Stanford Cancer Institute News

Fall 2018

Image courtesy of Garry Nolan, PhD

Visualizing more proteins through CODEX is a huge boon to research.

How Do You See the Inner Working of a Cancer Cell?

Stanford’s Cell Sciences Imaging Facility

Inside a tumor, molecules zip from place to place, and cluster in unusual areas, helping cancer cells multiply at a speedy pace. To stop tumors from growing, scientists often must aim to put the brakes on these molecules. Doing so requires visualizing the molecules and cells in the first place. Luckily for cancer researchers, modern microscopy has lots to offer them “It’s a very exciting time in imaging,” says Jon Mulholland, Director of the Stanford Cell Sciences Imaging Facility (CSIF).

At the CSIF, Stanford researchers can use state-of-the-art light and electron microscopes, get training on the microscopes, and get advice and assistance on how to prepare their samples and interpret their data. “Most biology labs have a light microscope,” says Mulholland. “What we offer is very high-end imaging that lets researchers do more unique and advanced things.”

Mulholland says that SCI faculty often rely on the central microscopy resource to study the very basic biology of cancer— if there’s a protein suspected of playing a role in cancer, for instance, they might use confocal or superresolution fluorescence microscopy to see exactly where in cancer cells the protein localizes and what other proteins it interacts with and when.

These questions are integral to how a cancer grows and spreads in the body.

Stanford scientists in the lab of SCI member Garry Nolan, PhD, developed the CODEX (co-detection by indexing) approach to be able to fluorescently label and visualize around 50 different proteins in a single tissue sample. In most classic fluorescence experiments, researchers are limited to seeing about five distinct proteins at once, each labeled with a different color fluorescent tag. For understanding cancer cells—which can have dozens of molecular players interacting in many ways— this leap to being able to see more proteins is a huge boon to research.

In CODEX, a tissue slice is labelled with up to 50 antibodies, each of which binds to a specific individual protein or other cellular target. Each antibody also has a short, unique DNA segment that acts as its barcode. Then, one of three fluorescent colors—green, orange or red—is assigned to each barcode.

Using iterative cycles, each revealing the locations of three proteins, researchers can work their way through all 50 targets to reveal their locations in a sample. Between each round of probing and visualization, the probes are washed away and a new set of three fluorescent-tagged complementary DNA tags is applied.

The whole CODEX system is engineered to fit inside a small microfluidics device that sits on the microscope— that means the sample
never shifts during sequential rounds, and each picture showing three probes can ultimately be combined into one image that shows many more proteins throughout the tissue.

“Since you can see where various classes of cells are simultaneously positioned, you can start to make hypotheses about interactions between those cells, and whole networks of cells,” says SCI member Peter Jackson, PhD, professor of microbiology and immunology, who has worked with CODEX in his own lab. This gives researchers insight into what molecular markers on cells might be useful to target with anti-cancer drugs. It also lets them compare how different classes of cancer cells change— or disappear—after different treatments, which can shed light on why some drugs work better for some patients than others. The setup for CODEX is expected to be available in the CSIF by the end of 2018, thanks to funding from multiple sources, including the SCI.

“It’s a really nice synergistic story about it all coming together,” says Mulholland.