“This will allow us to study all sorts of fundamental biological processes as a function of cell size,” says Daniel Needleman, a physicist and bioengineer at the University of California, Berkeley, and co-leader of the research team. “Now that we have the ability to make these measurements, we can actually ask: How variable is the growth of a cell? How sensitive is it to perturbations? How does growth depend on the nutrients or the environment that the cell is in? What happens to growth as cells become cancerous and stop responding to normal growth signals?”
“This is really a technical milestone in the field of single-cell biology,” adds Nevan Krogan, a quantitative biologist at the University of California, San Francisco (UCSF) and co-leader of the research team. “It will be transformative for the entire community, opening new possibilities for studying fundamental biology and disease mechanisms at the single-cell level.”
Needleman and Krogan are co-senior authors of a study describing the platform and its initial results, published today (May 12, 2022) in the journal Cell. While a handful of groups have measured the mass of populations of cells before, this group developed the first platform to weigh single cells in real-time as they grow.
They found that the growth rate of an individual cell is constant; that is, its mass increases steadily over time. Interestingly, this means that the metabolic rate of a cell per unit mass is decreasing as it grows. In other words, a smaller cell is more efficient at converting energy from its environment into growth than a larger cell. Moreover, the researchers showed that their methods could be used to measure the efficiency with which cells take in and convert external nutrients into growth.
“As a quantitative biologist, I've become passionate about using precise, quantitative approaches to study problems that were, until recently, too challenging or impossible to measure. To make a contribution, you have to build these new measuring tools,” says Krogan. “This effort required us to develop new experimental and computational approaches and bring together scientists with different backgrounds. It would not have been possible had we worked in isolation.”
Weighing the unweighable
The new platform—dubbed microfluidic weighing—combines microfluidics, which enables the precise manipulation of fluids at the submillimeter scale, with quantitative phase imaging, a relatively new microscopy technique that directly measures the mass of an object based on how much it bends light.
“The first technical challenge is simply manipulating and capturing cells,” explains Daniel Fletcher, a bioengineer at UC Berkeley and a co-author of the study, whose lab developed the microfluidic platform. “You don’t want hundreds of thousands of cells running through your system, because then you don't know which cell you’re measuring. But you also don't want to measure one cell at a time, because that would take too long. So, we trap tens or hundreds of cells at a time and flow media over them so that they're getting the nutrients they need to survive, but they stay trapped there. Then, the imaging team came in to optimize and implement quantitative phase imaging.”
To achieve quantitative phase imaging, the researchers shone a beam of light through a microchannel and onto the cells, capturing an image of the light as it emerged on the other side. If there were no cell in the channel, the wavefront of the light would be undisturbed. But when a cell is present, the light bends, slightly altering the wavefront. This change in wavefront can be computationally converted directly into the mass of the cell.
“By measuring the phase shift of the light as it passes through a cell, we infer the local index of refraction of the material, which is directly related to the cell's density,” explains study co-author Aydogan Ozcan, a professor of electrical engineering and computer science and director of the Integrated Optics Lab at UCLA. “Since we know the chemical composition of the cell and the density of its components, this enables us to precisely determine the cell’s mass.”
“These measurements are really sensitive,” says Needleman. “We can measure changes in the mass of a single cell corresponding to less than 1,000 water molecules being added to the cell.”
As cells in the microfluidic chamber absorbed nutrients from their surroundings, they expanded and grew in mass, as expected.
“But we noticed that the growth rate did not change as the cells became bigger,” says Needleman. “This means that the metabolic engine inside a small cell is actually more efficient at converting energy into growth than the engine of a larger cell.”
The team hopes that other scientists will adopt and further refine their technology to study the growth of many different types of cells under various conditions and environments, including disease conditions.
