Metaphase is the third phase of mitosis, the process that separates duplicated genetic material carried in the nucleus of a parent cell into two identical daughter cells. During metaphase, the cells chromosomes align themselves in the middle of the cell through a type of cellular "tug of war". Prior to metaphase, protein formations called kinetochores formed around the centromere. Long protein filaments called kinetochore microtubules extended from poles on either end of the cell and attached to the kinetochores. During metaphase, the kinetochore microtubules pull the sister chromatids back and forth until they align along the equator of the cell, called the equatorial plane.

There is an important checkpoint in the middle of mitosis, called the metaphase checkpoint, during which the cell ensures that it is ready to divide. Once the cell has established that all of the chromosomes are properly aligned and that the kinetochores are correctly attached, the cell enters the fourth phase of mitosis, known as anaphase.

In summary, during metaphase of mitosis, the chromosomes align themselves in the middle of the cell through the action of kinetochore microtubules, and the cell checks that all chromosomes are properly aligned before proceeding to anaphase.