Naslov
Dissection and characterization of microtubule bundles in the mitotic spindle using femtosecond laser ablation

Autori
Buđa, Renata ; Vukušić, Kruno ; Tolić, Iva Marija

Vrsta, podvrsta i kategorija rada
Poglavlja u knjigama, znanstveni

Knjiga
Cell Polarity and Morphogenesis / Methods in Cell Biology

Urednik/ci
Lecuti, Thomas

Izdavač
Elsevier

Godina
2017

Raspon stranica
81-101

ISBN
978-0-12-809373-3

ISSN
0091-679X

Ključne riječi
bridging fiber ; confocal laser scanning microscopy ; forces ; k-fiber ; laser cutting ; metaphase ; microtubules ; mitosis ; swept field confocal microscopy

Sažetak
The mitotic spindle is a highly organized and dynamic structure required for segregation of the genetic material into two daughter cells. Although most of the individual players involved in building the spindle have been characterized in vitro, a general understanding of how all of the spindle players act together in vivo is still missing. Hence, in recent years, experiments have focused on introducing mechanical perturbations of the spindle on a micron scale, thereby providing insight into its function and organization, as well as into forces acting in the spindle. Among different types of mechanical perturbations, optical ones are more flexible, less invasive, and more precise than other approaches. In this chapter, we describe a detailed protocol for cutting the microtubule bundles in human cells using a near-infrared femtosecond laser. This type of laser microsurgery provides the ability to precisely sever a single microtubule bundle while preserving spindle integrity and dynamics. Furthermore, we describe quantitative measurements obtained from the response of a severed microtubule bundle to laser ablation, which reveal the structure and function of individual parts of the spindle, such as the bridging fiber connecting sister k-fibers. Finally, the method described here can be easily combined with other quantitative techniques to address the complexity of the spindle.

Izvorni jezik
Engleski

Znanstvena područja
Biologija

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