Enzyme-Linked Immunoabsorbant Assay (ELISA)

Rhizobium and Legume Root Nodulation
Leguminous Plants
The Discovery of Symbiotic Nitrogen Fixation
Rhizobium Classification
Cultural Characteristics
Genetics
Flavonoids Induce Nodualation
Novel Approaches to Extend Root Nodulation
Ecology
Infection
Lectins
Structure of the Nodule
Function of the Nodule
Nitrogenase Hydrogenase Interrelationship
Stem Nodulating Legumes
Factors Affecting Nodulation
Temperature and Light
Hydrogen Ion Concentration
Mineral Nutrition

Enzyme-Linked Immunoabsorbant Assay (ELISA)

	Enzyme-Linked Immunoabsorbant Assay (ELISA) Currently ELISA is being used for 'nodule typing' to determine the 'nodule occupancy' of a selected strain introduced to seed surface or soil. For this purpose, nodules can be oven-dried and stored. The technique is particularly useful for field testing of mixed infections in nodules without the use of microscopes, facilitated by easily transportable ELISA kits.
There have been improvements to the technique by the use of fluorescent substrate and monoclonal antibodies. The method uses the earlier stated procedure for developing the required antisera in rabbits followed by bleeding and purifying the antisera. The antisera are then conjugated with alkaline phosphatase enzyme. The root system of the desired nodulated plant is gently pulled out of soil, washed, the nodules numbered and individually squashed in the wells of a microtitre plate. The plate is then incubated after the addition of the antisera enzyme conjugate in a substrate and colour developed with o-nitrophenol phosphate.
The traditional methods so far described which relate to antigen-antibody reactions and resistance to known antibiotics are expensive, labour intensive and lack sufficient specificity to identify strains of rhizobia in soil without doubt. There are however, some newer methods based on molecular biology that have advantages over earlier methods. These methods include the introduction of marker genes and identification of DNA sequences that are characteristic of particular strains.

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