The quantitation of ferritin in the seeds of staple crops: ELISA method development

Abstract

Biofortification, breeding of crops rich in bioavailable micronutrients, is a low‐cost sustainable strategy for combating iron deficiency anemia (IDA), a most common nutritional deficiency. Plant ferritin is believed to provide highly bioavailable iron, however a method to measure ferritin in seeds is currently lacking. The objective of this study was to develop a simple, reliable assay to quantitate the ferritin in several staple crops. A highly immunogenic 22 amino acid common sequence was identified among the ferritin sequences from beans, wheat, maize and rice to produce both anti‐ferritin polyclonal and monoclonal antibodies. A rapid method for the crude extraction of ferritin from the seeds was developed. Both antibodies shown cross reactivity with ferritin from all seeds tested; however the degree of reactivity was found to be dependent on the type or the amount of total protein extracted from each seed. Using our developed indirect ELISA assay with the polyclonal antibody and recombinant pea ferritin standard, ferritin content was found to be 10.2±1.0, 4.38±0.9, 1.2±0.3, 0.38±0.1, and 0.04±0.01 μg/g in red beans, white beans, wheat, maize, and brown rice, respectively. This simple and reliable assay will facilitate the rapid screening of large numbers of seeds and can be used by plant breeders to breed ferritin‐rich crops for providing highly bioavailable iron foods to combat IDA.