utilize
microtiter wells coated with an antibody with a high affinity for D-dimer. Incubation
with plasma results in the binding of any D-dimer present. A labeled antibody is then
added and the amount of bound labeled substance is determined by a colormetric reaction.
In spite of their high sensitivity and specificity, conventional ELISA assays are
expensive, labor intensive, and time consuming to perform. Therefore, they have not
been practical in most clinical situations, where rapidly available results are needed.
Latex agglutination (LA) assays use latex microparticles coated with monoclonal antibodies
specific for D-dimer. Incubation with plasma results in the formation of macroscopic
agglutinates. The sensitivity of the assay is usually adjusted to 1 g/ml during
the manufacture of the latex particles. Although conventional latex agglutination as-

D-Dimer Assays 2
Test
Methodology
says are inexpensive and rapid to perform, numerous studies have shown that they
lack the sensitivity to detect D-dimers in critical clinical situations, particularly in the
detection of pulmonary embolism and acute venous thrombosis.
Immunoturbidometric assays are automated microparticle assays in which a beam of
monochromatic light is passed through a suspension of latex microparticles coated by
covalent bonding with monoclonal antibodies specific for D-dimer. The wavelength of
the light (540 nm) is greater than the diameter of the latex microparticles and so the
solution of latex microparticles only slightly absorbs the light. When the plasma is
added to the suspension, any D-dimer present in the sample causes the latex microparticles
to agglutinate, becoming aggregates with diameters greater than the wavelength
of the light. This increases the absorbance of the light, which is measured photometrically,
and proportional to the amount of D-dimer present in the test sample.
These assays are cost effective, relatively rapid to perform, and have a sensitivity
comparable to conventional ELISA.
A commercially available whole blood assay for D-dimer uses a bispecific antibody specific
for D-dimer and a red blood cell antigen. A drop of whole blood is incubated with
the monoclonal antibody solution, causing visible agglutination of the red cells if D-
dimers are present. Many reports have indicated a high sensitivity for the assay, and it
is widely used in clinical settings.
Normal Values and
Critical Limits
The reporting standard for D-dimers varies with the test methodology and reagent
manufacturer. Latex and whole blood agglutination results are usually reported
as a D-dimer range (ng/mL). ELISA and immunoturbidimetric assays are
usually reported in fibrinogen equivalent units (FEU). One FEU is the quantity of
fibrinogen that was initially present before it was broken down. The actual
quantity of D-dimer is approximately half of an FEU (when 1.0 of fibrinogen g/
mL is broken down, 0.5 g/mL of D-Dimer remain).
Interferences
Rheumatoid factor may cause a false positive D-dimer assay. Lipemia may interfere
with immunoturbidimetric and ELISA assays.
Clinical Utilization
XDPs are cross-linked fibrin degradation products which arise directly from fibrin.
Thus, the measurement of XDPs, unlike total FDPs, is a specific measure of fibrinolysis.
Elevated D dimers are seen in DIC, pulmonary embolism, arterial and venous
thrombosis, septicemia, cirrhosis, carcinoma, sickle cell crisis, and following operative
procedures. Both FDPs and XDPs are present during late pregnancy and for approximately
48 hours post-surgery. During fibrinolytic therapy the FDP test is positive, while
the D-dimer test is negative in the absence of thrombolysis.
Disseminated intravascular coagulation (DIC, consumption coagulopathy) is one of the
most common and clinically important acquired disorders of hemostasis. In DIC, intravascular
activation of the coagulation system results in the widespread deposition of
fibrin microthrombi in the microcirculation, the consumption of platelets and clotting
factors, and activation of the fibrinolytic system. At the same time that thrombin converts
fibrinogen to fibrin, it also activated Factor XIII to form a plasma transglutaminase,
Factor XIIIa, which stabilizes fibrin by cross-linking the gamma chains of fibrinogen
in the region of the D-domain. Plasmin digests fibrin and fibrinogen to produce
fibrin(ogen) degradation (FDP) (or split, FSP) products (X,Y,D, and E), which are removed
from the circulation by the reticuloendothelial system.

D-Dimer Assays 3
Clinical Utilization
DIC is not a specific disease, but a sequalae of many pathologic conditions, including
acute intravascular hemolysis, hemolytic transfusion reactions, shock, hyperthermia,
extensive tissue damage, malignancies, obstetric complications, hyperthermia, snake
bites, etc. Prompt diagnosis and therapy of DIC is essential, since the associated hemorrhage,
small vessel thrombosis, and occasional large vessel thrombosis can lead to
the impairment of blood flow, ischemia, end-organ damage, and death. The clinical
signs and symptoms in DIC are variable and non-specific, and include fever, hypotension,
acidosis, proteinuria, hypoxia, petechiae and purpura, subcutaneous hematomas,
bleeding (surgical wound, traumatic wound, venipuncture), and arterial line oozing.
There is no laboratory assay pathognomonic of DIC. However, peripheral smear examination
is among the most helpful. In fulminent cases, this shows a characteristic
combination of schistocytes (red blood cell fragments), mild polychromatophilia, leukocytosis
with a left shift, thrombocytopenia, and large young platelets. However, the
diagnosis of DIC cannot be excluded by an absence of schistocytes, since these occur
in only 50% of patients. In addition to peripheral smear review, the prothrombin time
(PT), activated partial thromboplastin time (aPTT), platelet count, and serum fibrin/
fibrinogen degradation products are the best screening tests for DIC. The D-dimer assay
is more specific, and can be used to confirm the diagnosis.
The D-dimer has received much attention in recent years as a critical part of the
evaluation of emergency care patients with suspected pulmonary embolism (PE) or
deep venous thrombosis (DVT). Generally, numerous studies have shown that the
more advanced ELISA and immunoturbidometric assays have a sensitivity of 90% or
greater for PE or DVT in this setting, and a negative predictive value of 90% or greater
for the exclusion of disease. However, there is presently much debate about how the
results should be correlated with other findings, particularly radiographic studies.
The analysis of plasma D-dimers has been reported to be of diagnostic value in patients
with suspected complications of pregnancy such as pre-eclampsia and the HELLP
syndrome, to monitor anticoagulant and thrombolytic therapy, and to correlate with
disease severity in rheumatoid arthritis. CSF D-dimers have been reported positive in
patients with subarachnoid hemorrhage, but not in normal patients or those with
traumatic lumbar puncture.
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D-Dimer Assays 4
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(Cont’d)
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