Optical slice through a Medaka embryo (Nat. Methods 4, 311–313; 2007).

Biological systems are three-dimensional. Cells are rarely present as isolated monolayers, as in the Petri dish. However, there are technical challenges inherent to in vivo studies, and this is particularly true for imaging.

Biological tissue is opaque and scatters light effectively, and image quality deteriorates rapidly in thicker samples. Conventional confocal microscopy, for instance, cannot image deeper than a few hundred microns into a sample; a fruitfly larva, a mammalian embryo or a tissue slice may be several times thicker. Also, it is often desirable to image such samples in toto, to obtain a global view of tissue organization and gene expression; this too is challenging with conventional approaches. And although techniques such as magnetic resonance imaging (MRI) can image deep into biological samples, the images are low in resolution.

A gap therefore exists between high-resolution methods such as confocal microscopy and low-resolution, in vivo methods such as MRI. In recent years, several optical approaches have begun to fill this gap.

Light sheet microscopy, a classical method recently updated, can image a fluorescent signal at a depth of a few millimeters within a sample and has been usefully applied to small organisms and embryos. In the past year, a modified version of the approach was used to comprehensively image the cells, numbering in the thousands, in a zebrafish embryo during 24 hours of development (Science 322, 1065–1069; 2008).

Tomographic methods such as optical projection tomography (OPT), by contrast, rely on imaging the sample from many different angles and then using mathematical models to recalculate the original 3D information. OPT can image deeper into a sample (up to 10 mm), and it has been combined this past year with in vitro organ culture to image tissue movements and gene expression in the developing mouse limb bud (Nat. Methods 5, 609–612; 2008).

Although these and other volumetric approaches have yet to be taken up widely, this could change as the methods improve and as their capabilities become more commonly appreciated. The possibility of peering deep into organisms and watching biological processes unfold in 3D should be strong motivation.