Andreas Briel

Ultrasound derived imaging and quantification of cell adhesion molecules in experimental autoimmune encephalomyelitis (EAE) by Sensitive Particle Acoustic Quantification (SPAQ).

Molecular imaging requires, not only the identification of an appropriate marker, but also its quantitative analysis. We used the Sensitive Particle Acoustic Quantification (SPAQ) technology - a novel ultrasound technique - for detection and quantification of cell adhesion molecules in isolated tissue and in live animals. By conjugating gas-filled microparticles (MPs) with antibodies to intercellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion molecule 1 (VCAM-1), we were able to depict and quantify ICAM-1 and VCAM-1 in isolated brain and spinal cord from rats with autoimmune encephalomyelitis (EAE), an established inflammatory disease model of human multiple sclerosis (MS). Depiction and quantification of specific MPs were also feasible in living animals with AT-EAE with similar results. After treatment with methylprednisolone, the measured number of targeted anti-ICAM-1 and VCAM-1-MPs was significantly lower (P<0.01) compared to untreated animals demonstrating the high sensitivity of this imaging technique. Depending on the antibody linked to the surface of the MPs, the technique can be used to quantify the expression of any accessible antigen expressed on the luminal surface of endothelial cells and is therefore a promising tool for the non-invasive and dynamic assessment of disease-related molecules.

A Real-Time in Vitro Assay for Studying Functional Characteristics of Target-Specific Ultrasound Contrast Agents

AbstractPurpose. To develop an in vitro assay for studying the feasibility of specific targeting of ultrasound contrast agents (USCAs) for ultrasound diagnostics by employing the parallel plate flow chamber, which provides an environment that mimics some aspects of the in vivo conditions like shear rate and flow effects. Methods. USCAs based on air-filled microparticles (MP) were functionalized with specific antibodies using carbodiimide coupling chemistry and characterized by fluorescence activated cell sorter (FACS). The binding experiments were done by subjecting the MP to shear stress as they interact with the target-coated surface of the flow chamber. Results. A successive modification of MP with antibody and the glass surface with antigen was achieved and quantified. The binding studies showed specific attachment of targeted MP to EDB-FN (EDB domain of fibronectin) surface. The binding of MP via nonspecific interactions was minimal. The binding efficiency of antibody-loaded MP is dependent on the applied shear stress. An increase in the wall shear stress resulted in a decrease in binding efficiency. Binding efficiency was found to be correlated with the antibody density and antigen density on the interacting surfaces. Conclusions. The results indicate that the test system developed is reliable for characterizing targeted MP without any additional labeling and can be used as a functionality assay for studying the binding characteristic of USCA with respect to different parameters like density of targeting antibodies on the microparticle surface and of target protein. In addition, the microparticles can be studied in detail under different shear rates and flow conditions. Further studies concerning the in vitro-in vivo correlation will be necessary to further increase the value of this in vitro method.