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Open-source tool allows accurate measurement of 3D subcellular geometry

Nov 17, 2021 // Researchers develop software to consistently quantify spindle and chromatin morphology in 3D fluorescent data

Size and shape are important biological features. Morphometrics, the study of size and shape, have classically been performed on the level of organisms, tissues, and cells. Now, the continuous improvement of imaging techniques and data analysis allows for the accurate measurement of structures inside cells.

The function of cellular structures (organelles) is dependent on their geometry and proportionality to cell size. That´s why researchers are more and more interested in deciphering biological scaling laws at the micrometer level. „The mitotic spindle is a good example why length and shape of intracellular organelles matter.“, says Tobias Kletter, a PhD student and molecular biologist in the Reber Lab at IRI Life Sciences and first author of a new study, published today in Journal of Cell Biology.

The spindle is a short-lived organelle that cells construct when they divide. Tobias and his colleagues have searched for a way to retrieve precise data of the length, width and volume of the spindle and then to put these parameters in relation to the dividing cell.

Why is it so important to determine the size of the spindle exactly?

“During cell division, the job of the spindle is to distribute the duplicated chromosomes among the daughter cells,” Tobias explains. “Think of the spindle as a complex machine. It first unites all chromosomes before it pulls at them from opposing directions. Eventually, each chromosome splits into two copies and the spindle further moves these copies apart. The dividing cell takes great care that both daughter cells receive exactly one copy each.

If spindle size and shape are not set properly, this might lead to errors in the segregation process, which can have grave outcomes for the cell or, if it occurs early in development, for the entire embryo. Ultimately, we want to understand how spindle size and shape are regulated at a molecular level. First, however, we need reliable tools that allow us to quantify spindle size and shape at the cellular level.”

“Mitotic spindle width is a robust indicator of spindle volume”

The researchers have started to investigate how size and number of subcellular structures such as the spindle relate to cell size, a phenomenon known as "organelle scaling ". But what are the relevant morphometric measurements to precisely formulate spindle scaling phenomena?

To tackle this question, Tobias collaborated closely with image analysis expert Dr. Christian Tischer of the European Molecular Biology Laboratory (EMBL). Over the last two years, their cooperation resulted in the development of an open-source, user-friendly pipeline which they named "Spindle3D". It allows for the quantitative, consistent, and automated analysis of 3D fluorescent images of spindles and chromatin.

The researchers systematically analysed spindles in different cell types, including cancer cells, stem cells and one-cell embryos to derive geometric relations of spindle, chromatin, and cells. Taken together, “by measuring spindle width, we found that we can predict many other cellular size parameters. Tell me the width of the spindle and I’ll tell you the spindle’s mass and volume with great confidence. And I’ll predict the volume of the dividing cell that constructed the spindle, too. All of these features show a tight and interwoven geometrical relationship”, says Tobias. “However, we were surprised to see that, in comparison, the length of the spindle was correlating much less with the size of the cell or the dimensions of the chromosome plate. Still, many studies exclusively rely on spindle length as a proxy for spindle size. Our findings emphasize the need for proper 3D analysis.”

The analysis pipeline performs well on diverse data sets

According to Tobias, the biggest challenge in the development of Spindle3D was to design the algorithm to be as general as possible. The project was initially tailored to a specific cell type. To make it accessible for more researchers, the question arose whether this specific application could also be applied to other data sets. And in the best case, even to completely different cells or organisms.

The publication demonstrates that the analysis pipeline performs well on diverse data sets, including cow embryos and kangaroo rat cells. To get new insights, Tobias is counting on testimonials from his peers. “The project is a living project”, he says, “and we hope for a lot of feedback and suggestions from the community to be able to improve the software.”

Original publication

Tobias Kletter, Sebastian Reusch, Nils Dempewolf, Christian Tischer, Simone Reber. Volumetric morphometry reveals spindle width as the best predictor of mammalian spindle scaling. Journal of Cell Biology (2021). DOI: 10.1083/jcb.202106170

Further information

  • Sound of Science – The Science Podcast for Everyone

    Anke Wagner talks to Tobias Kletter (member of the Reber Lab) about challenges in research and the development of the Spindle3D software.


For further information and other requests please contact Tobias Kletter from the Reber Lab via email tobias.kletter(at)