An Adapted Protocol for Quantitative Rhizosphere Acidification Assay

Acidification of the rhizosphere is a key process in the homeostasis of multiple essential nutrients, including iron. Under iron deficiency, the release of protons from the roots helps solubilize and increase the accessibility of iron in the soil. Rhizosphere acidification has been widely examined in many iron homeostasis studies, generally using a qualitative method based on the color change of bromocresol purple, a pH indicator dye, near the roots. In this chapter, we introduce an adapted version of a rhizosphere acidification assay protocol that allows for the quantitative assessment of small pH changes in the rhizosphere. This colorimetric method also utilizes bromocresol purple, but the ratio of its absorbance at 434 nm and 588 nm is considered to quantify protons released into the assay solution. Furthermore, the assay is compatible with small sample volumes, such as those with young Arabidopsis seedlings.