Between May 1508 and the summer of 1511, he completed the "entrance half" of the Sistine chapel and ended this stage by painting the Creation of Eve and the scenes flanking this central panel.

The ignudi are young, nude males that Michelangelo painted as supporting figures at each corner of the five smaller narrative scenes along the center of the ceiling.

In the earliest frescoes painted by Michelangelo toward the entrance of the Sistine chapel, the ignudi are paired, and their poses are similar but with minor variations.

At the center of the composition, God is shown in contrapposto rising into the sky, with arms outstretched separating the light from the darkness.

For instance, Giorgio Vasari (1511–1574), Michelangelo's student and biographer, wrote in 1550:"Furthermore, to demonstrate the perfection of his art and the greatness of God, Michelangelo depicted God dividing the light from darkness in these scenes, where He is seen in all His majesty as He sustains himself alone with open arms with the demonstration of love and creative energy.

In an article published in the journal Neurosurgery in May 2010, Ian Suk, a medical illustrator, and Rafael Tamargo, a neurosurgeon, both from the Johns Hopkins University School of Medicine, suggested that Michelangelo concealed three neuroanatomical images in the Separation of Light from Darkness.

[6][7] Suk and Tamargo explained that Michelangelo started to dissect cadavers at the age of 17–19 years and continued his anatomical studies throughout his life.

Of note is that in an article published in the Journal of the American Medical Association in October 1990, Frank Meshberger, an obstetrician-gynecologist from Indiana, explained that Michelangelo similarly concealed an image of the brain in the shroud surrounding God in the Creation of Adam.

Alternatively, the concealed anatomical image in God's neck in the Separation of Light from Darkness has been interpreted as a goiter (an enlarged thyroid gland).

Along these lines, Gilson Barreto and Marcelo G. de Oliveira, in a book published in 2004,[11] analyzed the Separation of Light from Darkness and noted that God's upper chest and outstretched arms define a "U" shape, which they suggested is reminiscent of the hyoid bone, although they do not offer a thematic explanation for this structure in the fresco.