Overoxidized iron hematoxylin is a contributing factor to pale nuclear staining in Masson Trichrome. Which option identifies this cause?

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Nuclear staining intensity in Masson Trichrome comes from the iron hematoxylin complex that colors nuclei. When iron hematoxylin is overoxidized, the dye–mordant complex becomes unstable and loses color, so nuclei appear pale rather than the characteristic dark blue-black. The overoxidation essentially diminishes the staining power of the nuclear stain, leading to faint nuclear detail. Other scenarios, like omitting iron hematoxylin or staining time issues, can also affect intensity, but the specific cause described here is the overoxidation of the iron hematoxylin itself.