From: Elizabeth M. Van Cott, M.D., and Michael Laposata, M.D., Ph.D., "Coagulation." In: Jacobs DS et al, ed. The Laboratory Test Handbook, 5th Edition. Lexi-Comp, Cleveland, 2001; 327-358.
Index of Tests
Antiphospholipid Antibody (Lupus Anticoagulant and/or Anticardiolipin
Synonyms Lupus Inhibitor
Applies to Anticardiolipin Antibody; Antiphosphatidylserine
Antibodies; Beta-2-Glycoprotein I; Cardiolipin; Circulating Anticoagulant;
Lupus Anticoagulant; PTT
Abstract Antiphospholipid antibodies are associated with
an increased risk of thrombosis, thrombocytopenia, and recurrent
Specimen Lupus anticoagulant: plasma; anticardiolipin antibody:
Container Lupus anticoagulant: blue top (sodium citrate)
tube; anticardiolipin antibody: red top tube
Collection Routine venipuncture. If multiple tests are being
drawn, draw blue top tubes after any red top tubes but before any
lavender top (EDTA), green top (heparin), or gray top (oxalate/fluoride)
tubes. Immediately invert gently at least 4 times, mixing thoroughly.
Blue top tubes must be appropriately filled. Deliver immediately
to the laboratory.
Storage Instructions Separate plasma (or serum) from cells
as soon as possible. Plasma (or serum) may be stored on ice for
up to 4 hours; otherwise, store frozen. Platelet count must be <10
x 109/L in plasma prior to freezing, or false-negative
lupus anticoagulant results may occur.
Causes for Rejection Blue top specimen received more than
4 hours after collection; clotted specimen; blue top not filled;
patient on hirudin, danaparoid, or argatroban anticoagulation
Turnaround Time Lupus anticoagulant: less than 1 day if negative,
longer if positive because confirmatory assays need to be performed;
anticardiolipin: often several days because testing is batched.
Special Instructions Notify laboratory if patient is on heparin,
including subcutaneous low-dose heparin or low-molecular weight
heparin. In some assays, heparin may cause false-positive lupus
anticoagulant results. Therefore, heparin must first be removed
from the specimen by the laboratory. Other assays contain a heparin
neutralizer that tolerates specimens containing up to 1 unit/mL
heparin. Results can be interpreted correctly in patients on Coumadin®.
Hirudin, danaparoid, or argatroban anticoagulation may cause false-positive
results in some assays.
Reference Interval Negative for lupus anticoagulant; <15
units of anticardiolipin antibody
Use If an antiphospholipid antibody is suspected, assays
for both lupus anticoagulant and anticardiolipin antibody should
be performed. Used to evaluate hypercoagulable states, recurrent
miscarriage, thrombocytopenia, or prolonged PTT. Lupus anticoagulants
may or may not prolong the PTT.
Limitations Factor VIII inhibitors can cause false-positive
lupus anticoagulant1 tests (a factor VIII assay showing
normal factor VIII levels rules out this possibility). The transient
presence of antiphospholipid antibodies may accompany infections
or drugs; vide infra.
Anticardiolipin antibody: Enzyme-linked immunosorbent assay
(ELISA) using cardiolipin, a phospholipid, as the antigen. Newer
ELISA assays are available that test for anti-beta 2-glycoprotein
I antibodies or antiphosphatidylserine antibodies.
Lupus anticoagulant: To improve sensitivity, two screening
tests are suggested.2 These tests are clotting-time based
assays, such as the Russell viper venom time, PTT-based assays,
kaolin clotting time, or dilute prothrombin time (tissue thromboplastin
inhibition test). Lupus anticoagulants prolong various clotting
times in the laboratory because they bind to phospholipid and thereby
interfere with the ability of phospholipid to serve its essential
cofactor function in the coagulation cascade. Lupus anticoagulant
screening assays usually have a low concentration of phospholipid
to enhance sensitivity. Any abnormal (prolonged) screening result
is repeated after a 1:1 mixture of patient plasma with normal plasma
to demonstrate that the clotting time remains prolonged upon mixing.
Confirmatory assays are performed if the screening assay remains
abnormal after the 1:1 mixture. Confirmatory assays typically demonstrate
that upon addition of excess phospholipid, the clotting time shortens
toward normal. The "platelet neutralization procedure" is a confirmatory
assay in which the source of the excess phospholipid is freeze-thawed
platelets. Note: The routine PTT may or may not be prolonged,
depending on the amount of phospholipid in the reagent. In addition,
elevated factor VIII can normalize an otherwise prolonged PTT. PTT-based
lupus anticoagulant screening assays have a low concentration of
phospholipid to enhance sensitivity. When the PTT is prolonged,
a PTT mixing study may be a useful first test. When lupus anticoagulants
are present, the PTT remains prolonged upon mixing with an equal
volume of normal plasma.
Additional Information The two principal types of antiphospholipid
antibodies are lupus anticoagulants and anticardiolipin antibodies.
They are present in 0% to 5% of the general population and in 12%
or more of patients with thrombosis.3,4 Antiphospholipid
antibodies are acquired autoantibodies directed against phospholipid-protein
complexes. These antibodies are associated with an increased risk
for arterial or venous thrombosis,3,5 thrombocytopenia,6 and fetal loss.7 Associations with cardiac valve disease,
livedo reticularis, and other features are also recognized.8 The mechanism of thrombosis is not entirely clear, although a number
of mechanisms have been proposed. In a recent prospective study
involving individuals with antiphospholipid antibodies, the incidence
of thrombosis per year was 1% in individuals with no history
of thrombosis, 4% in patients with systemic lupus erythematosus,
5.5% in patients with a history of thrombosis, and 6% in individuals
with high titer IgG anticardiolipin antibody (>40 units).9
The diagnosis of antiphospholipid antibody syndrome requires
a positive test in the antiphospholipid antibody panel (lupus anticoagulant
and/or anticardiolipin antibody) on two separate occasions, at least
6 weeks apart, in the setting of thrombosis, thrombocytopenia, or
Anticardiolipin antibodies recognize cardiolipin bound to beta 2-glycoprotein I. Most lupus anticoagulants recognize
phospholipid bound to prothrombin, but others recognize phospholipid
bound to beta 2-glycoprotein I or other proteins. Rarely,
prothrombin levels become decreased as a result of a lupus anticoagulant,
and an increased risk for bleeding may develop.11 As
cardiolipin is the antigen used for syphilis screening tests (VDRL,
Venereal Disease Research Laboratories; and RPR, rapid plasma reagin),
false-positive syphilis tests may occur in patients with anticardiolipin
antibodies. Conversely, true syphilis infections can cause positive
anticardiolipin antibody test results.
Despite the prolonged clotting times, bleeding is not a typical
feature associated with these antibodies. Thrombocytopenia, if present,
is usually mild. Patients may have either a lupus anticoagulant
or an anticardiolipin antibody or they may have both antibodies.
A high percentage of patients with systemic lupus erythematosus
(SLE) or related autoimmune diseases have these antibodies. These
antibodies may also develop in patients without an underlying disorder.
The antibodies can appear transiently in association with certain
medications (eg, hydralazine, phenytoin) or infections. The human
immunodeficiency virus (HIV) is commonly associated with positive
tests for antiphospholipid antibodies.12 Infection-associated
antibodies may not be associated with clinical symptoms of antiphospholipid
antibody syndrome, and they tend to recognize phospholipid rather
than the phospholipid-protein complexes described above.13
Heparin treatment in patients with lupus anticoagulants can be
complicated by the fact that lupus anticoagulants may prolong the
baseline PTT and/or accentuate the PTT prolongation when heparin
is added. As such, heparin may be monitored with antifactor Xa assays.
If the antifactor Xa assay demonstrates that the heparinized PTT
is not affected by the lupus anticoagulant, cautious use of the
PTT may be considered for that patient.
1. Goudemand J, Caron C, De Prost D, et al, "Evaluation of Sensitivity
and Specificity of a Standardized Procedure Using Different Reagents
for the Detection of Lupus Anticoagulants,"Thromb Haemost,
2. Brandt JT, Triplett DA, Alving B, et al, "Criteria for the Diagnosis
of Lupus Anticoagulants: An Update,"Thromb Haemost, 1995,
3. Ginsburg KS, Liang MH, Newcomer L, et al, "Anticardiolipin Antibodies
and the Risk for Ischemic Stroke and Venous Thrombosis,"Ann Intern
Med, 1992, 117(12):997-1002.
4. Doig RG, O'Malley CJ, Dauer R, et al, "An Evaluation of 200
Consecutive Patients With Spontaneous or Recurrent Thrombosis for
Primary Hypercoagulable States,"Am J Clin Pathol, 1994, 102(6):797-801.
5. Vaarala O, Manttari M, Manninen V, et al, "Anticardiolipin Antibodies
and Risk of Myocardial Infarction in a Prospective Cohort of Middle-aged
Men,"Circulation, 1995, 91(1):23-7.
6. Galli M, Finazzi G and Barbui T, "Thrombocytopenia in the Antiphospholipid
Syndrome,"Br J Haematol, 1996, 93(1):1-5.
7. Yasuda M, Takakuwa K, Tokunaga A, et al, "Prospective Studies
of the Association Between Anticardiolipin Antibody and Outcome
of Pregnancy,"Obstet Gynecol, 1995, 86(4 Pt 1):555-9.
8. Hogan WJ, McBane RD, Santrach PJ, et al, "Antiphospholipid Syndrome
and Perioperative Hemostatic Management of Cardiac Valvular Surgery,"Mayo
Clin Proc, 2000, 75(9):971-6.
9. Finazzi G, Brancaccio V, Moia M, et al, "Natural History and
Risk Factors for Thrombosis in 360 Patients With Antiphospholipid
Antibodies: A Four-Year Prospective Study From the Italian Registry,"Am
J Med, 1996, 100(5):530-6.
10. Wilson WA, Gharavi AE, Koike T, et al, "International Consensus
Statement on Preliminary Classification Criteria for Definite Antiphospholipid
Syndrome,"Arthritis Rheum, 1999, 42(7):1309-11.
11. Bajaj SP, Rapaport SI, Fierer DS, et al, "A Mechanism for the
Hypoprothrombinemia of the Acquired Hypoprothrombinemia-Lupus Anticoagulant
Syndrome,"Blood, 1983, 61(4):684-92.
12. de Larranaga GF, Forastiero RR, Carreras LO, et al, "Different
Types of Antiphospholipid Antibodies in AIDS: A Comparison With
Syphilis and the Antiphospholipid Syndrome,"Thromb Res, 1999,
13. McNally T, Purdy G, Mackie IJ, et al, "The Use of an Antibeta-2-Glycoprotein-I
Assay for Discrimination Between Anticardiolipin Antibodies Associated
With Infection and Increased Risk of Thrombosis,"Br J Haematol,
Galli M and Barbui T, "Antiprothrombin Antibodies: Detection and
Clinical Significance in the Antiphospholipid Syndrome,"Blood,
Galli M, Finazzi G and Barbui T, "Antiphospholipid Antibodies:
Predictive Value of Laboratory Tests,"Thromb Haemost, 1997,
Martinez CE, Rivera GB, and Aguilar LD, "Anticardiolipin Antibodies
in Serum and Cerebrospinal Fluid From Patients With Systemic Lupus
Erythematosus,"J Investig Allergol Clin Immunol, 1997, 7(6):596-601.
Roubey RAS, "Antiphospholipid Antibody Syndrome,"Arthritis and
Allied Condition, Koopman WJ, ed, Baltimore, MD: Lippincott
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Triplett DA, "Protean Clinical Presentation of Antiphospholipid-Protein
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