Evaluation of different immunoassays for the detection of antiphospholipid antibodies: report of a wet workshop during the 13th International Congress on Antiphospholipid Antibodies.

BACKGROUND: The performance and standardization of anticardiolipin (aCL) and anti-ß2 glycoprotein I antibodies (aß2GPI) tests for the confirmation of diagnosis of antiphospholipid syndrome (APS) remain a matter of debate and concern. We evaluated the performance of different ELISAs and other new immunoassays for the detection of aCL and aß2GPI in a wet workshop at the 13th International Congress on Antiphospholipid Antibodies in Galveston, TX (April 13th, 2010, APLA 2010). METHODS: Aliquots of 26 un-identified APS or persistently aPL positive serum samples and 21 controls (9 from healthy individuals and 5 from patients with infectious diseases and 7 with various autoimmune diseases) were distributed to all participants/groups. All serum samples were evaluated in various aCL and aß2GPI ELISAs, a chemiluminescent immunoassay, a fluoro-enzyme immunoassay, and in a multiplexed immunoassay system. Monoclonal and polyclonal calibrators were also evaluated. RESULTS: Although not all the assays reported the titers of aCL and aß2GPI in the same units, the correlation of positive titers among the assays was good. All aCL and aß2GPI tests showed excellent clinical sensitivities, specificities and positive predictive values and good agreement with respect to the levels of the IgG and IgM antibodies, regardless of assay type, or whether tests were done using automated or "manual" systems. CONCLUSIONS: New methodologies for the detection of aPL look promising and comparable to currently approved ELISA tests. This study provides evidence of progress of efforts of harmonization of tests used to detect aPL.

Wannier-Stark states in finite superlattices.

Individual Wannier-Stark states are resolved in a current experiment over a wide electric-field range for a 5 and 4 period finite superlattice utilizing a hot-electron transistor. The observed field dependence of the tunneling transmission through the various states directly resembles the progressive localization of the wave functions. The basic transport through Wannier-Stark states in short-period superlattices is identified to be coherent. By tuning the Wannier-Stark state splitting with electric field into the optical phonon energy, the opening of new LO-phonon mediated transport paths is observed.

Resonant tunneling mediated by resonant emission of intersubband plasmons.

We study tunneling through resonant tunneling diodes (RTD) with very long emitter drift regions (up to 2 microm). In such diodes, charge accumulation occurs near the double barrier on the emitter side, in a self-induced potential pocket. This leads to a substantial enhancement of the wave function overlap between states of the pocket and the RTD, and, consequently, to increased off-resonant current mediated by various scattering processes. For RTD with the longest drift region (2 microm), an additional strong current peak is observed between the first and the second resonant peaks. We attribute this pronounced feature to the intersubband transitions mediated by resonant emission of intersubband plasmons.

Both immunization with protein and recombinant vaccinia virus can stimulate CTL specific for the E7 protein of human papilloma virus 16 in H-2d mice.

The transforming protein E7 of human papilloma virus type 16 can stimulate cytotoxic T lymphocytes (CTL) which can protect experimental animals against growth of E7 expressing tumour cells. In this study we compared CTL responses in mice immunized with either E7 protein in MF59 adjuvant or with recombinant vaccinia virus expressing E7 (Vac-E7). We have chosen H-2d mice because no E7-specific CTL responses have been described in this MHC haplotype. Immunization of these mice with Vac-E7 generated CTL which lysed target cells infected with Vac-E7 or transfected with the E7 gene. CTL from mice immunized with E7 protein in MF59 adjuvant showed specificity for the same target cells. Antibody blocking experiments revealed that both immunization with Vac-E7 and E7 protein stimulated CD8+ effector CTL. The find specificity of CTL induced by the two immunization protocols was similar. A major CTL epitope was mapped to the carboxyl terminal amino acids 48-98 of the E7 protein. Peptide isolation from E7 expressing cells followed by HPLC separation indicated that CTL induced by immunization with protein and Vac-E7 recognized the same HPLC purified peptide fractions. Together, the study suggests that vaccines based on protein can activate CTL with similar fine specificity to CTL induced by vaccines based on recombinant vaccinia virus.

Penetration of hydrogen peroxide from contact lenses or tear-side solutions into the aqueous humor.

Contact lenses were soaked in hydrogen peroxide (H2O2) solutions of 1 to 20 mM (34 to 680 ppm) and placed on isolated rabbit corneas to determine whether H2O2 could penetrate across the tissue into the artificial aqueous humor used to perfuse the endothelial surface. Corneas with intact epithelium allowed no H2O2 to cross into the perfusing fluid even with the lenses containing the highest (20 mM) concentration of peroxide. If the epithelium was removed a transient pulse of H2O2 appeared in the perfusing fluid only from lenses with 10 or 20 mM H2O2. The cornea metabolized H2O2 rapidly (the rate varying with the concentration) and thus the small quantities of H2O2 in the contact lenses (less than 400 nmol at 20 mM) are destroyed before diffusing across the entire thickness of the cornea. When the contact lens was replaced by a 0.8 ml saline containing H2O2, and renewed every 15 min, H2O2 crossed the intact cornea to the perfusing fluid when its concentration at the epithelium was between 3 and 4 mM. Should such larger quantities be presented to the epithelium (for example, in eye drops) the concentration, volume, and duration of exposure will determine whether H2O2 enters the anterior chamber. It is concluded that in the clinical situation of typical contact lens use in an eye with intact epithelium neither the corneal endothelium nor other intraocular tissues will be damaged by residual concentrations of H2O2 up to 680 ppm, whether in single or daily events.