Amperometric Biosensor Approaches for Quantification of Indole 3-Acetic Acid in Plant Stress Responses

Abstract

Amperometric biosensors are known to be sensitive, reliable, and inexpensive instruments for biomolecular detection. Recently, self-referencing amperometric biosensors have been utilized to quantify the endogenous apoplastic flux of plant hormone indole 3-acetic acid (IAA) in vivo. There is still a significant need for sampling and testing methods to measure IAA concentrations in whole tissue samples. In the present study we used nanomaterial-modified platinum microelectrodes for the detection of IAA extracted from whole plant tissues. The key to the use of the nanomaterials was to enhance the surface area and thus the limit of detection for IAA. The nanoscale electrochemical interface was modified by the application of a layer of platinum black, followed by silanization overnight and a coating of multiwalled carbon nanotubes. Electroanalytical characterization of sensor performance was evaluated using both IAA standards and IAA extracted from corn plant samples, using a three-electrode scheme (reference, sensing, and auxillary). We compared tissue concentrations in water- and salt-stressed corn seedlings and compared these results to values measured using established enzyme-linked immunosorbent assay (ELISA) protocols. We found that the values obtained from both methods were comparable. The data obtained from the IAA sensor suggested that the electroanalytical biosensor approach was slightly more reliable and sensitive. Our results demonstrate a novel nanomaterial biosensor approach for IAA quantification in plant tissue extracts. The results of the stress study clearly indicated that root and shoot elongation and growth had significant positive correlation with IAA content in the raw plant tissue extracts. Water and salt stress both reduced root and shoot growth, which may be due to overaccumulation of IAA, resulting in inhibition of elongation. In comparison to established methods, such as high-performance liquid chromatography (HPLC) or ELISA, our approach is simple and inexpensive. Unlike HPLC, our approach does not require any elaborate sample purification, nor does it require antibody development as needed for ELISA. In summary, this electroanalytical biosensor method can be effectively utilized for simple, cheap, and reliable detection of IAA extracted from plant samples.