 Kinetochore ablation experiment

In 1995, Conly L. Rieder, Richard W. Cole, Alexey Khodjakov and Greenfield Sluder tested whether an unattached kinetochore supplies the inhibitory signal delaying anaphase. Living PtK1 biological cells on glass coverslips were mounted in Rose chambers at $35$–$37{}^{\circ }\mathrm{C}$. An inverted microscope with a $60\times$ objective recorded differential interference contrast microscopy video. A pulsed $532\mathrm{nm}$ laser focused through that objective destroyed selected chromosome regions. The readout was chromosome movement and anaphase onset before and after surgery.^[ref]

Destroying the last unattached kinetochore permitted anaphase without that chromosome aligning. Cutting chromosome arms served as a damage control: progression still awaited attachment of the last naturally one-pole-attached chromosome. Correlative electron microscopy checked the absence of the targeted kinetochore. Together these results located an inhibitory activity in or near the unattached kinetochore, rather than establishing a requirement for alignment itself.^[ref]

This intervention localized control; it did not identify every signaling molecule or guarantee faithful inheritance after damage. The cultured PtK1 preparation differs from the hypothetical $2n=4$ cartoon. Neither the cartoon nor this experiment establishes identical control in every species.