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Updated: Oct 1, 2026
Measurement of Microtubule Dynamics by Spinning Disk Microscopy in Monopolar Mitotic Spindles
Published on: November 15, 2019
Determination of the mitotic index by microinjection of fluorescently labelled tubulin
Sandra L Schwindling1, Michael Faust, Mathias Montenarh
1Medical Biochemistry and Molecular Biology, University of the Saarland, Homburg, Germany.
Abstract:
The microneedle injection technique is one of the most established procedures for the introduction of proteins into living cells. To analyse injected proteins which are important in cell cycle progression it is often necessary to determine the mitotic index. Measuring the mitotic index after microinjection is complicated because only a limited number of cells of the whole cell population is microinjected. Therefore, we attempted to establish a new method to determine the mitotic index using microinjection of fluorescently labelled alpha/beta-tubulin into mammalian cells which allows to monitor the injected cells simultaneously with the determination of the mitotic index. We demonstrated that fluorescently labelled tubulin incorporates efficiently into the mitotic spindle apparatus. Fluorescence remains stable for several hours which is sufficient to observe the progression of cells through the M-phase of the cell cycle. The determination of the mitotic index with the method presented here gave similar results to those determined using other methods. With this method also different stages of mitosis can be visualized by analysing various steps of spindle formation. Thus, this rapid method allows the monitoring of the injected cells after microneedle injection and simultaneously the determination of the mitotic index.
Insights
Researchers developed a new method for determining the mitotic index in microinjected cells. This technique uses fluorescently labeled tubulin to simultaneously track injected cells and monitor cell cycle progression through mitosis.
Area of Science:
- Cell Biology
- Molecular Biology
- Biotechnology
Background:
- Microneedle injection is a standard method for introducing proteins into cells.
- Determining the mitotic index is crucial for analyzing cell cycle progression after microinjection.
- Current methods face challenges due to the limited number of microinjected cells.
Purpose of the Study:
- To establish a novel method for determining the mitotic index in microinjected mammalian cells.
- To enable simultaneous monitoring of injected cells and mitotic index determination.
- To improve the analysis of proteins involved in cell cycle progression.
Main Methods:
- Microinjection of fluorescently labeled alpha/beta-tubulin into mammalian cells.
- Utilizing the fluorescently labeled tubulin to visualize the mitotic spindle apparatus.
- Observing cellular progression through the M-phase of the cell cycle.
Main Results:
- Fluorescently labeled tubulin efficiently incorporates into the mitotic spindle.
- Stable fluorescence allows for several hours of observation during M-phase.
- The new method yields results comparable to established methods for mitotic index determination.
- Visualization of different mitotic stages through spindle formation analysis is possible.
Conclusions:
- The developed method provides a rapid and effective way to monitor microinjected cells.
- Simultaneous determination of the mitotic index is achieved, simplifying cell cycle analysis.
- This technique enhances the study of proteins affecting cell cycle progression via microinjection.
