Development of a method for the detection and quantification of total chitinase activity by digital analysis.
A method for the quantitative assessment of chitinase activity from crude plant extracts has been developed. Dilution series of commercial chitinase extracts and whole protein extracts from plants expressing Systemic Acquired Resistance (SAR) were assayed using this method. Using glycochitin as enzyme substrate, the activity assay is based on the affinity of fluorescent brightener 28 with undigested glycochitin. An agarose plate supports the substrate and the developed reaction plate is viewed under UV translumination. Digital analysis revealed that the chitinase activity measured using this method was found reproducible and reliable. Most importantly, it is fast and allows analysis of large number of samples with minimum effort.
Velasquez
Luis
L
Department of Plant Pathology, Michigan State University, East Lansing, MI 48824, USA.
Hammerschmidt
Ray
R
D001557:Benzenesulfonates / Q000737:chemistry*; D002686:Chitin / Q000031:analogs & derivatives* / Q000378:metabolism*; D002688:Chitinases / Q000032:analysis* / Q000302:isolation & purification / Q000378:metabolism; D018553:Cucumis sativus / Q000201:enzymology*; D007091:Image Processing, Computer-Assisted; D005779:Immunodiffusion / Q000379:methods*; D018515:Plant Leaves / Q000201:enzymology
D001557:Benzenesulfonates; D002686:Chitin; C007061:C.I. Fluorescent Brightening Agent 28; C031610:glycolchitin; D002688:Chitinases