Calcitonin (CT) assay is essential for recognizing medullary thyroid carcinoma (MTC), particularly occult familial MTC. In previous radioimmunoassays.
The technical assistance of Heike Moll and Helga Golling is gratefully acknowledged. We thank Ortrud Brand for valuable discussions. References 1. Windmueller HG, Spaeth AE. Uptake and metabolism of plasma glutamine by the small intestine. J Biol Chem 1974; 249:5070-9. 2. Fox AD, Kripke SA, DePaula J, Berman JM, Settle RG, Rombeau JL. Effect of a glutamine-supplemented enteral diet on
methotrexate-induced enterocolitis. J Parenter Enteral Nutr 1988;12:325-31. 3. Ollenschlager G, Jansen S, Schindler J, Rasokat H, SchrappeB#{225}cher M, Roth E. Plasma amino acid pattern of patients with H1V infection. Clin Chem 1988;34:1787-9. 4. Roth E, ZOchG, Schulz F, et al. Amino acid concentrations in plasma and skeletalmuscle of patients with acute hemorrhagic
necrotizing pancreatitis. Clin Chem 1985;31:1305-9. 5. Roth E, MUhlbacher F, Karner J, Steininger R, Schemper M, Funovics J. Liver amino acids in sepsis. Surgery 1985;97:436-42. 6. Redfield ER, Wright DC, Trasnont EC. The Walter Reedstaging classification. N Engi J Med 1983;314:131-2. 7. Waterlow JC, Fern EB. Free amino acidpools and theirregulation. In: Waterlow JC, Stephen JML, eds. Nitrogen metabolism
in man. London: Applied Science Publishers, 1981:1-16. 8. Adibi SA, Mercer DW. Protein digestion in human intestine as reflected in luminal, mucosal, and plasma amino acid concentrations after meal.J Clin Invest 1973;52:1586-94.
9. Roth E, MUhlbacher F, Karner J, Hamilton G, Funovics J. Free amino acid levels in muscle and liver of a patient with glucagonoma syndrome. Metabolism 1987;36:7-13. 1o_Windmueller HG. Glutamine utilization by the small intestine [Review]. Adv Enzymol 1982;53:201-37.
11. Marliss EB, Aoki TI’, PozefskyT, Most AS, Cahill GF. Muscle and splanchnic glutamine and glutamate metabolism in postabsorptive and starved man. J Clin Invest 1971;50:814-7. 12. Felig P, Wahren J, Karl I, Cerasi E, Luft R, Kipnis DM. Glutamine and glutamate metabolism in normal and diabetic subjects. Diabetes 1973;22:573-6. 13. Ollenschlager G, Roth E, Linkesch W, Jansen S, Simmel A, Modder B. Asparaginase-induced derangements of glutaminemetabolism-the pathogenetic basis for some drug-related sideeffects. Eur J Clin Invest 1988;18:512-6. 14. Roth E, Funovics J, Mtthlbacher F, et al. Metabolic disorders in severe abdominal sepsis: glutamine deficiency in skeletal muscle. Clin Nutr 1982;1:25-41. 15. Souba WW, Smith R, Wilmore DW. Glutamine metabolism by the intestinal tract [Review]. J Parenter Enthral Nutr 1985;9:60817. 16. Kovacevic Z, McGivan JD. Mitochrondrial metabolism of glutamine and glutamate and its physiological significance [Review].Physiol Rev 1983;63:547-605. 17. Ollenschlager G, Karner J, Karner-Hanusch J, Jansen S, Schindler J, Roth E. Plasma glutamate-a prognostic marker of cancer and of other immunodeficiency syndromes? Scand J Clin Lab Invest, in press.
CLIN. CHEM. 36/2, 381-383 (1990)
Monoclonal Immunoradiometric Assay of Calcitonin Improves Investigation of Familial Medullary Thyroid Carcinoma R. Perdrlsot,’ J. C. Blgorgne,2 D. Guliloteau,3 and P. JaIIet1 Calcitonin (CT) assay is essential for recognizing medullary thyroid carcinoma (MTC), particularly occult familial MTC. In previous radioimmunoassays of calcitonin, polyclonal antibodies were used. Here we evaluate a new two-site immunoradiometric assay (IRMA) of calcitonin based on use of monoclonal antibodies. We assayed samples from healthy subjects, patients with renal failure, and subjects from families affected by MTC. Basal values for healthy subjects were all 30 ngIL correspond to subjects with histologically confirmed MCT or micro-MCT. Polyclonal AlA performed in the same subjects failed to detect the moderate increase of CT that IRMA demonstrated. Preliminary results indicate that this new method may allow earlier detection of CT increase and thus improved diagnosis of MCT, particularly in familial screening. Monitoring surgical patients could also be improved by this new assay.
Addftlonal Keyphrases: screening . cancer - radioimmunoassay #{149} heritable disorders . pentagastrin stimulatIon test early detection Medullary
carcinoma ofthe thyroid (MCT), as described et al. (1), is transmitted by heredity in at least 25% of the cases. Early diagnosis and treatment of hereditary cases relies on assay of calcitonin (CT) after stimulation with pentagastrmn. Patients with MCT have been detected because ofan increased basal value or, at least, by a significant increase in CT concentrations in serum after pentagastrin ilijection. Until now, in CT assays polyclonal antibodies have been used (2-4). Motto et al. (5) recently described an immuneradiometric assay (IRMA) involving use of two monoclonal antibodies. The aim of the present study was to evaluate this new method and to attempt to answer two questions: can this IRMA recognize microscopic lesions of MCT better than the usual radioimmunoassay (RIA), and what are the criteria for interpreting results of the pentagastrmn test with mMA? by Hazard
MaterIals and Methods 1 Laboratoire Joliot-Curie and2 Service de Mddecine C, CHRU, 1 avenue H#{244}tel-Dieu, 49033 Angers Cedex, France. 3 Laboratoire de Biophysique M#{233}dicale, UER M#{233}decine, 2 Bd
Tonnell#{233}, 37032 Tours Cedex, France. Received September 21, 1989; accepted November 16, 1989.
Assays Inununoreactive CT was measured simultaneously for every patient with the monoclonal antibody IRMA and with one of the three RIAs described below, in all of which CLINICAL
CHEMISTRY,
Vol. 36, No. 2, 1990 381
polyclonal
antibodies
were used.
The methodology of the three RIAs is similar. Synthetic CT labeled with 1251 is recognized by the antibody, then the free and bound fractions of 1251-labeled CT are separated by using dextran-coated charcoal or a second antibody. The specific antibodies used in two ofthe methods are obtained from sheep immunized with synthetic monomeric CT (Ab M732 and Ab 600017); the third method is a commercial kit method (CIS International, Gifsur Yvette, France) in which rabbit antibodies (Ab3) are used. All three antibodies are directed mainly against determinants in the middle part ofthe molecule. For all three assays the lowest detectable concentration of CT is about 25 ng/L. IRMA. The assay of Motto et ii. (5) is now available commercially (ELSA-hCT; CIS Bioindustries, Gif sur Yvette, France). In it are used two monoclonal antibodies, directed against different determinants. The second antibody is labeled with 1251 A single incubation of both antibodies for 24 h is carried out in the presence of plasma heated to 56 #{176}C. The antibodies recognize the 11-17 and 24-32 amino acid regions of the monomeric CT. Only the intact monomeric form of CT may be determined with this method. The lowest detectable concentration is 2.5 ngIL. RIAs.
Subjects Healthy subjects. Basal concentrations of CT were deterin 83 control subjects, ages 7 to 60 years. A pentagastrin stimulation test (6) was performed in 18 healthy volunteers, ages 20 to 35 years; they received a slow intravenous injection of 0.5 g of pentagastrin (‘Peptavlon”; Imperial Chemical Industry Pharma, Eughien, France) per kilogram body weight during 3 mm, and CT was assayed in their blood sampled 5 mm before and 0, 3,5, and 10 mm after this injection. For RIA methods, the response is expressed as the ratio ofthe maximum peak to the basal value; the response is considered abnormal when this ratio exceeds 2.5. Patients with renal failure. The basal CT concentrations were determined in 13 chronic hemodialyzed patients without known thyroid disease. MCTpatients (with clinicaiprimitive or recurrent tumor). The CT basal concentration was determined in seven patients presenting with histologically confirmed medullary carcinoma, appearing clinically as one or several thyroid nodules. The follow-up of 21 patients who had already had surgery for MCT revealed a marked increase of CT in the first two years after surgery, indicating recurrence of the mined
disease.
Family study. We also assayed samples from six children from one family who underwent a pentagastrin stimulation test. The mother had a histologically confirmed MCT that was recognized because of a clinically detectable thyroid nodule.
MCT patients with clinically palpable tumors. In these patients the basal values were always increased, whatever the assay (RIA or IRMA), although RIA usually gave higher values than IRMA. Pentagastrin Stimulation Tests Healthy subjects. With the iRMA, nine of 18 subjects showed no increase of calcitomn after pentagastrmn administration. The other nine subjects gave a response >5 ngIL: in two the peak was
