An ELISA system to detect anti-factor VIII antibodies without interference by lupus anticoagulants. Preliminary data in hemophilia A patients.
OBJECTIVE
Difficulties in identifying the coexistence of neutralizing anti-factor VIII antibodies (anti-fVIII) and lupus anticoagulant (LA) are mainly due to the interference of LA on anti-fVIII assays. Our aim was to reveal the presence of anti-fVIII using a system that is not affected by LA.


METHODS
We developed an enzyme-linked immunosorbent assay (ELISA) method that uses phospholipid-free recombinant factor VIII as the antigen. A monoclonal anti-fVIII was tested as a positive control, excluding non-specific binding by using two unrelated monoclonal antibodies. The ELISA was performed on hemophilic plasmas with anti-fVIII and negative LA (n=12) or without inhibitors (n=12). Two hemophilic plasmas with LA and presumably anti-fVIII were also assayed. Positive LA (n=12) and normal (n=10) plasmas were tested as negative controls.


RESULTS
All (12/12) plasmas with anti-fVIII and 5/12 hemophilic plasmas without inhibitors were positive; LA and normal plasma controls were negative.


CONCLUSIONS
Results presented here show that LA does not interfere with the anti-fVIII ELISA: However, the assay detects both neutralizing and non-neutralizing anti-fVIII antibodies, therefore a neutralizing effect must be confirmed through functional tests.
Blanco
A N
AN
Instituto de Investigaciones Hematológicas "Mariano R. Castex", Academia Nacional de Medicina, Pacheco de Melo 3081, 1425 Buenos Aires, Argentina. blanco@connmed.com.ar
Peirano
A A
AA
Grosso
S H
SH
Gennari
L C
LC
Bianco
R P
RP
Lazzari
M A
MA
D000906:Antibodies / Q000097:blood / Q000276:immunology*; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay; D005169:Factor VIII / Q000276:immunology*; D006467:Hemophilia A / Q000097:blood / Q000276:immunology*; D006801:Humans; D016682:Lupus Coagulation Inhibitor / Q000276:immunology; D011994:Recombinant Proteins / Q000276:immunology; D012680:Sensitivity and Specificity
D000906:Antibodies; D016682:Lupus Coagulation Inhibitor; D011994:Recombinant Proteins; D005169:Factor VIII