A Review of Fluorescence Methods for Assessing Labile Iron in Cells and Biological Fluids

Espósito, Breno Pannia; Epsztejn, Silvina; Breuer, William; Cabantchik, Z. Ioav

doi:10.1006/abio.2002.5611

Cited by 218 publications

(184 citation statements)

References 101 publications

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“…That the processes monitored by endosomal Oxyburst-green or cytosolic CALG were associated with NTBI and not with TBI was deduced from the fact that addition of apotransferrin to media containing native or artificial NTBI abrogated the processes in a similar way as hydroxyethylstarch-DFO. We also found that the proposed steps of NTBI uptake into RAW cells were largely recapitulated with thalassemia major sera probed with CALG, that reversibly binds iron, 20,21 including components of plasma NTBI, and is endocytosed commensurately with NTBI levels ( Figure 3). However, a key question is to what extent CALG added to sera reports NTBI ingress into cells rather than promotes NTBI ingress by binding to plasma NTBI, whether present as low-molecular weight complexes or bound to plasma proteins.…”

mentioning

confidence: 59%

“…Cells were subsequently washed with HEPES-buffered saline (HBS, 130 mM NaCl, 20 mM Hepes, pH 7.4) and bathed at 37°C in DMEM-HEPES containing 0.5 mM probenecid (to minimize probe leakage). 13,19,20 To assess fluorescence properties, the cells were analyzed either microscopically or by flow cytometry (following release by trypsinization). Epi-fluorescence microscopy analysis of CALG was carried out using EXC: 488nm and Em: 520 nm and for CALB EXC: 390 nm and Em: 430 nm).…”

Section: Measurement Of Cytosolic Calg or Calb Fluorescence In Cellsmentioning

confidence: 99%

“…For tracing ingress of labile iron into the cytosol we used CALG, a green fluorescent metal sensor that is loaded into cells via its CALG-AM precursor. 13,20 The probe undergoes swift and stoichiometric quenching by interacting with labile iron and recovers its fluorescence when challenged with iron chelators. Using CALG-laden RAW macrophages and fluorescence microscopy live imaging, we found that ferric compounds such as ferric citrate, considered a major component of NTBI in iron overloaded sera, 11,12 failed to evoke changes in cytosol fluorescence over a period of 90 min, unless the medium was supplemented with human serum albumin (HSA) ( Figure 1 shows data for only two time points: 0 and 40 min).…”

Section: Ntbi Uptake Into Cells As Revealed With Cytosolic Iron Markersmentioning

confidence: 99%

“…After washing of cells, the fluorescence of Oxyburst-green was monitored before and 10 min after addition of H2O2 (50 μM). Finally, as CALG-Fe complexes are essentially dissociable, 20,21 we considered the possibility that during the incubation of cells with CALG-supplemented thalassemia major sera, a fraction of the endosomal CALG-Fe undergoes dissociation followed by Fe translocation into the cytosol via NRAMP1 and/or 2 (DMT1), the two principal iron transporters in endocytic vesicles. That possibility was examined in two sublines of RAW cells over-expressing functional NRAMP1 (RAW 37) or non-functional antisense NRAMP1 (RAW 21).…”

Section: Ntbi Uptake and The Status Of Iron In Thalassemia Major Seramentioning

confidence: 99%

See 3 more Smart Citations

The role of endocytic pathways in cellular uptake of plasma non-transferrin iron

Sohn¹,

Ghoti²,

Breuer³

et al. 2011

Haematologica

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mentioning

confidence: 59%

Section: Measurement Of Cytosolic Calg or Calb Fluorescence In Cellsmentioning

confidence: 99%

Section: Ntbi Uptake Into Cells As Revealed With Cytosolic Iron Markersmentioning

confidence: 99%

Section: Ntbi Uptake and The Status Of Iron In Thalassemia Major Seramentioning

confidence: 99%

See 2 more Smart Citations

The role of endocytic pathways in cellular uptake of plasma non-transferrin iron

Sohn¹,

Ghoti²,

Breuer³

et al. 2011

Haematologica

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show abstract

“…There are two possibilities to measure NTBI: Indirect measurement involves the chemical reactivity of NTBI, for instance, the bleomycin method (39 -41), whereas direct measurement typically involves the use of scavenging molecules to mobilize loose, nonspecifically bound iron (42,43). The results of such tests largely depend on the design of the assay.…”

mentioning

confidence: 99%

Non–Transferrin-Bound Iron in the Serum of Hemodialysis Patients Who Receive Ferric Saccharate

Scheiber-Mojdehkar

Lutzky²,

Schaufler³

et al. 2004

Journal of the American Society of Nephrology

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Abstract. Intravenous iron (iv.Fe) is used to optimize response to recombinant human erythropoietin (r-HuEPO) in ESRD, but no consensus exists with respect to the best regimen to avoid transferrin "oversaturation," oxidative stress, and the occurrence of non-transferrin-bound iron (NTBI). Iv.Fe was stopped for 1 wk in 35 hemodialysis (HD) patients who were routinely receiving iv.Fe and r-HuEPO. The iv.Fe group received 100 mg of ferric saccharate (Venofer) at the end of the first HD session, whereas the time-control group was treated under the same conditions but received no iv.Fe. Serum samples were taken before the first HD session, immediately and 60 min after iv.Fe administration, and before the next HD session. Sera were analyzed for NTBI and peroxides; transferrin saturation was analyzed by urea-PAGE and Western blot. In an in vitro model system with HepG2 cells, the effects of ESRD serum on the labile iron pool (LIP) were assayed using the fluorescence calcein assay. NTBI significantly increased after iv.Fe-administration and returned to baseline values before the next HDsession. There was a shift from apo-to monoferric transferrin, but no "oversaturation" of transferrin after iv.Fe-treatment. Peroxides increased in both groups after HD. Hemodialysis decreased bioavailable iron for the LIP in HepG2-cells, whereas serum of iv.Fe-treated HD patients highly increased the LIP in these cells. A total of 100 mg of iv.Fe led to NTBI generation but not to an oversaturation of transferrin. Peroxide concentrations significantly increased during HD but were not correlated to iv.Fe administration and seemed to result from other sources of oxidative stress related to HD. NTBI can enter liver cells and increase the potentially harmful LIP.

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Biological and Biomedical Aspects of Metal Phenolates

Ming

2014

Patai's Chemistry of Functional Groups

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The phenol functionality is involved in diverse biological processes and functions, serving as a proton donor and as a metal‐binding ligand. It is commonly found in many biochemicals (such as siderophores and the amino acids tyrosine and tryptophan), pharmaceuticals (including antibiotics, metal‐chelating agents, and diagnostic agents), and natural products (such as polyphenols, flavonoids, and salicylic acid). In association with other functional groups, the phenol moiety forms various types of metal‐binding sites of diverse structures and functions, for example, for iron binding and transport, in enzymes for oxygenation of substrates, and for biotransformation of aromatic compounds. Many environmentally hazardous aromatic compounds such as bisphenol A, PCB, and DDT can be degraded by tyrosinate(phenolate)‐containing metalloenzymes from microorganisms, which provides a clue for bioremediation of these toxic substances. Moreover, the phenol‐containing wet‐adhesive byssal proteins from some clams and mussels has stimulated further development of adhesive materials for medical and other applications; and the study of binding of transferrin with nanoparticles affords better understanding of protein–mineral interfacial interactions. In addition to these naturally occurring metal‐phenolate centers, there are many metal complexes of phenol and derivatives that exhibit various catalytic activities and serve as structural and/or functional model systems for tyrosinate‐containing metalloproteins.

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A Review of Fluorescence Methods for Assessing Labile Iron in Cells and Biological Fluids

Cited by 218 publications

References 101 publications

The role of endocytic pathways in cellular uptake of plasma non-transferrin iron

The role of endocytic pathways in cellular uptake of plasma non-transferrin iron

Non–Transferrin-Bound Iron in the Serum of Hemodialysis Patients Who Receive Ferric Saccharate

Biological and Biomedical Aspects of Metal Phenolates

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