Measurement of isoagglutinins in immunoglobulins for intravenous application by flow cytometry.

Isoagglutinins present in intravenous immunoglobulin (IVIG) products have been linked to haemolysis. Therefore, accurately assessing isoagglutinin content in IVIG products is important. The standard European Pharmacopoeia (Ph.Eur.) direct assay is limited by low precision. Here, we describe the development of a fluorescence-activated cell sorting (FACS) method for assessing isoagglutinin levels. Serially diluted IVIG samples were incubated with red blood cells (RBCs), RBC-bound anti-A and anti-B antibodies were detected using a fluorescently-labelled antibody and the median fluorescence intensity of samples was assessed by FACS. Results were compared with the Ph.Eur. direct assay. The method was used to determine isoagglutinins in commercial products produced with and without isoagglutinin reduction steps. Assay precision, reported as the coefficient of variation, for the FACS method was 14% and 8% for anti-A and anti-B, respectively versus 33% and 20% with the Ph.Eur. direct assay. Application of the method on commercially available IVIGs revealed differences in isoagglutinin content between products produced with and without isoagglutinin reduction steps. This FACS assay allows for quantification of isoagglutinin concentrations in IVIGs with higher precision than the Ph.Eur. direct assay. Also the FACS assay confirms differences in isoagglutinin levels between IVIG products and the efficacy of isoagglutinin reduction measures.

Transient magnetorheological response of magnetoactive elastomers to step and pyramid excitations.

Transient rheological response of magnetoactive elastomers is experimentally studied using dynamic torsion at a fixed oscillation frequency in temporally stepwise changing magnetic fields and oscillation amplitudes. For step magnetic-field excitations, at least three exponential functions are required to reasonably describe the time behavior of the storage shear modulus over long time scales (>10(3) s). The deduced characteristic time constants of the corresponding rearrangement processes of the filler network differ approximately by one order of magnitude: τ1 ≲ 10(1) s, τ2 ∼ 10(2) s, and τ3 ∼ 10(3) s. The sudden imposition of the external magnetic field activates a very fast rearrangement process with the characteristic time under 10 s, which cannot be determined more precisely due to the measurement conditions. Even more peculiar transient behavior has been observed during pyramid excitations, when either the external magnetic field was first stepwise increased and then decreased in a staircase manner at a fixed strain amplitude γ or the strain amplitude γ was first stepwise increased and then decreased in a staircase manner at a fixed magnetic field. In particular, the so-called "cross-over effect" has been identified in both dynamical loading programs. This cross-over effect seems to be promoted by the application of the external magnetic field. The experimental results are discussed in the context of the specific rearrangement of the magnetic filler network under the simultaneous action of the external magnetic field and shear deformation. Striking similarities of the observed phenomena to the structural relaxation processes in glassy materials and to the jamming transition of granular materials are pointed out. The obtained results are important for fundamental understanding of material behavior in magnetic fields as well as for the development of devices on the basis of magnetoactive elastomeric materials.

Novel formulation of a reconstituted high-density lipoprotein (CSL112) dramatically enhances ABCA1-dependent cholesterol efflux.

OBJECTIVE: The ability of high-density lipoprotein (HDL) to remove cholesterol from atherosclerotic plaque is thought to underlie its inverse correlation with cardiovascular risk. Our objective was to produce and characterize a human apolipoprotein AI (apoA-I) product optimized to treat clinical atherosclerotic disease. APPROACH AND RESULTS: A new formulation of full length, plasma-derived human apoA-I termed CSL112 was designed to maximize the cholesterol efflux from cells and exhibit favorable pharmacological properties. CSL112 is a disc-shaped particle that strongly elevates cholesterol esterification and shows good pharmacokinetics in rabbits. Infusion of CSL112 into rabbits caused a strong and immediate increase in the ATP binding cassette transporter A1 (ABCA1)-dependent efflux capacity of plasma, an increase in plasma unesterified cholesterol and rapid subsequent cholesterol esterification. In the presence of human plasma, CSL112 was significantly more potent than native HDL at enhancing cholesterol efflux from macrophages, and the efflux elevation was predominantly via the ABCA1 transporter. Consistent with this observation, addition of CSL112 to plasma led to generation of high levels of HDL-VS, a favorable substrate for ABCA1. The lipid profile of plasma did not affect these behaviors. In studies with whole human blood, CSL112 reduced expression of intercellular adhesion molecule 1 and cytokine secretion, and as with cholesterol efflux, these activities were substantially greater than those of native HDL assayed in parallel. CONCLUSIONS: CSL112 has favorable pharmacological properties and strongly elevates the ability of plasma to withdraw cholesterol from cells. Preferential elevation of ABCA1-dependent efflux may target atherosclerotic plaque for cholesterol removal and this property makes CSL112 a promising candidate therapy for acute coronary syndrome.