© GEORGE RETSECK
Analyzing the molecular constituents of a homogenized piece of animal or plant tissue may provide clues about the nature of the component cells’ functions. But it may also obscure striking functional differences between individual cells.
Not surprisingly, then, the study of single cells is a burgeoning area of research. While single-cell genomics and transcriptomics have benefited from DNA’s and RNA’s inherent ability to be amplified, single-cell proteomics and metabolomics must rely solely on miniscule quantities of starting material.
Researchers have thus been developing a variety of techniques for tackling such tiny volumes. The teams...
With the aid of a microscope, Masujima uses the micromanipulator to direct a tiny capillary needle to a stabilized piece of plant tissue. He then pierces an individual cell and sucks up its contents. The few hundred femtoliters of cell innards are mixed with an ionization solvent and zapped with high voltage electricity, which shoots the material into a mass spectrometer for immediate analysis.
The approach, which could be used for plant or animal cells, yields hundreds to thousands of molecular signals (spectral peaks) per cell. However, says Sixue Chen of the University of Florida, the big challenge in mass spectrometry is determining the identities of the molecules responsible for those peaks—the majority of which are unknown. Nonetheless, he says, the publication of Masujima’s extensive protocol “is very useful to the research community to move this type of analysis forward.” (Nature Protocols, 10:1445-56, 2015)
MASS-SPEC TECHNIQUE FOR SINGLE-CELL ANALYSIS
|HOW IT WORKS
Matrix-assisted laser desorption/ionization (MALDI)
Single cells are mixed with a matrix material, applied to a metal plate, and irradiated. The resulting vapor is then ionized and shot into a mass spectrometer.
No. Pretreatment with matrix destroys cells.
A wide range, from small molecules to macromolecules such as proteins
Live MS nano-
|Individual cell contents are aspirated, mixed with ionizing solution, and sprayed into a mass spectrometer.
|At the point of sampling, yes. However, cell does not survive aspiration.
|Mainly small molecules (e.g., metabolites, amino acids, sugars, lipids).
|No. Contents only, avoiding noise from membrane lipids