Ultrasound is one of the most widely used imaging techniques in medicine, but up until recently it hardly played a role in imaging the tiniest structures of our bodies such as cells. "Clinical ultrasound, like the kind used for pregnancy scans, creates real-time images of body parts", first author Baptiste Heiles explains. "It allows diagnosis of various diseases, or to monitor a developing baby. However, what is going on at a microscopic level remains hidden."
Imaging living cells in 3D
Now, a team of scientists from TU Delft, the Netherlands Institute for Neuroscience and Caltech managed to image specifically labelled cells in 3D with ultrasound. For the first time, they imaged living cells inside whole organs across volumes the size of a sugar cube. In comparison, current light-based microscopes often require imaging of non-living samples, Heiles says. "The sample or organ of interest has to be removed and processed, and you lose the ability to track activity of cells over time".
Artist's impression of the new ultrasound technique, called nonlinear sound sheet microscopy. The blue lines represent two ultrasound waves intersecting in an X-shape pattern to scan cells and capillaries in body tissue. The cells and capillary vessels are labelled with ultrasound probes (in cyan and pink), making them light up in ultrasound images. (Credits: Maayan Harel, Maresca Lab)
The present leading technology to image how living cells behave in 3D, for example during the development of embryos, is called light sheet microscopy. This method is limited to translucent or thin specimens because light cannot penetrate deeper than 1 mm in opaque tissue. "Ultrasound can image centimetres deep in opaque mammal tissue, allowing non-invasive imaging of whole organs. This gives us information about how cells behave in their natural environment, something that light-based methods can't do in larger, living tissues", lead researcher David Maresca says.
Labelling capillaries and cells with sound-reflecting probes
Key to this innovation in ultrasound imaging—a method called Nonlinear sound sheet microscopy—was the discovery of a sound-reflecting probe made in the Shapiro Lab at Caltech. Heiles: "This probe is a nanoscale gas-filled vesicle that lights up in ultrasound images, making cells visible. These vesicles have a protein shell and we can engineer them to tune their brightness in images. We used these gas vesicles to track cancer cells."
Brain imaging
In addition to revealing cells, the team used ultrasound and microbubbles as probes circulating in the blood stream to detect brain capillaries. Heiles: "To our knowledge, nonlinear sound sheet microscopy is the first technique capable of observing capillaries in living brains. This breakthrough has tremendous potential to diagnose small vessel diseases in patients." Since microbubble probes are already approved for human use, this technique could be deployed in hospitals in a few years.
Potential for cancer research
Beyond clinical practice, sound-sheet microscopy can greatly benefit biological research and the development of new cancer treatments in particular, according to Maresca. "Our imaging technique can distinguish healthy versus cancer tissue. Furthermore, it can visualise the necrotic core of a tumour; the centre of the tumour where cells start dying due to a lack of oxygen. Thus, it could assist in monitoring the progression of cancer and the response to treatment."
