Arthritis & Rheumatism, Volume 62,
November 2010 Abstract Supplement

Abstracts of the American College of
Rheumatology/Association of Rheumatology Health Professionals
Annual Scientific Meeting
Atlanta, Georgia November 6-11, 2010.


Evaluation of the Performance of Monoclonal and Polyclonal Antibody Standards in Different Assays for the Detection of Antiphospholipid (aPL) Antibodies: Report of a Wet Workshop at the 13th International Congress on Antiphospholipid Antibodies.

Forastiero8,  Ricardo, Papalardo6,  Elizabeth, Watkins1,  Michael, Nguyen1,  Hoang, Crisostomo1,  Maria, Vandam1,  Wendy, Hardy1,  JOel

Bio-Rad Laboratories, Hercules, CA
INOVA Diagnostics, San Diego, Ca
Instrumentation Laboratories, Bedford, MA
Phadia GMBH, Freiburg, Germany
TherTest, Lombard, IL
Univ of TX Med Branch, Galveston, Galveston, TX
Univ of TX Med Branch, Galveston, TX
Universidad Favaloro, Buenos Aires, Argentina

Background:

The standardization of tests used in the diagnosis of Antiphospholipid Syndrome (APS) [anticardiolipin (aCL) and the anti-b2glycoprotein I (ab2GPI)] is still a matter of debate and concern. One important topic of discussion is whether to use monoclonal (MoAbs) or polyclonal antibody preparations for the calibration and/or quality control of these assays. However, the proper cross-validation and evaluation of the available preparations is lacking.

Objective:

to evaluate the performance of various MoAbs and polyclonal antibody calibrators for the detection of aCL and ab2GPI antibodies (IgG, IgM) at a wet workshop at the 13th International Congress on Antiphospholipid Antibodies.

Methods:

Polyclonal standards G2 and M3 for IgG, IgM aCL [Louisville APL Diagnostics, Inc.(LAPL)], MoAbs IgG and IgM: a) HCAL and EY2C9 [Center for Diseases Control (CDC)]; b) AbyD05045 and AbyD03892 (Phadia); c) "Sapporo" (INOVA) were distributed to the groups for evaluation. All serum and calibrator preparations were from the same lot and were tested in the following aCL and ab2GPI IgG and IgM assays: QUANTA LITE™ (INOVA), TheraTest, Phadia, Bio-Rad (ELISA PhD™ lx and Multiplex BioPlex™ 2200), in the APhL ELISA®[an assay that utilizes a mixture of negatively charged phospholipids and b2GPI (LAPL)] and in a new chemiluminescent assay HemosIL® AcuStar Antiphosphilipid assay panel [Instrumentation Laboratory, IL)]. Calibrators and kits were assayed as per the manufacturers' instructions. The workshop was open to congress participants who registered for the event and the groups evaluated the calibrators at least on two occasions.

Results:

All groups reported values of the polyclonal aPL calibrators within the expected range including the new chemiluminescent and the multiplex assays. Excellent correlation and linearity was observed with two monoclonal preparations (AbyD05045 IgG and IgM and Sapporo INOVA IgG and IgM). As expected, AbyD3092 showed negative results in various tests (this antibody has aCL activity only). The "Sapporo" and the "CDC" monoclonal antibodies, although originally derived from the same hybridoma sources showed apparent different concentration, specificity and presentation (liquid affinity purified Ig for the Sapporo and freeze-dried hybridoma supernatant for the CDC). A large variability of the results of the CDC MoAbs was reported by the groups at this workshop when tested at the same final concentration (HCAL: from 8.2 to 75.3 GPL units for IgG aCL and 20.9 to 424.5 Units for IgG ab2GPI; EY2C9: from 30.4 to 145.3 MPL units for IgM aCL and from 0 to 520.4 Units for IgM ab2GPI). Participants reported difficulties with the reconstitution of the CDC MoAb freeze-dried material, indicating problems with the stability and variability between vials of the preparations. Those included not only differences among any given assay but also within the same test, run in more than one occasion.

Conclusions:

The LAPL polyclonal and the Phadia and INOVA monoclonal preparations are suitable sources for calibration of aCL and ab2GPI assays. Based on results obtained at this wet workshop, the use of the CDC MoAbs is questionable and not recommended at this time.

To cite this abstract, please use the following information:
Forastiero, Ricardo, Papalardo, Elizabeth, Watkins, Michael, Nguyen, Hoang, Crisostomo, Maria, Vandam, Wendy, et al; Evaluation of the Performance of Monoclonal and Polyclonal Antibody Standards in Different Assays for the Detection of Antiphospholipid (aPL) Antibodies: Report of a Wet Workshop at the 13th International Congress on Antiphospholipid Antibodies. [abstract]. Arthritis Rheum 2010;62 Suppl 10 :2251
DOI: 10.1002/art.30014

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