Clinical Science and Molecular Medicine (1976) 51, 399-402.
A study of changes in wholebody calcium, phosphorus, sodium and nitrogen by neutron activation analysis in vivo in rats on a calciumdeficient diet K. BODDY,' R. LINDSAY,? I. HOLLOWAY,* D. A. S. SMITH,? A. ELLIOTT,* I. ROBERTSON* A N D D. GLAROS* Nuclear Medicine Unit, Scottish Universities Research and Reactor Centre, East Kilbride, and
? Department of Medicine, Bone Metabolism Research Unit, Western Infirmary, Glasgow, Scotland (Received 29 April 1976)
such changes in rats subjected to variations in the mineral constituents of their diet.
S-arY
1. A method of measuring changes in the total body content of calcium, phosphorus, nitrogen and sodium in rats by activation analysis in vivo is described. 2. The change in the body content of the elements has been measured in rats on a calcium-deficient diet and in control animals, the body nitrogen being used to represent lean body mass for normalization. 3. There were significant differences in Ca/N and P/N but not in Ca/P ratios between the animals on a deficient diet and control animals at the end of the dietary period.
Materials and methods
The total body content of phosphorus and nitrogen of twelve female Wistar rats was measured by total body neutron activation analysis in vivo, a facility developed for clinical studies (Boddy, Holloway, Elliott, Glaros, Robertson & East, 1972; Boddy, Holloway & Elliott, 1973) being used. A measurement of the natural body radioactivity of each animal was made by counting in a fixed position with a high-sensitivity shadow-shield wholebody counter (Boddy et al., 1972; Boddy, Holloway, Elliott & Robertson, 1975). The animals were then irradiated unilaterally with 14 MeV neutrons for 60 s at 12 cm SSD with a Philips type 18602 sealed tube neutron generator. Sixty seconds later, the radioactivity induced in the body, 28A1(from phosphorus) and 'N, was measured by wholebody counting for 600 s live time. On the following day, when the 28A1and 3N radioactivity had decayed to negligible levels, the total body content of calcium and sodium was measured by a similar procedure but using neutrons from a nuclear reactor as developed for clinial studies (Boddy, 1966; Boddy & Alexander, 1967). The irradiation time was 300 s and counting of the induced radioactive isotopes 49Caand 24Na began 90 s later. The body weight of each animal was noted. The animals were divided randomly into two groups of six. For 44 days, the control group r e
Key words: activation analysis in vivo, calcium, nitrogen, phosphorus, sodium. Introduction In the investigation of animals subject to changes in diet or to metabolic experiments, the measurement of changes in the total body content of calcium, phosphorus and sodium may be important. However, conventional balance techniques are tedious and inevitably are subject to cumulative errors. An alternative method is to measure the wholebody constituents in vivo at the beginning of the study and at intervals thereafter by the use of total body neutron activation analysis in vivo. The present paper reports the use of this technique to measure Correspondence: Dr Keith Boddy, Nuclear Medicine Unit, Scottish Universities Research and Reactor Centre, East Kilbride G75 OQU, Scotland.
399
K . Boddy et at.
400
ceived a normal laboratory diet containing 794 mg of Ca, 539 mg of P and 95 mg of Mg/100 g. Over the same period, the second group (the 'diet group') received a standard calcium-deficient diet (Nutritional Biochemicals, U.S.A.), containing 17.3 mg of Ca, 197 mg of P and 1012 mg of Mg/100 g. The high magnesium content of the experimental diet tended to induce diarrhoea and 2 days after commencement methylcellulose (10 g/100 g of food) was added, which effectively stopped the diarrhoea in these animals. The total body content of phosphorus, nitrogen, calcium and sodium was then remeasured as before. Paired results were compared, by using Wilcoxon's signed ranks test, and mean values with Wilcoxon's sum of ranks test.
1.144and 1.212counts/mgofCa(mean 1.178 k0.034 countslmg of Ca) and 33.757 and 34.023 counts/mg of P (mean 33.89k0.13 countslmg of P). These values are in good agreement. Effect of diet
Results Precision of the technique Repeated measurements of 13N on a rat 'phantom', comprising a polyethylene bottle containing 250 ml of ammonium nitrate solution, gave a coefficient of variation of 5 % . The difference in radioactivity counting rate per gram of element between a 'phantom' containing 150 ml of solution and that containing 250 ml was 3%. This 'weight range' essentially spanned that of the animals used. As a more direct check on the precision of the technique, at the end of the experimental period the carcasses of two of the animals were analysed chemically for calcium and phosphorus. The radioactivity counting-rates for these elements in uiuo, which are activated respectively by thermal and fast neutrons, were compared in each carcass with the body contents. The weights of the carcasses were 202 g and 247 g and gave corresponding values of
The results, expressed as the mean values and statistical significances, are summarized in Table 1. The mean body weights of the control and diet groups were not significantly different at the beginning of the study. By the end of the experiment there was a significant change in both groups although the mean weight of the group on the deficient diet was less than that of the control group. Consequently, the calcium, phosphorus and sodium results were normalized to the corresponding 3N result in each animal. As discussed below, this is equivalent to normalizing to lean body mass (Pace & Rathbun, 1945). As only relative changes were of interest, all of the results are expressed in arbitrary units representing corrected net counts normalized for neutron dose and are not molar quantities. It should be noted, particularly, that the Ca/P ratio values are also normalized arbitrary values and d o not represent molar ratios. In the initial measurements, there was no significant difference between the diet and control groups in the mean values for body weight, body nitrogen nor in the Ca/N, P/N, Na/N and Ca/P ratios. These findings were to be expected. In five of the six animals on the deficient diet there was a decrease in the Ca/N ratio over the dietary period whereas in five of the six control animals this ratio increased. However, in neither group did the change achieve statistical significance (P> 0.05). When the mean values in the two groups were
TABLE 1, Changes in body composition Mean values are shown. NS = not significant; S = significant. ~
~
Pre-diet
~
~
~
~~~
Post-diet
Group Wt.
Na/N
Ca/N
P/N
Ca/P
(€9 Diet Control
189.0 186.2 NS
Wt.
Na/N
CalN
PIN
CalP
3.035"' 2.776'" NS
2.240 2.584
3.345'" 3.866")
S
S
6.688 6,68O(l) NS
(9)
1.833 1.686 NS
2.490 2.413 NS (l)
4442 4.156 NS
6.161 5.813 NS
200.5"' 235.1'l)
S
Significant difference post-diet and pre-diet.
Changes in dietary-deficient rats
compared, the Ca/N ratio was significantly less in the dietary-deficient group than in the control animals (P< 0.05). The P/N ratio decreased significantly (P< 0.05) in both groups of animals and the mean value in the dietary-deficient group was significantly less (P< 0.05)than in the control animals. In both groups of animals, the Na/N ratio increased significantly (P< 0.05) over the dietary period, the mean increase in both cases being close to 65%. Five of the six animals in each group showed an increase in the Ca/P ratio but the increase attained statistical significance (P‘0.05) only in the control animals. The mean values of the Ca/P ratio in the two groups were not significantly different (P> 0.05). Discussion Total body activation analysis in viuo was first demonstrated in animals by Mayneord, Martin & Layne (1949)with high-energy X-rays. Subsequently, by using neutron irradiation, the technique has been successfully used in man by several groups (Anderson, Osborn, Tomlinson, Newton, Rundo, Salmon & Smith, 1964; Chamberlain, Fremlin, Peters & Philip, 1968;Palmer, Nelp, Murano & Rich, 1968; Cohn, Dombrowski & Fairchild, 1970;Boddy et al., 1972). The feasibility of measuring the total body content of various elements in small animals by neutron activation analysis in vivo has been reported by several workers (Nagai, Fujii, Muto & Inouye, 1969; Williams, Cargol, Pailthorp & Nelp, 1970; Biggin & Morgan, 1971 ; Weber & Andrews, 1972; Chasteland & Comar, 1972;Shukla & Cohn, 1973; Dombrowski, Wallach, Shukla & Cohn, 1973; Palmer, 1973). However, although a number of clinical investigations have been described in man, we are not aware of any previous analogous studies in animals applying these techniques to a specific medical problem. The aims of the present study were to explore the usefulness of the technique in sequential studies of the body composition of animals and, in particular, to examine the influence of a calcium-deficient diet on skeletal composition in rats. The precision of the method was shown to be satisfactory. There seems little doubt of the effectivenessof the technique since it can provide a simultaneous measurement of total body calcium, phosphorus, nitrogen and sodium in a semi-automated fashion without killing the animals.
401
The statistical significance of sequential changes can be evaluated and a study concluded at an appropriate time. A further important advantage of the method is that changes in mineral content can be related to body nitrogen, as the I3N counting rate, and, hence, to lean body mass (Pace & Rathbun, 1945). This is especially useful in growing animals where dietary or therapeuticregimens may cause nonspecific changes in the rate of growth without proportional changes in skeletal mass. For example, in the present study, there was a significantly smaller change in the body weight of the diet animals than in the control animals and analysis of the counting rates per se of calcium, phosphorus and sodium could be misleading. An apparent ‘deficiency’ might simply be a corollary of the reduced overall generalized growth of the diet animals. For this reason, we have preferred to interpret the results relative to nitrogen which correlated closely with lean body mass (Pace & Rathbun, 1945). The Ca/N and P/N ratios were reduced in a remarkably similar proportion in the diet animals compared with the control animals. These findings represent therefore a true failure of animals to gain mineral on the low-calcium and low-phosphorus and high-magnesium diet. There was an increase in Ca/P ratio (although these do not represent molar quantities), which was less pronounced in the diet animals. This could imply a change in skeletal or soft tissue composition with age in favour of calcium compared with phosphorus, and the effect would be less marked in the animals on a diet relatively deficient in calcium. There was a marked change in the Na/N ratio of about 65% over the dietary period, which was similar in both the diet and the control animals. The dietary intake of sodium was similar in the two groups. However, the method measures the total body content and does not distinguish between uptake into soft tissue and bone, which might have been different in the two groups of animals. Acknowledgments This work was supported by grants and studentships (A.E. and I.R.) from the Medical Research Council and financial assistance from the .National Fund for Research into Crippling Diseases, which are gratefully acknowledged. We thank Professor G . M. Wilson and Professor H. W. Wilson for their interest and encouragement.
402
K. Boddy et al.
References ANDERSON, J., OSBORN, S.B., TOMLINSON, R.W.S., NEWTON, D., RUNDO,J., SALMON, L. & SMITH,J.W. (1964) Neutron activation analysis in man in vivo: a new technique in medical investigation. Lancer, ii, 1201-1205. BIGGIN,H.C. & MORGAN, W.D. (1971) Fast neutron activation analysis of the major body elements. Journal of Nuclear Medicine, 12, 808-814. BODDY,K. (1966) In vivo activation analysis of iodine in the thyroid gland-a preliminary study. In: Proceedings of the 7th Syniposiuni on Radioactive Isotopes in Clinical Medicine and Research, p. 377. Urban and Schwarzenberg, Munich. BODDY, K. &ALEXANDER, W.D. (1967) Clinical experience of in vivo activation analysis of iodine in the thyroid gland: an assessment of the problems. In: Proceedings of the Syniposiurn on Nuclear Activation Techniques in the Life Sciences, p. 583. I.A.E.A., Vienna. BODDY,K., HOLLOWAY, I. & ELLIOTT,A. (1973) Preliminary results of measuring total body calcium with a new facility for total body in vivo activation analysis. In: Proceedings of the Panel on in vivo Neutron Activation Analysis, p. 163. I.A.E.A., Vienna. BODDY,K., HOLLOWAY, I., ELLIOTT,A., GLAROS,D., ROBERTSON, I. & EAST,B.W. (1972) Low cost facilities for partial body and total body in vivo activation analysis in the clinical environment. In: Proceedings of the Symposium on Niiclear Acrivation Techniques in the Life Sciences, p. 589. I.A.E.A., Vienna. BODDY,K., HOLLOWAY, I., ELLIOTT,A. & ROBERTSON, I. (1975) A high sensitivity dual-detector shadow-shield whole-body counter for clinical use, particularly total body in vivo activation analysis. Physics in Medicine and Biology, 20, 295-304. CHAMBERLAIN, M.J., FREMLIN, J.H., PETERS, D.K. & PHILIP, H. (1968) Total body calcium by whole-body neutron activation: new technique for study of bone disease. British Medical Journal, ii, 581-585. CHASTELAND, M. & COMAR,D. (1972) Dosage par radioactivation neutronique in vivo de quelques elements con-
tenus dans I'organisme du rat. International Journal of Applied Radiation and Isotopes, 23,209-218. COHN,S.H., DOMBROWSKI, C.S. & FAIRCHILD, R.G. (1970) In vivo neutron activation analysis of calcium content in man. International Journal of Applied Radiation and I.OtOpeS, 21, 127-1 37. DOMBROWSKI, C.S., WALLACH, S., SHUKLA, K.K. & COHN, S.H. (1973) Determination of whole-body magnesium by in vivo neutron activation. International Joiirnal of Nuclear Medicine and Biology, 1, 15-21. MAYNEORD, W.V., MARTIN,J.H. & LAYNE,D.A. (1949) Production ofradioactivity in animal tissues by high energy X-rays. Nature (London), 164,728-730. NAGAI,T., FUJII,I., MUTO,H. & INOUYE, T. (1969) Totalbody nitrogen and protein determined by in vivo fastneutron activation analysis. Journal of Niiclear Medicine, 10, 192-196. PACE,N. & RATHBUN, E.N. (1945) Studies on body composition. The body water and chemically combined nitrogen content in relation to fat content. Journal of Biological Chemistry, 158, 685-691. PALMER, H.E. (1973) Feasibility of determining total-body calcium in animals and humans by measuring 37Ar in expired air after neutron irradiation. Journal of Nuclear Medicine, 14, 522-527. PALMER, H.E., NELP,W.B., MURANO, R. &RICH,C.R. (1968) The feasibility of in vivo neutron activation analysis of total body calciumand other elements of body composition. Physics in Medicine and Biology, 13, 269-279. SHUKLA, K.K. & COHN,S.H. (1973) Measurement ofcalcium in rats by total body neutron activation analysis. International Journal of Nuclear Medicine and Biology, 1,73-18. WEBER, D.A. & ANDREWS, H.L. (1972) Neutron activation analysis of calcium in biological samples. Journal of Nuclear Medicine, 13, 293-299. WILLIAMS, J.L., CARGOL, L.H., PAILTHORP, K.G. & NELP, W.B. (1970) Neutron activation analysis of total-body phosphorus in mice. Journal of Nuclear Medicine, 11. 576-579.
A study of changes in wholebody calcium, phosphorus, sodium and nitrogen by neutron activation analysis in vivo in rats on a calciumdeficient diet K. BODDY,' R. LINDSAY,? I. HOLLOWAY,* D. A. S. SMITH,? A. ELLIOTT,* I. ROBERTSON* A N D D. GLAROS* Nuclear Medicine Unit, Scottish Universities Research and Reactor Centre, East Kilbride, and
? Department of Medicine, Bone Metabolism Research Unit, Western Infirmary, Glasgow, Scotland (Received 29 April 1976)
such changes in rats subjected to variations in the mineral constituents of their diet.
S-arY
1. A method of measuring changes in the total body content of calcium, phosphorus, nitrogen and sodium in rats by activation analysis in vivo is described. 2. The change in the body content of the elements has been measured in rats on a calcium-deficient diet and in control animals, the body nitrogen being used to represent lean body mass for normalization. 3. There were significant differences in Ca/N and P/N but not in Ca/P ratios between the animals on a deficient diet and control animals at the end of the dietary period.
Materials and methods
The total body content of phosphorus and nitrogen of twelve female Wistar rats was measured by total body neutron activation analysis in vivo, a facility developed for clinical studies (Boddy, Holloway, Elliott, Glaros, Robertson & East, 1972; Boddy, Holloway & Elliott, 1973) being used. A measurement of the natural body radioactivity of each animal was made by counting in a fixed position with a high-sensitivity shadow-shield wholebody counter (Boddy et al., 1972; Boddy, Holloway, Elliott & Robertson, 1975). The animals were then irradiated unilaterally with 14 MeV neutrons for 60 s at 12 cm SSD with a Philips type 18602 sealed tube neutron generator. Sixty seconds later, the radioactivity induced in the body, 28A1(from phosphorus) and 'N, was measured by wholebody counting for 600 s live time. On the following day, when the 28A1and 3N radioactivity had decayed to negligible levels, the total body content of calcium and sodium was measured by a similar procedure but using neutrons from a nuclear reactor as developed for clinial studies (Boddy, 1966; Boddy & Alexander, 1967). The irradiation time was 300 s and counting of the induced radioactive isotopes 49Caand 24Na began 90 s later. The body weight of each animal was noted. The animals were divided randomly into two groups of six. For 44 days, the control group r e
Key words: activation analysis in vivo, calcium, nitrogen, phosphorus, sodium. Introduction In the investigation of animals subject to changes in diet or to metabolic experiments, the measurement of changes in the total body content of calcium, phosphorus and sodium may be important. However, conventional balance techniques are tedious and inevitably are subject to cumulative errors. An alternative method is to measure the wholebody constituents in vivo at the beginning of the study and at intervals thereafter by the use of total body neutron activation analysis in vivo. The present paper reports the use of this technique to measure Correspondence: Dr Keith Boddy, Nuclear Medicine Unit, Scottish Universities Research and Reactor Centre, East Kilbride G75 OQU, Scotland.
399
K . Boddy et at.
400
ceived a normal laboratory diet containing 794 mg of Ca, 539 mg of P and 95 mg of Mg/100 g. Over the same period, the second group (the 'diet group') received a standard calcium-deficient diet (Nutritional Biochemicals, U.S.A.), containing 17.3 mg of Ca, 197 mg of P and 1012 mg of Mg/100 g. The high magnesium content of the experimental diet tended to induce diarrhoea and 2 days after commencement methylcellulose (10 g/100 g of food) was added, which effectively stopped the diarrhoea in these animals. The total body content of phosphorus, nitrogen, calcium and sodium was then remeasured as before. Paired results were compared, by using Wilcoxon's signed ranks test, and mean values with Wilcoxon's sum of ranks test.
1.144and 1.212counts/mgofCa(mean 1.178 k0.034 countslmg of Ca) and 33.757 and 34.023 counts/mg of P (mean 33.89k0.13 countslmg of P). These values are in good agreement. Effect of diet
Results Precision of the technique Repeated measurements of 13N on a rat 'phantom', comprising a polyethylene bottle containing 250 ml of ammonium nitrate solution, gave a coefficient of variation of 5 % . The difference in radioactivity counting rate per gram of element between a 'phantom' containing 150 ml of solution and that containing 250 ml was 3%. This 'weight range' essentially spanned that of the animals used. As a more direct check on the precision of the technique, at the end of the experimental period the carcasses of two of the animals were analysed chemically for calcium and phosphorus. The radioactivity counting-rates for these elements in uiuo, which are activated respectively by thermal and fast neutrons, were compared in each carcass with the body contents. The weights of the carcasses were 202 g and 247 g and gave corresponding values of
The results, expressed as the mean values and statistical significances, are summarized in Table 1. The mean body weights of the control and diet groups were not significantly different at the beginning of the study. By the end of the experiment there was a significant change in both groups although the mean weight of the group on the deficient diet was less than that of the control group. Consequently, the calcium, phosphorus and sodium results were normalized to the corresponding 3N result in each animal. As discussed below, this is equivalent to normalizing to lean body mass (Pace & Rathbun, 1945). As only relative changes were of interest, all of the results are expressed in arbitrary units representing corrected net counts normalized for neutron dose and are not molar quantities. It should be noted, particularly, that the Ca/P ratio values are also normalized arbitrary values and d o not represent molar ratios. In the initial measurements, there was no significant difference between the diet and control groups in the mean values for body weight, body nitrogen nor in the Ca/N, P/N, Na/N and Ca/P ratios. These findings were to be expected. In five of the six animals on the deficient diet there was a decrease in the Ca/N ratio over the dietary period whereas in five of the six control animals this ratio increased. However, in neither group did the change achieve statistical significance (P> 0.05). When the mean values in the two groups were
TABLE 1, Changes in body composition Mean values are shown. NS = not significant; S = significant. ~
~
Pre-diet
~
~
~
~~~
Post-diet
Group Wt.
Na/N
Ca/N
P/N
Ca/P
(€9 Diet Control
189.0 186.2 NS
Wt.
Na/N
CalN
PIN
CalP
3.035"' 2.776'" NS
2.240 2.584
3.345'" 3.866")
S
S
6.688 6,68O(l) NS
(9)
1.833 1.686 NS
2.490 2.413 NS (l)
4442 4.156 NS
6.161 5.813 NS
200.5"' 235.1'l)
S
Significant difference post-diet and pre-diet.
Changes in dietary-deficient rats
compared, the Ca/N ratio was significantly less in the dietary-deficient group than in the control animals (P< 0.05). The P/N ratio decreased significantly (P< 0.05) in both groups of animals and the mean value in the dietary-deficient group was significantly less (P< 0.05)than in the control animals. In both groups of animals, the Na/N ratio increased significantly (P< 0.05) over the dietary period, the mean increase in both cases being close to 65%. Five of the six animals in each group showed an increase in the Ca/P ratio but the increase attained statistical significance (P‘0.05) only in the control animals. The mean values of the Ca/P ratio in the two groups were not significantly different (P> 0.05). Discussion Total body activation analysis in viuo was first demonstrated in animals by Mayneord, Martin & Layne (1949)with high-energy X-rays. Subsequently, by using neutron irradiation, the technique has been successfully used in man by several groups (Anderson, Osborn, Tomlinson, Newton, Rundo, Salmon & Smith, 1964; Chamberlain, Fremlin, Peters & Philip, 1968;Palmer, Nelp, Murano & Rich, 1968; Cohn, Dombrowski & Fairchild, 1970;Boddy et al., 1972). The feasibility of measuring the total body content of various elements in small animals by neutron activation analysis in vivo has been reported by several workers (Nagai, Fujii, Muto & Inouye, 1969; Williams, Cargol, Pailthorp & Nelp, 1970; Biggin & Morgan, 1971 ; Weber & Andrews, 1972; Chasteland & Comar, 1972;Shukla & Cohn, 1973; Dombrowski, Wallach, Shukla & Cohn, 1973; Palmer, 1973). However, although a number of clinical investigations have been described in man, we are not aware of any previous analogous studies in animals applying these techniques to a specific medical problem. The aims of the present study were to explore the usefulness of the technique in sequential studies of the body composition of animals and, in particular, to examine the influence of a calcium-deficient diet on skeletal composition in rats. The precision of the method was shown to be satisfactory. There seems little doubt of the effectivenessof the technique since it can provide a simultaneous measurement of total body calcium, phosphorus, nitrogen and sodium in a semi-automated fashion without killing the animals.
401
The statistical significance of sequential changes can be evaluated and a study concluded at an appropriate time. A further important advantage of the method is that changes in mineral content can be related to body nitrogen, as the I3N counting rate, and, hence, to lean body mass (Pace & Rathbun, 1945). This is especially useful in growing animals where dietary or therapeuticregimens may cause nonspecific changes in the rate of growth without proportional changes in skeletal mass. For example, in the present study, there was a significantly smaller change in the body weight of the diet animals than in the control animals and analysis of the counting rates per se of calcium, phosphorus and sodium could be misleading. An apparent ‘deficiency’ might simply be a corollary of the reduced overall generalized growth of the diet animals. For this reason, we have preferred to interpret the results relative to nitrogen which correlated closely with lean body mass (Pace & Rathbun, 1945). The Ca/N and P/N ratios were reduced in a remarkably similar proportion in the diet animals compared with the control animals. These findings represent therefore a true failure of animals to gain mineral on the low-calcium and low-phosphorus and high-magnesium diet. There was an increase in Ca/P ratio (although these do not represent molar quantities), which was less pronounced in the diet animals. This could imply a change in skeletal or soft tissue composition with age in favour of calcium compared with phosphorus, and the effect would be less marked in the animals on a diet relatively deficient in calcium. There was a marked change in the Na/N ratio of about 65% over the dietary period, which was similar in both the diet and the control animals. The dietary intake of sodium was similar in the two groups. However, the method measures the total body content and does not distinguish between uptake into soft tissue and bone, which might have been different in the two groups of animals. Acknowledgments This work was supported by grants and studentships (A.E. and I.R.) from the Medical Research Council and financial assistance from the .National Fund for Research into Crippling Diseases, which are gratefully acknowledged. We thank Professor G . M. Wilson and Professor H. W. Wilson for their interest and encouragement.
402
K. Boddy et al.
References ANDERSON, J., OSBORN, S.B., TOMLINSON, R.W.S., NEWTON, D., RUNDO,J., SALMON, L. & SMITH,J.W. (1964) Neutron activation analysis in man in vivo: a new technique in medical investigation. Lancer, ii, 1201-1205. BIGGIN,H.C. & MORGAN, W.D. (1971) Fast neutron activation analysis of the major body elements. Journal of Nuclear Medicine, 12, 808-814. BODDY,K. (1966) In vivo activation analysis of iodine in the thyroid gland-a preliminary study. In: Proceedings of the 7th Syniposiuni on Radioactive Isotopes in Clinical Medicine and Research, p. 377. Urban and Schwarzenberg, Munich. BODDY, K. &ALEXANDER, W.D. (1967) Clinical experience of in vivo activation analysis of iodine in the thyroid gland: an assessment of the problems. In: Proceedings of the Syniposiurn on Nuclear Activation Techniques in the Life Sciences, p. 583. I.A.E.A., Vienna. BODDY,K., HOLLOWAY, I. & ELLIOTT,A. (1973) Preliminary results of measuring total body calcium with a new facility for total body in vivo activation analysis. In: Proceedings of the Panel on in vivo Neutron Activation Analysis, p. 163. I.A.E.A., Vienna. BODDY,K., HOLLOWAY, I., ELLIOTT,A., GLAROS,D., ROBERTSON, I. & EAST,B.W. (1972) Low cost facilities for partial body and total body in vivo activation analysis in the clinical environment. In: Proceedings of the Symposium on Niiclear Acrivation Techniques in the Life Sciences, p. 589. I.A.E.A., Vienna. BODDY,K., HOLLOWAY, I., ELLIOTT,A. & ROBERTSON, I. (1975) A high sensitivity dual-detector shadow-shield whole-body counter for clinical use, particularly total body in vivo activation analysis. Physics in Medicine and Biology, 20, 295-304. CHAMBERLAIN, M.J., FREMLIN, J.H., PETERS, D.K. & PHILIP, H. (1968) Total body calcium by whole-body neutron activation: new technique for study of bone disease. British Medical Journal, ii, 581-585. CHASTELAND, M. & COMAR,D. (1972) Dosage par radioactivation neutronique in vivo de quelques elements con-
tenus dans I'organisme du rat. International Journal of Applied Radiation and Isotopes, 23,209-218. COHN,S.H., DOMBROWSKI, C.S. & FAIRCHILD, R.G. (1970) In vivo neutron activation analysis of calcium content in man. International Journal of Applied Radiation and I.OtOpeS, 21, 127-1 37. DOMBROWSKI, C.S., WALLACH, S., SHUKLA, K.K. & COHN, S.H. (1973) Determination of whole-body magnesium by in vivo neutron activation. International Joiirnal of Nuclear Medicine and Biology, 1, 15-21. MAYNEORD, W.V., MARTIN,J.H. & LAYNE,D.A. (1949) Production ofradioactivity in animal tissues by high energy X-rays. Nature (London), 164,728-730. NAGAI,T., FUJII,I., MUTO,H. & INOUYE, T. (1969) Totalbody nitrogen and protein determined by in vivo fastneutron activation analysis. Journal of Niiclear Medicine, 10, 192-196. PACE,N. & RATHBUN, E.N. (1945) Studies on body composition. The body water and chemically combined nitrogen content in relation to fat content. Journal of Biological Chemistry, 158, 685-691. PALMER, H.E. (1973) Feasibility of determining total-body calcium in animals and humans by measuring 37Ar in expired air after neutron irradiation. Journal of Nuclear Medicine, 14, 522-527. PALMER, H.E., NELP,W.B., MURANO, R. &RICH,C.R. (1968) The feasibility of in vivo neutron activation analysis of total body calciumand other elements of body composition. Physics in Medicine and Biology, 13, 269-279. SHUKLA, K.K. & COHN,S.H. (1973) Measurement ofcalcium in rats by total body neutron activation analysis. International Journal of Nuclear Medicine and Biology, 1,73-18. WEBER, D.A. & ANDREWS, H.L. (1972) Neutron activation analysis of calcium in biological samples. Journal of Nuclear Medicine, 13, 293-299. WILLIAMS, J.L., CARGOL, L.H., PAILTHORP, K.G. & NELP, W.B. (1970) Neutron activation analysis of total-body phosphorus in mice. Journal of Nuclear Medicine, 11. 576-579.
