1015 TESTS OF GROWTH HORMONE RESERVE
SIR,-I. Gil-Ad and colleagues (Aug. 11, p.278) highlight a number of important principles which require consideration when comparing the efficacy of tests of growth hormone (GH) reserve with insulin induced hypoglycaemia. They have convincingly demonstrated a GH response to oral clonidine but then conclude from their data that clonidine is a more potent stimulus of GH release than insulin and may, therefore, be especially useful in the diagnosis of GH-deficient states. More information about the patients and results are necessary before this claim can be accepted. From the clinical viewpoint of substantiating or refuting a diagnosis of GH deficiency, the only useful measurement is the peak GH level attained after stimulation. Gil-Ad et al. do not provide individual peak values after clonidine and insulin stimulation, and this omission and the much greater SEMs after clonidine than after insulin or arginine mean that we cannot exclude the possibility that some "normal" patients may have failed to achieve, after clonidine administration, a peak GH level representing adequate GH reserve for the laboratory concerned. All tests of GH reserve may fail to induce adequate peak responses in normal subjects, and such patients can only be assumed to have adequate GH reserves after examination of the height and height velocity percentiles; however, Gil-Ad et al. cite no auxological measurements, and it would be especially interesting to know the height and height velocity percentiles, on and off GH treatment, of those in the hypopituitary group. Clonidine, like insulin, arginine, and levodopa is failing to release GH in these patients, some of whom may have adequate GH reserves, especially if they are prepubertal. In evaluating GH response to intravenous insulin the criteria of adequate hypoglycaemia must be defined. In a recent study in our department, in which 0.2 units/kg body weight (double the dose in Gil-Ad’s study) of soluble insulin was administered i.v. to sixty short adolescent subjects, only fortyeight achieved either a blood-glucose below 2.3 mmol/1 or a 60% reduction of glucose from the pre-test value, these being our criteria of satisfactory hypoglycxmia, correlating well with an incremental rise of serum-cortisol of >250 nmol/1. Unless adequate hypoglycsemia was achieved a true comparison of the two tests cannot be done-and this point may, in part, explain the discrepancy between Gil-Ad’s results and those of Dr Ferrari and colleagues (Oct. 13, p.796). We accept that the subjects did not seem anxious but we feel that measurement of serum prolactin levels is not a satisfactory way of assessing "stress": prolactin levels do rise due to the stress of major surgery or insulin-induced hypoglycaemia1 but they increase in only a small proportion of patients stressed by hospital attendance or by venepuncture.2,3 Since the prolactin responses to stimulation seem greater in females than in males2 it is a pity that prolactin was measured only in males. We have compared GH responses to i.v. metoclopramide and insulin in adolescent males and preferred to use basal serum cortisol rather than prolactin, as a biochemical index of stress4 because the adrenal response to stress is universally accepted.5 University Departments of Medicine and Clinical Biochemistry,
Royal Infirmary, Glasgow G4 0SF
H. N. COHEN G. H. BEASTALL J. A. THOMSON
1. Noel
GL, Suh HK, Stone JG, Frantz AG. Human prolactin and growth horrelease during surgery and other conditions of stress. J Clin Endocrinol Metab 1972; 35: 840-51. 2. Cowden EA, Ratcliffe WA, Beastall GH, Ratcliffe JG. Laboratory assessment of prolactin status. Ann Clin Biochem 1979; 16: 113-21. 3. Koninekx P. Stress hyperprolactinæmia in clinical practice. Lancet 1978; i: mone
273. 4. Cohen HN,
Hay ID, Thomson JA, Logue F, Ratcliffe WA, Beastall GH. Metoclopramide stimulation: A test of growth hormone reserve in adolesmales. Clin Endocrinol 1979; 11: 89-93. CL. Adrenal steroids and disease. London: Pitman Medical, 1972: 199-226.
cent
5
Cope
ANTIBODY INTERFERENCE IN RADIOIMMUNOASSAYS
SIR,-Gunilla Hedenborg and colleagues (Oct. 6, p. 755) report the interesting observation that heterophilic antibodies against rabbit immunoglobulin may cause falsely high results in a TSH radioimmunoassay. Other examples of antibody interference may go unrecognised. For instance, about a third of human sera gave falsely high results in an assay for the milk protein lactalbumin because they contained a cross-reacting antibody to bovine lactalbumin which binds the 1251-labelled human lactalbumin tracer. 1,2 Presumably the antibody arises as a result of dietary exposure. Interference can be prevented by adding bovine lactalbumin since the assay antibody (rabbit anti-human lactalbumin) recognises antigenic sites present on the human but not the bovine protein. It is well known that insulin antibodies can seriously affect insulin radioimmunoassays.3 In other instances the presence of antibodies reacting with one of the assay components may not be suspected. Such a possibility is worth considering during the development of new assays, especially if the results seem anomalous. Department of Therapeutics and Clinical Pharmacology, Medical School, University of Birmingham, Birmingham B15 2TH
K. L. WOODS
ACTUAL OR STANDARD BICARBONATE?
SIR,-May I confirm Dr Howorth’s explanation (Oct. 20, p. 849) that blood-gas analysers measure bicarbonate with a preprogrammed calculator. It is a simple matter to check the accuracy of this unit by substituting recorded indices in the following formula based on Weil:4 Conc. of hydrion (nmol I"’)xactual bicarbonate (mmol 1’’) ——————————————————————————————.—————24
PC02 (mm Hg) In practice, the figure 24 should not vary by more than ± 1.0. This formula seems to be used by the manufacturers and assumes, as Dr Howorth states, that the pK remains unchanged in the presence of quite wide variation of the other factors. Department of Biochemistry, Morriston Hospital, Swansea, SA6 6NL
E. B. LOVE
PLASMA HYDROLASES IN CYSTIC FIBROSIS
SIR,-Dr Hosli and Esther Vogt (Sept. 15, p. 543) reported that the a-mannosidase and acid phosphatase activities of cystic fibrosis (CF) plasma assayed at pH S-4 are abnormally thermolabile at 411and 36-3°C, respectively, and that the levels of residual activity after heat inactivation clearly distinguish between CF patients, CF carriers, and normal sub-
jects. We have repeated these experiments under identical conditions and found no differences between the three groups. After heating for 200 min at the given temperatures, the ranges of residual activity in diluted plasma were as shown in the table. In additional experiments, no differences were observed when 1. Woods KL, Heath DA. The interference of endogenous antibodies to bovine lactalbumin in the radioimmunoassay of human lactalbumin in serum. Clin Chim Acta 1978; 84: 207-11. 2. Stevens U, Laurence DJR, Ormerod MG. Antibodies to lactalbumin interfere with its radioimmunoassay in human plasma. Clin Chim Acta 1978; 87: 149-57. 3. Moxness KE, Molnar GD, Taylor WF, Owen CA, Ackerman E, Rosevear JW. Studies of diabetic instability. I. Immunoassay of human insulin in plasma containing antibodies to pork and beef insulins. Metabolism 1971; 20: 1074-82. 4. Weil WB. Acid-base phenomena and the hydrogen ion. J Pediat 1973; 83: 359-71.
SIR,-I. Gil-Ad and colleagues (Aug. 11, p.278) highlight a number of important principles which require consideration when comparing the efficacy of tests of growth hormone (GH) reserve with insulin induced hypoglycaemia. They have convincingly demonstrated a GH response to oral clonidine but then conclude from their data that clonidine is a more potent stimulus of GH release than insulin and may, therefore, be especially useful in the diagnosis of GH-deficient states. More information about the patients and results are necessary before this claim can be accepted. From the clinical viewpoint of substantiating or refuting a diagnosis of GH deficiency, the only useful measurement is the peak GH level attained after stimulation. Gil-Ad et al. do not provide individual peak values after clonidine and insulin stimulation, and this omission and the much greater SEMs after clonidine than after insulin or arginine mean that we cannot exclude the possibility that some "normal" patients may have failed to achieve, after clonidine administration, a peak GH level representing adequate GH reserve for the laboratory concerned. All tests of GH reserve may fail to induce adequate peak responses in normal subjects, and such patients can only be assumed to have adequate GH reserves after examination of the height and height velocity percentiles; however, Gil-Ad et al. cite no auxological measurements, and it would be especially interesting to know the height and height velocity percentiles, on and off GH treatment, of those in the hypopituitary group. Clonidine, like insulin, arginine, and levodopa is failing to release GH in these patients, some of whom may have adequate GH reserves, especially if they are prepubertal. In evaluating GH response to intravenous insulin the criteria of adequate hypoglycaemia must be defined. In a recent study in our department, in which 0.2 units/kg body weight (double the dose in Gil-Ad’s study) of soluble insulin was administered i.v. to sixty short adolescent subjects, only fortyeight achieved either a blood-glucose below 2.3 mmol/1 or a 60% reduction of glucose from the pre-test value, these being our criteria of satisfactory hypoglycxmia, correlating well with an incremental rise of serum-cortisol of >250 nmol/1. Unless adequate hypoglycsemia was achieved a true comparison of the two tests cannot be done-and this point may, in part, explain the discrepancy between Gil-Ad’s results and those of Dr Ferrari and colleagues (Oct. 13, p.796). We accept that the subjects did not seem anxious but we feel that measurement of serum prolactin levels is not a satisfactory way of assessing "stress": prolactin levels do rise due to the stress of major surgery or insulin-induced hypoglycaemia1 but they increase in only a small proportion of patients stressed by hospital attendance or by venepuncture.2,3 Since the prolactin responses to stimulation seem greater in females than in males2 it is a pity that prolactin was measured only in males. We have compared GH responses to i.v. metoclopramide and insulin in adolescent males and preferred to use basal serum cortisol rather than prolactin, as a biochemical index of stress4 because the adrenal response to stress is universally accepted.5 University Departments of Medicine and Clinical Biochemistry,
Royal Infirmary, Glasgow G4 0SF
H. N. COHEN G. H. BEASTALL J. A. THOMSON
1. Noel
GL, Suh HK, Stone JG, Frantz AG. Human prolactin and growth horrelease during surgery and other conditions of stress. J Clin Endocrinol Metab 1972; 35: 840-51. 2. Cowden EA, Ratcliffe WA, Beastall GH, Ratcliffe JG. Laboratory assessment of prolactin status. Ann Clin Biochem 1979; 16: 113-21. 3. Koninekx P. Stress hyperprolactinæmia in clinical practice. Lancet 1978; i: mone
273. 4. Cohen HN,
Hay ID, Thomson JA, Logue F, Ratcliffe WA, Beastall GH. Metoclopramide stimulation: A test of growth hormone reserve in adolesmales. Clin Endocrinol 1979; 11: 89-93. CL. Adrenal steroids and disease. London: Pitman Medical, 1972: 199-226.
cent
5
Cope
ANTIBODY INTERFERENCE IN RADIOIMMUNOASSAYS
SIR,-Gunilla Hedenborg and colleagues (Oct. 6, p. 755) report the interesting observation that heterophilic antibodies against rabbit immunoglobulin may cause falsely high results in a TSH radioimmunoassay. Other examples of antibody interference may go unrecognised. For instance, about a third of human sera gave falsely high results in an assay for the milk protein lactalbumin because they contained a cross-reacting antibody to bovine lactalbumin which binds the 1251-labelled human lactalbumin tracer. 1,2 Presumably the antibody arises as a result of dietary exposure. Interference can be prevented by adding bovine lactalbumin since the assay antibody (rabbit anti-human lactalbumin) recognises antigenic sites present on the human but not the bovine protein. It is well known that insulin antibodies can seriously affect insulin radioimmunoassays.3 In other instances the presence of antibodies reacting with one of the assay components may not be suspected. Such a possibility is worth considering during the development of new assays, especially if the results seem anomalous. Department of Therapeutics and Clinical Pharmacology, Medical School, University of Birmingham, Birmingham B15 2TH
K. L. WOODS
ACTUAL OR STANDARD BICARBONATE?
SIR,-May I confirm Dr Howorth’s explanation (Oct. 20, p. 849) that blood-gas analysers measure bicarbonate with a preprogrammed calculator. It is a simple matter to check the accuracy of this unit by substituting recorded indices in the following formula based on Weil:4 Conc. of hydrion (nmol I"’)xactual bicarbonate (mmol 1’’) ——————————————————————————————.—————24
PC02 (mm Hg) In practice, the figure 24 should not vary by more than ± 1.0. This formula seems to be used by the manufacturers and assumes, as Dr Howorth states, that the pK remains unchanged in the presence of quite wide variation of the other factors. Department of Biochemistry, Morriston Hospital, Swansea, SA6 6NL
E. B. LOVE
PLASMA HYDROLASES IN CYSTIC FIBROSIS
SIR,-Dr Hosli and Esther Vogt (Sept. 15, p. 543) reported that the a-mannosidase and acid phosphatase activities of cystic fibrosis (CF) plasma assayed at pH S-4 are abnormally thermolabile at 411and 36-3°C, respectively, and that the levels of residual activity after heat inactivation clearly distinguish between CF patients, CF carriers, and normal sub-
jects. We have repeated these experiments under identical conditions and found no differences between the three groups. After heating for 200 min at the given temperatures, the ranges of residual activity in diluted plasma were as shown in the table. In additional experiments, no differences were observed when 1. Woods KL, Heath DA. The interference of endogenous antibodies to bovine lactalbumin in the radioimmunoassay of human lactalbumin in serum. Clin Chim Acta 1978; 84: 207-11. 2. Stevens U, Laurence DJR, Ormerod MG. Antibodies to lactalbumin interfere with its radioimmunoassay in human plasma. Clin Chim Acta 1978; 87: 149-57. 3. Moxness KE, Molnar GD, Taylor WF, Owen CA, Ackerman E, Rosevear JW. Studies of diabetic instability. I. Immunoassay of human insulin in plasma containing antibodies to pork and beef insulins. Metabolism 1971; 20: 1074-82. 4. Weil WB. Acid-base phenomena and the hydrogen ion. J Pediat 1973; 83: 359-71.
