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Abstract

Background

Iron is ubiquitously distributed in biology, only a miniscule amount exists as free is capable of catalysing production of highly toxic reactive hydroxyl radicle. This free iron also called; labile iron, non-transferrin bound iron or catalytic iron (CI). CI is measured by bleomycin detectable iron assay. The assay as described originally was difficult to perform accurately and reproducibly due to variations of pH in the assay mixture and due to the lack of properly diluted iron standards.

Methods

In our laboratory we modified the assay for serum/plasma so that the variations of pH in assay medium were constantly between 7.4 and 7.6 using acid diluted iron standards by multiple treatments of Chelex resin which is alkaline.

Results

Intra assay CV for low, medium, and high levels of catalytic iron was 0.05%, 0.61% and 0.68% whereas the interassay CV was 0.06%, 0.96% and 0.28% respectively. The modified assay is highly sensitive being able to detect levels as low as 0.1 μmoles/l. In patients on maintenance haemodialysis CI measured by the original assay failed to detect any catalytic iron in almost all of these samples whereas by modified method it was measurable in all patients with a mean of 0.66 ± 0.10 μmoles/l. Normal values for catalytic iron in subjects having no comorbidities measured by modified method is 0.11 ± 0.06 μmoles/l.

Conclusions

The modified assay is reproducible and more sensitive than original assay and has been validated in several clinical studies.

Keywords

Catalytic iron non-transferrin bound iron labile iron reactive oxygen species acute coronary syndrome acute kidney injury major adverse cardiac events major adverse kidney events mortality

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References

Gutteridge

Rowley

Halliwell

. Superoxide-dependent formation of hydroxyl radicals in the presence of iron salts. Detection of “free” iron in biological systems by using bleomycin-dependent degradation of DNA. Biochem J 1981; 199(1): 263–265.

Gutteridge

Hou

. Iron complexes and their reactivity in the bleomycin assay for radical-promoting loosely-bound iron. Free Radic Res Commun 1986; 2(3): 143–151.

Evans

Halliwell

. Measurement of iron and copper in biological systems: bleomycin and copper-phenanthroline assays. Methods Enzymol 1994; 233: 82–92.

Lele

Shah

McCullough

, et al. Serum catalytic iron as a novel biomarker of vascular injury in acute coronary syndromes. EuroIntervention 2009; 5(3): 336–342.

Steen

Cannon

Lele

, et al. Prognostic evaluation of catalytic iron in patients with acute coronary syndromes. Clin Cardiol 2013; 36(3): 139–145.

Lele

Mukhopadhyay

Mardikar

, et al. Impact of catalytic iron on mortality in patients with acute coronary syndrome exposed to iodinated radiocontrast-The Iscom Study. Am Heart J 2013; 165(5): 744–751.

Leaf

Rajapurkar

Lele

, et al. Plasma catalytic iron, AKI, and death among critically ill patients. Clin J Am Soc Nephrol 2014; 9(11): 1849–1856.

Leaf

Rajapurkar

Lele

, et al. Increased plasma catalytic iron in patients may mediate acute kidney injury and death following cardiac surgery. Kidney Int 2015; 87(5): 1046–1054.

Leaf

Rajapurkar

Lele

, et al. Iron, hepcidin, and death in human AKI. J Am Soc Nephrol 2019; 30(3): 493–504.

10.

Fuernau

Traeder

Lele

, et al. Catalytic iron in acute myocardial infarction complicated by cardiogenic shock - a biomarker substudy of the IABP-SHOCK II-trial. Int J Cardiol 2017; 227: 83–88.

11.

Chakurkar

Rajapurkar

Lele

, et al. Increased serum catalytic iron may mediate tissue injury and death in patients with COVID-19. Sci Rep 2021; 11: 19618.

12.

Van Coillie

Van San

Goetschalckx

, et al. Targeting ferroptosis protects against experimental (multi) organ dysfunction and death. Nat Commun 2022; 13: 1046.

13.

Yadav

Ghosh

Divyaveer

, et al. Serum catalytic iron and progression of chronic kidney disease: findings from the ICKD study. Nephrol Dial Transplant 2022; 37: 1879–1887.

14.

Peleman

Van Coillie

Ligthart

, et al. Ferroptosis and pyroptosis signatures in critical COVID-19 patients. Cell Death Differ 2023; 30(9): 2066–2077.

15.

Linnet

Boyd

. Selection and analytical evaluation of methods-with statistical techniques. In: Burtis

Ashwood

Burns

(eds). Teitz textbook of clinical chemistry and molecular diagnostics. 4th ed. St. Louis, MO: Saunders, 2006, pp. 353–407.

16.

National Committee for Clinical Laboratory Standards (NCCLS) . User evaluation of precision performance of clinical chemistry devices. NCCLS Tentative Guideline EP5-T. Wayne, PA: NCCLS, 1984.

17.

Ehrenfeld

Rodriguez

Hecht

, et al. Copper(I)-bleomycin: structurally unique complex that mediates oxidative DNA strand scission. Biochemistry 1985; 24(1): 81–92. DOI: 10.1021/bi00322a013.

18.

Suzuki

Kuwahara

Sugiura

. Copper-bleomycin has no significant DNA cleavage activity. Biochemistry 1985; 24(18): 4719–4721. DOI: 10.1021/bi00339a001.

19.

Ehrenfeld

Shipley

Heimbrook

, et al. Copper-dependent cleavage of DNA by bleomycin. Biochemistry 1987; 26(3): 931–942. DOI: 10.1021/bi00377a038.

20.

Takahashi

Takita

Umezawa

. The nature of thiol-compounds which trap cuprous ion reductively liberated from bleomycin-Cu(II) in cells. J Antibiot (Tokyo) 1987; 40(3): 348–353. DOI: 10.7164/antibiotics.40.348.

21.

Surgenor

Koechlin

Strong

. Chemical, clinical, and immunological studies on the products of human plasma fractionation. XXXVII. The metal-combining globulin of human plasma. J Clin Investig 1949; 28(1): 73–78.

A simpler and more sensitive modified catalytic (bleomycin detectable) iron assay

Abstract

Background

Methods

Results

Conclusions

Keywords

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References