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Multi-Photon Laser Ablation of Cytoplasmic Microtubule Organizing Centers in Mouse Oocytes
Multi-Photon Laser Ablation of Cytoplasmic Microtubule Organizing Centers in Mouse Oocytes
JoVE Journal
Developmental Biology
This content is Free Access.
JoVE Journal Developmental Biology
Multi-Photon Laser Ablation of Cytoplasmic Microtubule Organizing Centers in Mouse Oocytes

Multi-Photon Laser Ablation of Cytoplasmic Microtubule Organizing Centers in Mouse Oocytes

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08:24 min

November 11, 2022

DOI:

08:24 min
November 11, 2022

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Transcript

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This protocol offers an excellent strategy to selectively ablate cytoplasmic MTOCs, which are critical regulators for spindle positioning in mouse oocytes.Therefore contributing to our understanding of molecular mechanisms that regulates oocyte meiosis.This technique allows to perform targeted ablations within live oocytes with high spatial accuracy and with minimal phototoxic effects on the cell.This protocol can be used to induce a selective mechanical perturbation of different instructors without altering the cell viability.This protocol requires the technical skills to ablate all cytoplasmic MTOCs in the shortest possible time to avoid compromising the oocytes’viability.So practicing this technique several times before carrying out the actual experiment is essential.To begin supplement the CZB medium with glutamine and milrinone and place it in the incubator.Prepare the collection and culture dishes.Make four micro drops of collection medium and two micro drops of culture medium in 60 and 35 mm Petri dishes, respectively.And cover them with mineral oil.Keep the collection dish on the slide warmer, and incubate at 37

Summary

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An optimized protocol is presented that enables the depletion of cytoplasmic microtubule organizing centers in mouse oocytes during metaphase I using a near-infrared femtosecond laser.

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