1238
Anal. Chem. 1986, 58, 1238-1241
Actinide Separations for a Spectrometry Using Neodymium Fluoride Coprecipitation Forest D. H i n d m a n
U S . Department of Energy, Radiological and Environmental Sciences Laboratory, 785 DOE Place, Idaho Falls, Idaho 83402
A procedure was developed In which a 1O-g soli sample was fused in potassium fluoride followed by a pyrosulfate fusion. The flux was dissolved in dilute hydrochloric acld and thorium, uranium, plutonium, and americium were copreclpltaled on barium sulfate. The barium sulfate was dissolved and precipitated in the presence of dlethyienetriaminepentaacetic acid (DTPA) to separate the actjnides from barium sulfate. The filtrate was decomposed, fused in a pyrosulfate flux, and dissolved in dilute nitrlc acid. Thorium was separated from the other actinides by coprecipitation on ceric Iodate. Uranium, plutonium, and americium were separated sequentlaiiy, one from the other, by oxidizing them to their highest oxidation states followed by a series of coprecipltations of the nucildes. The isolated nuclides were coprecipltated on neodymium fluoride, filtered, and counted by a! spectrometry. The recoveries ranged from 85 % to 95 %. The resolution for full width at half maximum (fwhm) was between 60 and 70 keV. Decontamination factors ranged between lo3 and lo4.
A developing trend in this laboratory, and in others as well, is toward doing more analyses with fewer people. T o meet this challenge the radiochemist needs procedures that are more rapid, more economical, and more efficient. The procedure described herein fulfills the above needs and yields good decontamination factors, high recoveries, and excellent resolution of the a spectra for uranium, plutonium, americium, and thorium. All four elements above are separated from one another by coprecipitation techniques that have eliminated the solvent extraction procedures used in the past ( 1 , 2 ) . The extraction procedures in general for americium ( I , 2 ) do not separate it from the rare earths. This separation is normally made with a column procedure ( 3 ) after the extractions are completed. In this procedure, however, the americium is oxidized to the highest valence state where the rare earths can be precipitated as fluorides and separated from it by centrifugation ( 4 ) . A considerable savings in time is achieved by eliminating both the solvent extractions for all four elements and the column procedure for americium. Thorium is separated from the other three elements with an iodate coprecipitation technique. If uranium is to be determined from the same sample, polonium, protactinium, and platinum must be separated from it. The iodate coprecipitation mentioned above removes all three elements along with the thorium. If americium is also being determined from this same sample, cerium must be separated from it. Like polonium, protactinium, and platinum, the cerium is precipitated along with the thorium. The four potential interferences are eliminated a t no extra cost in time or effort while the thorium determination is carried out. Time has also been saved in reagent preparation because most of the reagents used in this procedure are simple aqueous solutions of very soluble inorganic compounds.
EXPERIMENTAL SECTION Instrumentation. The a spectrometry system has been described previously (2). Reagents. All solutions were stored in polypropylene bottles. Neodymium Chloride (10 mg of NdlrnL). Twenty-five milliliters of 12 M hydrochloricacid and 1.17 g of neodymium oxide were heated on a hotplate until the neodymium oxide was in solution. The solution was cooled and diluted to 100 mL. Neodymium Chloride (0.5 mg of NdlmL). Five milliliters of neodymium chloride (10 mg of Nd/mL) was diluted to 100 mL with water. Neodymium Perchlorate (0.5 mg of NdlmL). Five milliliters of neodymium chloride (10 mg of Nd/mL) was heated to perchloric acid fumes in 5 mL of 12 M perchloric acid. The solution was cooled, diluted to 100 mL with water, and treated with 10-20 mg of solid potassium dichromate. Tracer Solutions. The uranium, plutonium, americium, and throium tracer solutions were purified (5) and standardized (2) as described previously. However, thorium-229 rather than thorium-234 was used in this work. Carbon Suspension (6). A 47-mm GA-6 Metricel filter (Gelman Sciences Co., Ann Arbor, MI) was fumed for about 5 min in 5 mL of 18 M sulfuric acid. The suspension was cooled and diluted to 50 mL with water. Substrate Suspension (6). One milliliter of neodymium chloride (10mg of Nd/mL) and 20 mL of 12 M hydrochloric acid were diluted to 400 mL with water. Ten milliliters of 29 M hydrofluoric acid and 1to 2 mL of carbon suspension were added with swirling after each addition and the suspension was diluted to 500 mL with water. Each day before use, the substrate suspension was placed in a sonic bath for 15 min. Ammonium Hydroxide-DTPA. Thirty grams of diethylenetriaminepentaacetic acid (DTPA) was added to 800 mL of water, dissolved with 120 mL of 15 M ammonium hydroxide, and the mixture was diluted to 1000 mL. Reprecipitating Solution. One-hundred-thirty-fivegrams of anhydrous potassium sulfate was dissolved with heat in 800 mL of water and 50 mL of 12 M hydrochloricacid. The solution was cooled and diluted to 1000 mL. Potassium and Sodium Sulfate. Seventy-five grams of potassium sulfate and 75 g of sodium sulfate were dissolved in 800 mL of water with heat, if needed, and the resulting solution was diluted to 1000 mL with water. Ammonium Iodate. Twenty grams of ammonium iodate and 20 mL of 16 M nitric acid were added to 800 mL of water and the mixture was heated to boiling, cooled, and filtered through a 47-mm Metricel GA-6 filter. The filtrate was diluted to 1000 mL with water. Ammonium Fluoride. Eighty grams of ammonium fluoride was dissolved and diluted to 1000 mL. The portion needed each day for use in the precipitation of the rare earths from americium was treated with enough solid ammonium persulfate t o give a solution approximately 5% in ammonium persulfate. Cerium Nitrate (10 mg of Ce/mL). Three and one-tenth grams of cerous nitrate hexahydrate (Allied Chemical Co., thorium free) was dissolved in 100 mL of 1%nitric acid. Zirconyl Perchlorate (10 mg of ZrlmL). Two and nine-tenths grams of zirconyl nitrate dihydrate (Fisher Scientific Co., thorium free) was fumed in 10 mL of 12 M perchloric acid to 5 mL and the solution was then cooled and diluted to 100 mL with water.
This article not subject to U S . Copyright. Published 1986 by the American Chemical Society
ANALYTICAL CHEMISTRY, VOL. 58, NO. 6, MAY 1986
If the solution was turbid, it was filtered through a 47-mm GA-6 filter. The turbidity was detected by the Tyndall beam generated using a flashlight in a dark room. Procedure. Sample Decomposition (2). Ten grams of soil was placed in a 250-mL platinum dish, treated with 10 mL of 16 M nitric acid, the appropriate tracers (5), and 10 mL of 29 M hydrofluoric acid. After the mixture was evaporated to a thin paste on an asbestos-covered hot plate, 30 g of anhydrous potassium fluoride was mixed into the sample with a stirring rod made of Teflon. The sample was then heated to a fusion over a Fisher blast burner. The dish was placed on a hot plate and allowed to cool to the temperature of the hot plate. Thirty-five milliliters of 18 M sulfuric acid was added in successive small portions as quickly as the frothing in the dish would allow. When the reaction stopped, 20 g of anhydrous sodium sulfate was added and the sample was heated to a pyrosulfate fusion over the blast burner. Coprecipitation on Barium Sulfate (7).Five-hundred milliliters of water and 150 mL of 12 M hydrochloric acid were heated to boiling in a 1000-mL beaker containing about five silicon carbide boiling chips. The fusion cake was dissolved in the boiling solution. When uranium, plutonium, americium, and thorium were to be determined, the boiling solution was treated with just enough solid stannous chloride to reduce the yellow ferric iron in the sample to give a colorless solution. Four drops of 1%safranine-0 were added followed by ten drops of 20% titanium trichloride. When americium, plutonium, and thorium were to be determined, the fusion cake was dissolved as described above and treated with 10 mL of 2.5% potassium metabisulfite. The boiling was continued for 15 min. Either treatment of the sample above was followed by the addition of 100 g of anhydrous potassium sulfate to the boiling solution. As soon as the potassium sulfate was dissolved and the sample was boiling again, four 10-mL portions of 0.5% barium chloride were added with stirring, and the sample was boiled for 5 min after each addition. The sample was filtered through a 47-mm Metricel GA-6 filter. The transfer was completed and the filter was rinsed with 0.5% sulfuric acid wash solution. The precipitate and filter were placed in a 100-mL, thick-walled, round-bottomed, glass centrifuge tube. Reprecipitation of the Nuclides (2). The filter and precipitate were dissolved in 4 mL of sulfuric acid and about 1mL of 16 M nitric acid by heating the centrifuge tube over the blast burner. When the nitric acid was boiled away and the sulfuric acid just began to fume, 2 drops of 1:l 16 M nitric acid-12 M perchloric acid were added and the sample was again heated just to sulfuric acid fumes. When uranium was to be reprecipitated, 100 mg of solid hydrazine sulfate was added and the solution was heated to sulfuric acid fumes yielding a colorless solution. Repeated treatments may be necessary with the hydrazine sulfate to achieve a colorless solution. The sample was cooled and the barium sulfate reprecipitated with 60 mL of reprecipitating solution containing 5 drops of 20% titanium trichloride. If uranium was not to be reprecipitated, an adddition of 5 drops of 25% potassium metabisulfite replaced the titanium trichloride with no hydrazine sulfate added. The sample was heated in a hot water bath for 10 min with occasional swirling. The sample was cooled in a cold water bath and centrifuged at 2000 rpm for 5 min. The supernate was decanted and discarded. The precipitate was completely broken up and suspended in 25 mL of 0.5% sulfuric acid solution with the aid of a vortex mixer. The sample was centrifuged again and the supernate discarded. Elimination of Barium Sulfate. The sample was suspended in 40 mL of ammonium hydroxide-DTPA solution using a vortex mixer. The sample was then heated in a water bath until the barium sulfate was in solution. The sample was heated for 10 rnin more, treated with 5 mL of 17 M acetic acid and 2 mL of potassium and sodium sulfate. The solution was then heated for 5 min, cooled, and filtered through a 47-mm Metricel GA-6 filter. The filtrate was caught in a 250-mL Erlenmeyer flask and treated with 10 mL of 18 M sulfuric acid. Three glass boiling beads were added and the sample was evaporated to sulfuric acid fumes on the bare hot plate. At this point black drops of solution condensed on the walls of the flask. Two milliliters of 12 M perchloric acid was added and. the sample evaporated once more to sulfuric acid fumes. When a colorless solution was obtained, the sample was
1239
heated over the blast burner until no more white fumes could be blown from the flask. A heavy wire gauze was used to protect the flask from thermal shock while heating. The sample was cooled first on the hot plate and then on an asbestos board, again to prevent breakage from thermal shock. The sample was treated with 1mL of 18 M sulfuric acid, fused once more, and cooled as described above. Coprecipitation of Plus Four Elements on Ceric Iodate. Twenty milliliters of 2% nitric acid was added to the fused sample in the 250-mL Erlenmeyer flask. The sample was heated gently on an asbestos-covered hot plate until it was dissolved. The sample was then transferred to a 50-mL round-bottomed glass centrifuge tube. A minimum of water was used to complete the transfer. One-half milliliter of cerium nitrate (10 mg of Ce/mL), 2 drops of 1%manganous sulfate hexahydrate, and approximately 0.25 g of solid ammonium persulfate were added. The sample was heated in a hot water bath until the yellow ceric ion color and the purple permanganate color were fully developed. The heating was continued for 10 min and the sample was treated once more with 0.25 g of ammonium persulfate. The heating was continued for 5 min and the sample was cooled in a cold water bath. Ceric iodate was precipitated with the addition of 10 mL of ammonium iodate solution, the sample was allowed to stand for 5 min, and the mixture was centrifuged for 5 min at 2000 rpm. The supernate was transferred, without rinsing the centrifuge tube, back to the original 250-mL Erlenmeyer flask and saved for uranium, plutonium, and americium determinations. The ceric iodate precipitate in the centrifuge tube was saved for thorium determination. Uranium Determination. Five milliliters of 12 M hydrochloric acid, 1 mL of 18 M sulfuric acid, and three glass beads were added to the supernate from the ceric iodate step. The sample was evaporated to dryness on an asbestos-covered hot plate, treated with 5 drops of 18 M sulfuric acid, and heated to a clear melt over the blast burner until bubbles atopped forming. The sample was cooled very slowly on the hot plate and placed on an asbestos board to minimize thermal shock to the flask. Twenty milliliters of 2% hydrochloric acid was added to the sample, which was heated slowly to boiling on the asbestos-covered hot plate. The sample was boiled slowly for 5 min and transferred to a 50-mL round-bottomed glass centrifuge tube using a minimum of water to complete the transfer. After 0.5 mL of neodymium chloride (10 mg of Nd/mL) was added, the sample was cooled in a cold water bath. The solution was then treated with 3 mL of ammonium fluoride solution, allowed to stand for 10 min, and centrifuged for 5 min at 2000 rpm. One milliliter of neodymium chloride (0.5 mg of Nd/mL) was added to the solution and swirled gently to avoid disturbing the precipitate in the bottom of the centrifuge tube. The sample was allowed to stand for 10 min, before centrifuging. The supernate was decanted gently, without rinsing the glass tube, into a 50-mLround-bottomed polycarbonate centrifuge tube and saved for uranium determination. Two milliliters of 1 2 M perchloric acid and 2 drops of 1% potassium dichromate were added to the precipitate, and the sample was heated to perchloric acid fumes on the asbestoscovered hot plate. This was done without delay to minimize the contact time of fluoride with glass. This solution was saved for plutonium determination. Two milliliters of 12 M hydrochloric acid, 1 drop of 0.1% safranine-0, and 2 drops of 20% titanium trichloride were added to the uranium sample in the polycarbonate centrifuge tube. One-tenth milliliter of neodymium chloride (0.5 mg of Nd/mL) and 5 mL of 29 M hydrofluoric acid were added in that order, with swirling after each addition. The uranium sample was then heated in a hot water bath for 10 min and cooled. A 25-mm Tuffryn HT-200 membrane filter (Gelman Sciences, Ann Arbor, MI) or equivalent was mounted on a stainless steel support in a polysulfone twist-lock funnel (Gelman Sciences, Ann Arbor, MI). With vacuum applied, about 2 mL of 80% ethanol was drawn through the filter. As the filter went dry, the following solutions were added, in order, to the center of the filter: 5 mL of substrate suspension (freshly treated for 15 min in the sonic bath), the sample (after vigorous swirling), a hot 5 mL of 20% hydrofluoric acid-10% perchloric acid rinse of the centrifuge tube, a 5 mL water rinse of the filter, and a 2 mL 80% ethanol rinse of the filter. The filter was dried for 5 min under a heat lamp
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ANALYTICAL CHEMISTRY, VOL. 58, NO. 6, MAY 1986
at a distance of 12-16 in. and submitted to a spectrometry for the determination of uranium. This filtration procedure is used for each nuclide fraction being determined. Plutonium Determination. The perchloric acid solution in the glass centrifuge tube was heated to boiling on the asbestos-covered hot plate for 15 min. The solution was cooled, treated with 20 mL of 1% perchloric acid-O.Ol% potassium dichromate solution, and cooled in a cold water bath. Five milliliters of 29 M hydrofluoric acid containing 2 drops of 1%potasium dichromate was added with swirling, the sample was allowed to stand for 10 min and centrifuged at 2000 rpm for 5 min. One milliliter of neodymium perchlorate (0.5 mg of Nd/mL) was added and the sample was swirled gently to avoid disturbing the precipitate in the bottom of the centrifuge tube. The sample was allowed to stand for another 10 min and centrifuged again. The supernate was gently decanted, without rinsing the glass tube, into a 50-mL round-bottomed polycarbonate centrifuge tube and saved for the plutonium determination. The precipitate was treated with 1mL of 18 M sulfuric acid and 2 mL of potassium and sodium sulfate solution. The sample was heated to a clear melt over the blast burner, cooled, and saved for the americium determination. The fusion was completed without delay to minimize the contact time of fluoride with glass. The plutonium sample in the polycarbonate centrifuge tube was treated with 1 drop of 30% hydrogen peroxide and heated in a hot water bath for 10 min. One-tenth milliliter of neodymium chloride (0.5 mq of Nd/mL) was added with swirling and the solution was heated 10 min and cooled. The sample was filtered onto an HT-200 membrane filter. Americium Determination (4). The melt in the glass centrifuge tube was treated with 20 mL of 2% nitric acid, 1 drop of 0.2% silver nitrate, and 1drop of 1% manganous sulfate hexahydrate and heated in a hot water bath until the melt was in solution. Approximately 0.25 g of solid ammonium persulfate was added with swirling and the sample was heated in the hot water bath to the purple permanganate color. The heating was continued for 10 min more. Another 0.25 g of ammonium persulfate was added with swirlingwandthe sample was heated for 5 min more. The sample was cooled in a cold water bath, treated with 3 mL of ammonium fluoride solution (see reagent preparation), allowed to stand for 10 min, and centrifuged at 2000 rpm for 5 min. The supernate was decanted carefully, without rinsing the centrifuge tube, into a 50-mL round-bottomed polycarbonate centrifuge tube and the precipitate was discarded. One milliliter of 50% manganous sulfate hexahydrate solution was added to the supernate in the polycarbonate centrifuge tube and the sample was allowed to stand for 10 min. One-tenth milliliter of neodymium perchlorate (0.5 mg of Nd/mL) and 5 mL of 29 M hydrofluoric acid (pretreated with 2 drops of 1% potassium dichromate) were added to the sample, in that order, with swirling after each addition. The sample was allowed to stand at room temperature for 10 min and filtered onto an HT-200 membrane filter. Thorium Determination. The ceric iodate precipitate in the 50-mL round-bottomed glass centrifuge tube was suspended in 1mL of 12 M hydrochloric acid and 2 mL of 12 M perchloric acid. The sample was heated to perchloric acid fumes and fumed at this temperature for 5 min. The sample was cooled, diluted with 20 mL of 1%perchloric acid, and treated with 2 drops of 25% sodium nitrite and 0.5 mL of zirconyl perchlorate (10 mg of Zr/mL). Zirconium iodate was precipitated with the addition of 10 mL of ammonium iodate and the mixture was allowed to stand for 5 rnin and centrifuged for 5 min at 2000 rpm. The supernate was discarded and the precipitate was dissolved and reprecipitated starting with the addition of 1mL of 12 M hydrochloricacid and 2 mL of 12 M perchloric acid. The supernate from the second centrifugation was discarded and the precipitate was dissolved in 1 mL of 12 M hydrochloric acid and 2 mL of 1 2 M perchloric acid. One-tenth milliliter of 29 M hydrofluoric acid was added and heated to perchloric acid fumes on an asbestos-covered hot plate. The fuming was continued for 5 rnin and the solution was cooled and transferred to a 50-mL round-bottomed polycarbonate centrifuge tube using 20 mL of 1% perchloric acid to complete the transfer. One-tenth milliliter of neodymium chloride (0.5 mg of Nd/mL) and 5 mL of 29 M hydrofluoric acid were added, in this order, with swirling after each addition. The sample was
heated for 10 rnin in a hot water bath, cooled, and filtered onto an HT-200 membrane filter.
RESULTS AND DISCUSSION When uranium-232 tracer must be used in the determination of uranium, the tracer must be separated from the thorium-228 daughter before being introduced to the sample (5). If this is not done, high thorium blanks will be obtained during the determination of thorium. During the development of the thorium section of this procedure, silver catalyst was used in the persulfate oxidation of uranium, plutonium, and americium to 6+. Polonium, platinum, and cerium were oxidized to 4+. The 4+ elements were then coprecipitated on zirconium iodate rather than on ceric iodate. This approach to the separation worked very well, but the silver ended up in the final uranium fraction and introduced a precipitate during the reduction of uranium with titanium trichloride. When silver was omitted from the persulfate oxidation, the unwanted precipitate was eliminated. However, part of the americium coprecipitated on the zirconium iodate. When zirconium was replaced with cerium, the americium was completely oxidized to 6+ and remained in solution along with uranium (VI) and plutonium(V1). The ceric iodate carried thorium, polonium, platinum, protactinium, and cerium, with the yellow ceric and purple permanganate colors serving as an indicator for the oxidation with persulfate. Even though protactinium coprecipitates on ceric iodate along with thorium, i t does not coprecipitate with thorium on the 50 pg of neodymium precipitated as neodymium fluoride. In this way both uranium and thorium are free of spectral interference from protactinium. Unlike protactinium, however, neptunium is a spectral interference to uranium-234, thorium-230, and plutonium-242 tracer but not to plutonium-236 tracer. The complete elimination of polonium-210 from the final uranium fraction is very important because the polonium-210will precipitate along with the uranium and contribute a counts directly under the uranium-232 tracer peak giving a calculated uranium recovery that is too high. The uranium results will then be too low. If platinum is not separated from uranium, platinum metal will precipitate with uranium and be filtered onto the HT-200 filter with the uranium. This additional mass will serve as an adsorber and cause poor resolution. If cerium is not precipitated completely as ceric iodate, part of the cerium will follow americium into the final americium fraction. When americium is oxidized to americium(V1) by the silver-catalyzed persulfate reaction, any cerium with the americium will be oxidized t o cerium(1V). When the rare earths are precipitated as fluorides, the americium(V1) will remain in solution and the cerium(1V) will be only partially precipitated as the fluoride. The rest of the cerium(1V) will follow the americium onto the HT-200 filter. Again, the additional mass will contribute to poor resolution of the americium peaks. Even though the thorium determination is placed a t the end of this procedure, the thorium determination should be completed on the same day thorium is separated from barium sulfate. This is where thorium is separated from its radium daughters. Any time delay in the determination would allow the daughters to grow back and interfere spectrally with thorium. No more than 50 pg of uranium and 100 wg of thorium should be taken for analysis. The mass from larger quantities of either element will cause poor resolution by self-adsorption. As mentioned above, radium is separated from the other nuclides on barium sulfate precipitated from a DTPA solution. If barium-133 is introduced as a tracer for radium at the beginning of this procedure, the radium can be determined with a de-emanation procedure (8).
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Anal. Chem. 1986, 58, 1241-1245
Both glass beads and silicon carbide boiling chips break up in the high-temperature fusion used in this procedure but the glass beads are free of metals that form insoluble fluotides. The silicon carbide boiling chips are not. Neodymium fluoride was preferred as a carrier over either cerium fluoride or lanthanum fluoride because neodymium is more soluble in the small pyrosulfate fusions used throughout this procedure. The lanthanum reagents seemed to have more radioactive contaminants in them than neodymium. However, if the neodymium or cerium reagents require purification, it can be done ( I ) . In the uranium, plutonium, and americium determinations, neodymium must be added to fluoride solutions to coprecipitate the nuclides. T o do this, rapid mixing and slow precipitation are required. Precipitation before complete mixing must be avoided as much as possible. Neodymium precipitates much slower than either cerium or lanthanum. With only 50 bg of neodymium being added, the precipitation appears to be a homogeneous one. Precipitations of this kind can be more easily observed in a darkened room with a flashlight beam directed up through the bottom of the clear 50-mL roundbottomed polycarbonate centrifuge tube (Tyndall beam effect). The HT-100 filters used previously (6) are no longer available. However, the HT-200 filter, with twice the pore size, can be made to work as well if the substrate suspension is placed in a sonic bath for 15 min each day before use. This treatment probably breaks up the particles of neodymium fluoride and the finely divided precipitate plugs the pores in the HT-200 filter and helps it to have the apparent pore size of an HT-100 filter.
If a sonic bath is not available, HT-200 filters will work nicely if the vacuum is decreased to give a flow rate of 1drop/s through the filter. Nothing else needs to be changed. The recoveries ranged from 85% to 95%. The resolution for full width a t half maximum (fwhm) was between 60 and 70 keV. Decontamination factors ranged between lo3 and lo4.
ACKNOWLEDGMENT The author wishes to thank K. W. Puphal for useful suggestions in the development of this procedure. The author also thanks R. L. Williams for the many a spectrometry measurements needed to complete this work. Registry No. DTPA, 67-43-6; neodymium fluoride, 13709-42-7; ceric iodate, 13813-99-5;thorium, 7440-29-1;uranium, 7440-61-1; plutonium, 7440-07-5; americium, 7440-35-9.
LITERATURE CITED Bernabee, R. P.; Perclval, D. R.; Hlndman, F. D. Anal. Chem. 1980, 52, 2351-2353. Sill, C. W.; Puphal, K. W.; Hindman, F. D. Anal. Chem. 1974, 4 6 , 1725-1737.
Filer, T. D. Anal. Chem. 1974, 4 6 , 608-610. Moore, F. L. Anal. Chem. 1983, 35, 715-719. Sill, C. W. Anal. Chem. 1974, 46, 1426-1431. Hindman, F, D. Anal. Chem. 1983, 66, 2460-2461. Puphal, K. W., Department of Energy, IDO-12096, 1982. Martin, D. B., private communication, Radiological and Environmental Sclences Laboratory, Department of Energy, 550 Second St., Idaho Falls, I D 83401, November 15, 1980.
RECEIVED for review August 12, 1985. Resubmitted October 8, 1985. Accepted December 24, 1985. Use of commercial product names is for accuracy in technical reporting and does not constitute endorsement of the product by the United States Government.
Drug Antibody Measurement by Homogeneous Enzyme Immunoassay with Amperometric Detection Celeste A. Broyles and Garry A. Rechnitz*
Department of Chemistry, University of Delaware, Newark, Delaware 19716
A homogeneous enzyme Immunoassay Is described for the determination of drug antlbodles. The method Is based upon inhlbitlon of the enzyme actlvity of an enzyme-antigen conjugate by the correspondlng antlbody that Is to be measured. The activity Is monitored by amperometrlc detection of fhe rate of NADH oxidation at a platinum electrode. The technique Is Illustrated by uslng the lldocalne-antl-lldocalne system and ylelds callbration curves at nanogram levels of antlbody with absolute sensltlvlty dependent upon the original amount of enzyme-antlgen conjugate. Interferences such as protein adsorption and antl-qulnidine cross-reactivity are shown to have mlnlmal effect on the assay, whlch has a precislon of 5%.
The determination of immunochemicals, both qualitative and quantitative, continues to engage researchers in diverse branches of chemistry and biology (1,2). Radioimmunoassay and advances in immunodiffusion techniques have increased the sensitivity of analytical methodology to detect antibodies 0003-2700/86/0358-1241$01.50/0
and antigens (3, 4). A more recent development, enzyme immunoassay (EIA) (5) has found wide application. Numerous variations of EIA, such as enzyme-linked immunosorbent assay (6)for determining both antibodies and antigens and enzyme multiplied immunoassay technique (EMIT) (7) for measuring antigens, have been described. Immunoassays are categorized as either heterogeneous, requiring a separation step during the analytical procedure, or the more desirable homogeneous assay, involving no separation step. While homogeneous EIA (HEIA) has been successfully employed for determining haptens (7), it has been less widely applied to antibody measurements, especially at the nanogram levels of interest. In this paper, we propose an electrochemical homogeneous enzyme immunoassay for drug antibodies using amperometric detection. Electrochemical homogeneous assays for antibodies have been reported using ion-selective electrodes with complement-induced lysis of liposomes (8)and sheep red blood cells (9) and using a Ag2S electrode (IO)to monitor denaturation of the protein. None of these assays was shown to be suitable at nanogram per milliliter concentrations, and all are rather 0 1986 American Chemlcal Society
