Establishment of a reliable protocol for the quantification of an oxidative burst in suspension-cultured wheat cells upon elicitation

Spent growth medium of Pantoea agglomerans primes wheat suspension cells for augmented accumulation of hydrogen peroxide and enhanced peroxidase activity upon elicitation

“…The generation of H 2 O 2 upon addition of elicitor-active substances (a mixture of chitosan polymers or chitin hexamer) was measured by luminol-dependent chemiluminescence [200 l aliquot of the cell suspension; 600 l of 50 mM potassium phosphate buVer, pH 7.9; 100 l of 1.21 mM luminol in the above potassium phosphate buVer; 100 l of 14 mM potassium hexacyanoferrate(III); 10 s-integration time; 430 nm] according to Warm and Laties (1982) as described previously (Ortmann et al 2004). The maximum concentrations of H 2 O 2 produced by the wheat cells using the same concentration of elicitor varied over longer periods of time, most likely as a consequence of diVerent vitality of the cells (Ortmann et al 2004).…”

“…The maximum concentrations of H 2 O 2 produced by the wheat cells using the same concentration of elicitor varied over longer periods of time, most likely as a consequence of diVerent vitality of the cells (Ortmann et al 2004). Therefore, the appropriate concentration of elicitor was determined for each experiment in a test sample, and a positive control (elicitation of naive control cells) was always performed.…”

“…This rapid release of H 2 O 2 after treatment with pathogens or pathogen-derived elicitors has been described as a key event in the orchestration of various cellular defense responses (Levine et al 1994). We have established recently a protocol, allowing for the Wrst time the reliable quantiWcation of H 2 O 2 generated during an oxidative burst in monocot cells (Ortmann et al 2004). We chose chitin hexamer and partially acetylated chitosan polymers as known elicitors of disease resistance mechanisms in wheat (Vander et al 1998), namely a rapid and transient oxidative burst (Ortmann et al 2004) and increased extracellular peroxidase activity (Gotthardt and Grambow 1992).…”

“…We have established recently a protocol, allowing for the Wrst time the reliable quantiWcation of H 2 O 2 generated during an oxidative burst in monocot cells (Ortmann et al 2004). We chose chitin hexamer and partially acetylated chitosan polymers as known elicitors of disease resistance mechanisms in wheat (Vander et al 1998), namely a rapid and transient oxidative burst (Ortmann et al 2004) and increased extracellular peroxidase activity (Gotthardt and Grambow 1992). To our knowledge, the observed eVect of the EPS is the Wrst example of priming by a speciWc component of bacterial origin in a monocot plant.…”

Development of a Colorimetric Assay for Determining the Amount of H2O2Generated in Tobacco Cells in Response to Elicitors and Its Application to Study of the Structure-Activity Relationship of Flagellin-Derived Peptides

“…16) Various methods have been used to measure the amount of H 2 O 2 generated in plant cells during an oxidative burst. The luminol-dependent chemiluminescence method is often used for such measurements, [17][18][19] but although highly sensitive, it is inconvenient in that it requires special equipment, a luminometer, to monitor the reactions. A fluorometric assay utilizing scopoletin has also been used to measure H 2 O 2 .…”