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The Use of Two Radioactive Isotopes of Iron in Tracer Studies of Erythrocytes

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The Use of Two Radioactive Isotopes of Iron in Tracer Studies of Erythrocytes THE USE OF TWO RADIOACTIVE ISOTOPES OF IRON IN TRACER STUDIES OF ERYTHROCYTES Wendell C. Peacock, … , Soma Weiss, John G. Gibson 2nd J Clin Invest. 1946;25(4):605-615. https://doi.org/10.1172/JCI101743. Research Article Find the latest version: https://jci.me/101743/pdf THE USE OF TWO RADIOACTIVE ISOTOPES OF IRON IN TRACER STUDIES OF ERYTHROCYTES 1 By WENDELL C. PEACOCK, ROBLEY D. EVANS, JOHN W. IRVINE, JR., WILFRED M. GOOD, ARTHUR F. KIP, SOMA WEISS,2 AND JOHN G. GIBSON, 2ND (From the Radioactivity Center, Massachusetts Institute of Technology, Cambridge; the Medical Clinic of the Peter Bent Brigham Hospital, and Department of Medicine, Harvard Medical School, Boston) (Received for publication January 18, 1946) INTRODUCTION Ross (6) used these mixed isotopes, but their There are 2 radioactive isotopes of iron suit- radioactivity detection apparatus was sensitive able for use as biological tracers. One of these, only to the radiations from Fe59. The nuclear Fe59, has a half life of 47 days and emits gamma reactions are Fe58 (d, p), Fe59 (Fe58, a stable rays and low energy beta rays. The other, Fe55, isotope, bombarded with deuterons, d, forms, has a half life of about 5 years and emits low through the liberation of a proton, p, Fe59) and energy x-rays only. Fe54 (d, p) Fe55. When iron is bombarded, This paper describes how these 2 isotopes are these 2 isotopes are produced simultaneously, and prepared for red blood cell tracer studies, how cannot be isolated thereafter in a state of radio- biological samples containing the radioactive iron active purity. are prepared for measurements, how the measure- By deuteron bombardment of manganese, the ments are performed, and how the results are reaction Mn55 (d, 2n) Fe55 occurs and only the reported to collaborating medical research teams long-lived radioactive iron isotope is obtained. engaged in various physiological and clinical The manganese targets are prepared from an al- studies. loy of manganese (90 per cent Mn, 8 per cent Cu, Fe59 has been in use for a number of years for 2 per cent Si) which has suitable mechanical and metabolism and red cell studies (1 to 5) and a thermal properties. This alloy is cast into %-inch method of preparing samples for measurement slabs and machined down to %2-inch thickness. with a thin window beta ray counter has been Targets 'A x 1 inch are cut from this material described by Ross (6). The longer lived iso- and silver soldered to the oscillating probe target tope, Fe55, on the other hand, appears not to described by Livingston (7). Evaporation of the have been used previously. target face limits the deuteron beam current to Each isotope has characteristics that make it 100 /La at 14 MEV. Under these conditions our particularly useful in certain problems; the com- cyclotron yields of Fe55 have been approximately bined use of both tracers offers solutions of % of a microcurie per microampere hour. problems which cannot be obtained by the employ- The shorter-lived isotope, Fe59, can be produced ment of either isotope alone. in pure form by the bombardment of cobalt with neutrons, through the nuclear reaction Co59 (n, p) Fe59. PREPARATION OF THE ISOTOPES Deuteron bombardment of cobalt also yields Fe59 as a by-product of the fast neutrons always a. Nuclear bombardment. Bombardment of present with deuteron reactions. Because thin iron with deuterons yields both isotopes mixed. cobalt targets are necessary for thermal conduc- The earlier work of Hahn et al. (1 to 5) and tion, and the total amount of cobalt is small (1 to 2 grams), this is not an efficient method of pro- 1 The work described in this paper was done under a ducing high Fe59 activities. If a comparatively contract, recommended by the Committee on Medical Re- large piece of cobalt (about 40 grams) is soft search, between the Office of Scientific Research and De- the of a velopment and the Massachusetts Institute of Technology, soldered to back beryllium target the in collaboration with the Peter Bent Brigham Hospital. larger amount of cobalt, higher permissible beam 2Deceased, January 31, 1942. currents on Be, and much higher neutron yield 605 606 PEACOCK, EVANS, IRVINE, GOOD, KIP, WEISS, AND GIBSON from the Be9 (d, n) BI0 reaction, give much better The final step in the preparation of the iron is yields of Fe59. Using beryllium, our yields have to synthesize it into a compound suitable for in- been approximately 0.023 microcuries per micro- travenous use in animals and humans. Ferric ampere hour. ammonium citrate, in doses described below, has -b. Radiochemical preparation. Regardless of been found safe for intravenous injection. After the mode of formation of the radioactive iron, the dissolving the Fe2O3 in concentrated HCI and purification and preparation of material for use is diluting, Fe(OH)3 is again precipitated with essentially the same in all cases. The target mate- NH4OH and is well washed with distilled water. rial Fe, Mn, or Co is dissolved in concentrated The precipitate is dissolved in a solution contain- hydrochloric acid, diluted and filtered. If man- ing 3 moles of citric acid for each mole of iron ganese and cobalt of high purity are used it is present. Warming the mixture hastens the solu- necessary to add 1 to 10 mgm. of inactive Fe+++ tion of the precipitate. When the precipitate is as a carrier. Usually there is enough iron in the completely dissolved the solution is brought to a target so that such additions are unnecessary. pH of 7.0 with NH4OH and filtered into a clean After oxidation of the iron from Fe++ to Fe+++ bottle. with H202 the iron is precipitated as Fe (OH)3 This solution can be diluted with neutral dis- with pyridine (8). tilled water to obtain any desired concentration The precipitate of Fe (OH)3 is dissolved in 6 M of iron. It withstands sterilization by autoclaving HC1, and 10 mgm. each of Mn, Co and Zn added if sealed in glass ampouls, but heating in open con- as carriers to aid in eliminating the radioactive iso- tainers may result in the formation of a precipi- topes of these elements from the iron. Ferric tate. Ferric ammonium citrate is photosensitive hydroxide is again precipitated with pyridine, and should be kept in the dark to avoid precipita- leaving the Mn, Co and Zn in solution. This tion. The compound does not inhibit the growth precipitation procedure is repeated 2 to 4 times of common molds. until the activity in the filtrate is low. After elimination of these 3 elements, the hy- PREPARATION OF RADIOACTIVE RED CELLS drogen sulphide group is eliminated by adding 50 When radioactive ferric ammonium citrate is' mgm. of Cu++ as carrier for this group, and pre- injected intravenously into animals or humans it cipitating the sulphide with H2S from a 0.1 M is rapidly removed from the blood stream. A HCl solution. The preciptate is filtered off and large portion of the iron is retained in the body discarded. The filtrate is boiled to eliminate HKS, tissues concerned with iron storage. Hahn et al. Fe++ is oxidized to Fe... with H2O,, and Fe (OH)3 (10) gave ferric ammonium citrate to dogs and is preciptated with NH4OH. This precipitate assayed liver and spleen for both ferritin and is dissolved in 1.5 M H2SO4, and 10 mgm. of in- radioactive iron, and found a relatively high level dium is added (radioactive indium is formed from of radioactivity in the ferritin iron. It therefore the cadmium in the silver solder used). Pre- seems highly probable that the radio-iron becomes cipitation of Fe... with cupferron leaves the in- intimately mixed with all of the body iron stores dium behind in the filtrate .(9). The cupferron present when the isotope was injected, and is precipitate is carefully ignited, the Fe2O3 dis- hence available for hematopoietic needs. solved in concentrated HCl, diluted to 3 M, in- It is generally believed that hemoglobin is dium carrier added and the precipitation with "laid down" inside the developing erythrocyte. cupferron repeated. This precipitate is ignited, If some of the iron atoms involved in this syn- dissolved in concentrated HCl, diluted, filtered thesis are radioactive, a proportionate number of (to remove unburned carbon which usually is them will become an integral part of the new present after igniting the cupferron precipitate) hemoglobin molecule within the newly developed and the iron reprecipitated as Fe(OH)3 with cell. When the cell is released into the circula- NH4OH. This preciptate is carefully ignited in tion, its presence in the blood stream can be de- a weighed crucible and the Fe2O3 weighed to de- tected as long as it remains morphologically in- termine the weight of iron present, tact. When the tagged red cell is destroyed, its RADIOACTIVE ISOTOPES OF IRON IN STUDIES OF RED CELLS 607 hemoglobin derived iron is very rapidly removed PREPARATION OF BLOOD SAMPLES FOR from the plasma, to be reused, to some extent at RADIOACTIVE MEASUREMENT least, in the synthesis of new hemoglobin. Both of the iron isotopes have radiations that In general, each red cell contains approximately are readily absorbed in a thick sample of ma- 0.1 per cent iron, or about a thousand million terial. Thus, for maximum detection efficiency atoms of iron. In the radioactive donors, between all of the iron must be separated from extraneous, 1 and 10 of these iron atoms contains a radioac- material, and the metal plated as a thin uniform of the iron atoms in tive nucleus, the vast majority film on a metal planchet.
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