Auxin Immunolocalization in Plant Tissues

The major naturally occurring auxin, indol-3 acetic acid (IAA), coordinates many growth and differentiation processes by modulating gene expression during plant development. The sites of IAA biosynthesis and its polar transport (PAT) routes determine auxin accumulation and distribution during growth. From many studies on the model plant Arabidopsis thaliana over the last years, it has become evident that the expression and sub-cellular localization of multiple transport proteins are required to initiate and maintain directional auxin flows within plant organs and tissues, creating the auxin concentration gradients that regulate plant development. For this reason, the understanding of auxin dependent pattern formation also relies on the possibility to directly visualize auxin concentration and distribution in the tissues. The production and isolation of antibodies highly specific for IAA provide the means to detect and localize free IAA in different plant species during tissues differentiation and organ development. The immunolocalization protocol presented here uses a monoclonal anti-IAA specific antibody that can be used to visualize auxin accumulation in different organs and tissues during plant development. We successfully used this protocol to determine IAA maxima during kernel and inflorescence development in maize, highlighting also alterations in auxin accumulation patterns in a mutant with reduced auxin accumulation capacity and in plants treated with an inhibitor of auxin transport.