A high-throughput method for the quantification of iron saturation in lactoferrin preparations |
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Authors: | Grzegorz Majka Klaudyna ?piewak Katarzyna Kurpiewska Piotr Heczko Gra?yna Stochel Magdalena Strus Ma?gorzata Brindell |
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Institution: | 1. Department of Inorganic Chemistry, Faculty of Chemistry, Jagiellonian University, Ingardena 3, 30-060, Krakow, Poland 2. Chair of Microbiology, Jagiellonian University Medical College, Czysta 18, 31-121, Krakow, Poland 3. Department of Crystal Chemistry and Crystal Physics, Faculty of Chemistry, Jagiellonian University, Ingardena 3, 30-060, Krakow, Poland
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Abstract: | Lactoferrin is considered as a part of the innate immune system that plays a crucial role in preventing bacterial growth, mostly via an iron sequestration mechanism. Recent data show that bovine lactoferrin prevents late-onset sepsis in preterm very low birth weight neonates by serving as an iron chelator for some bacterial strains; thus, it is very important to control the iron saturation level during diet supplementation. An accurate estimation of lactoferrin iron saturation is essential not only because of its clinical applications but also for a wide range of biochemical experiments. A comprehensive method for the quantification of iron saturation in lactoferrin preparations was developed to obtain a calibration curve enabling the determination of iron saturation levels relying exclusively on the defined ratio of absorbances at 280 and 466 nm (A 280/466). To achieve this goal, selected techniques such as spectrophotometry, ELISA, and ICP-MS were combined. The ability to obtain samples of lactoferrin with determination of its iron content in a simple and fast way has been proven to be very useful. Furthermore, a similar approach could easily be implemented to facilitate the determination of iron saturation level for other metalloproteins in which metal binding results in the appearance of a distinct band in the visible part of the spectrum. |
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