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Ferrous iron [Fe(II)] analysis by ferrozine assay

WB Wei Bai
TR Tahina Onina Ranaivoarisoa
RS Rajesh Singh
KR Karthikeyan Rengasamy
AB Arpita Bose
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The Fe(II) concentration measurement was done using 10 μL of culture mixed with 90 μL 1 M HCl in a 96-well plate inside the anaerobic chamber (filled with 5% H2/75% N2/20% CO2, Coy Laboratory, Grass Lake, MI). After the plate was removed from the anaerobic chamber, 100 μL of ferrozine (0.1% (w/v) ferrozine in 50% ammonium acetate) was added to the sample. Then the 96-well plate was covered with foil and incubated at room temperature for 10 min. before the absorbance was measured at 562 nm. The absorbance was then converted to Fe(II) concentration based on a standard curve generated by measuring the absorbance from 0 mM, 1 mM, 2.5 mM and, 5 mM Fe(II).

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