High frequency ultrasonic imaging and attachment of contrast agents under flow conditions

Background: Targeting of ultrasound contrast agents is an area of research which is continuously expanding. Contrast agents can be targeted to attach to markers expressed at areas of interest within the body. Areas of inflammation and molecules expressed at these areas are associated with unstable plaque in arteries. Imaging techniques to distinguish between unstable and stable plaques are currently unavailable.

Previously a flow chamber was described which allowed high frequency ultrasonic imaging of contrast agents attached to surfaces while subjected to flow. A lipid based contrast agent which can be targeted to molecules expressed at areas of inflammation has been developed and attached to agar, under static conditions, using avidin and biotin. It was shown that the contrast agent remained attached to the agar under shear stresses of up to 3.4 Pa. The work presented here has involved attaching the contrast agent under flow conditions.

Aim: To determine if the variation of biotin in the contrast agent shell affects attachment under flow conditions.

Methods: Streptavidin coated agar samples were placed in the flow chamber. Biotinylated lipid contrast agents, 0.5 ml in 50 ml saline, were circulated through the flow chamber for 2 hours at a flow rate of 1.65 ml min(-1). The samples were imaged with 40 MHz intravascular ultrasound. Images and RF data were captured before and after circulation of the contrast agent. Contrast microbubbles containing 1%, 3% and 5% biotin were tested. Saline and un-biotinylated contrast agent were also circulated.

Results: On the circulation of saline and un-biotinylated contrast agent over the streptavidin coated agar in the flow chamber, no enhancement was seen in the IVUS images. On circulation of the 1% biotinylated agent, enhancement of the agar surface was seen. After circulation of the 3% and 5% biotinylated contrast agents, again enhancement of the agar surface was seen, however, there was better coverage across the surface of the agar than for the 1% biotin agent. The raw RF data was used to determine the mean backscatter power from the agar surface, across the surface there was no significant increase seen after circulation of 1% biotin-agent. However, there was an increase in the mean backscatter power for the 3% and 5% biotinylated contrast agents.

Conclusions: Variations in the concentration of biotin within the shell can affect the attaching potential of targeted contrast agent. The agent was seen to attach to an agar surface under low flow rates, with better coverage of the agar sample with increasing levels of biotin.