title: Macrocalcitonin Is a Novel Pitfall in the Routine of Serum Calcitonin Immunoassay.
abstract: BACKGROUND
  Calcitonin (CT) is a sensitive marker of medullary thyroid carcinoma (MTC) and is used for primary diagnosis and follow-up after thyroidectomy. However, persistently elevated CT is observed even after complete surgical removal without evidence of a recurrent or persistent tumor.
  
  
  OBJECTIVE
  To investigate the presence of assay interference in the serum CT of MTC patients who are apparently without a structural disease.
  
  
  METHODS
  We studied three index MTC cases for CT assay interference and 14 patients with metastatic MTC. The CT level was measured using an immunofluorometric assay. Screening for assay interference was performed by determination of CT levels before and after serum treatment with polyethylene glycol. Additionally, samples were analyzed by chromatography on ultra-performance liquid chromatography and protein A-Sepharose.
  
  
  RESULTS
  Patients with biochemical and structural disease showed CT mean recovery of 84.1% after polyethylene glycol treatment, whereas patients suspected of interference showed recovery from 2-7%. The elution profile on UPLC showed that the immunometric CT from these three patients behaved like a high molecular mass aggregate (>300 kDa). Additionally, when these samples were applied to the protein A-Sepharose, CT immunoreactivity was retained on the column and was only released after lowering the pH.
  
  
  CONCLUSIONS
  For the first time, our results show the presence of a novel pitfall in the CT immunoassay: "macrocalcitonin." Its etiology, frequency, and meaning remain to be defined, but its recognition is of interest and can help clinicians avoid unnecessary diagnostic investigations and treatment during the follow-up of MTC.
authors.affiliation: Thyroid Disease Center and Laboratory of Molecular and Translational Endocrinology (T.G.A., T.S.K., J.H.Y., I.S.K., S.C.L., C.P.C., M.R.D.d.S., R.M.B.M., J.G.H.V., J.R.M.M.), Division of Endocrinology, Department of Medicine, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04039-032 São Paulo, SP, Brazil; and Molecular Biology Division (T.G.A., M.C.Z.M., A.M., J.R.M.M.), Department of Biochemistry, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04044-020 São Paulo, SP, Brazil.
authors.forename: Thalita G
authors.initials: TG
authors.lastname: Alves
authors.affiliation: Thyroid Disease Center and Laboratory of Molecular and Translational Endocrinology (T.G.A., T.S.K., J.H.Y., I.S.K., S.C.L., C.P.C., M.R.D.d.S., R.M.B.M., J.G.H.V., J.R.M.M.), Division of Endocrinology, Department of Medicine, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04039-032 São Paulo, SP, Brazil; and Molecular Biology Division (T.G.A., M.C.Z.M., A.M., J.R.M.M.), Department of Biochemistry, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04044-020 São Paulo, SP, Brazil.
authors.forename: Teresa S
authors.initials: TS
authors.lastname: Kasamatsu
authors.affiliation: Thyroid Disease Center and Laboratory of Molecular and Translational Endocrinology (T.G.A., T.S.K., J.H.Y., I.S.K., S.C.L., C.P.C., M.R.D.d.S., R.M.B.M., J.G.H.V., J.R.M.M.), Division of Endocrinology, Department of Medicine, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04039-032 São Paulo, SP, Brazil; and Molecular Biology Division (T.G.A., M.C.Z.M., A.M., J.R.M.M.), Department of Biochemistry, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04044-020 São Paulo, SP, Brazil.
authors.forename: Ji H
authors.initials: JH
authors.lastname: Yang
authors.affiliation: Thyroid Disease Center and Laboratory of Molecular and Translational Endocrinology (T.G.A., T.S.K., J.H.Y., I.S.K., S.C.L., C.P.C., M.R.D.d.S., R.M.B.M., J.G.H.V., J.R.M.M.), Division of Endocrinology, Department of Medicine, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04039-032 São Paulo, SP, Brazil; and Molecular Biology Division (T.G.A., M.C.Z.M., A.M., J.R.M.M.), Department of Biochemistry, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04044-020 São Paulo, SP, Brazil.
authors.forename: Maria Cecília Z
authors.initials: MC
authors.lastname: Meneghetti
authors.affiliation: Thyroid Disease Center and Laboratory of Molecular and Translational Endocrinology (T.G.A., T.S.K., J.H.Y., I.S.K., S.C.L., C.P.C., M.R.D.d.S., R.M.B.M., J.G.H.V., J.R.M.M.), Division of Endocrinology, Department of Medicine, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04039-032 São Paulo, SP, Brazil; and Molecular Biology Division (T.G.A., M.C.Z.M., A.M., J.R.M.M.), Department of Biochemistry, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04044-020 São Paulo, SP, Brazil.
authors.forename: Aline
authors.initials: A
authors.lastname: Mendes
authors.affiliation: Thyroid Disease Center and Laboratory of Molecular and Translational Endocrinology (T.G.A., T.S.K., J.H.Y., I.S.K., S.C.L., C.P.C., M.R.D.d.S., R.M.B.M., J.G.H.V., J.R.M.M.), Division of Endocrinology, Department of Medicine, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04039-032 São Paulo, SP, Brazil; and Molecular Biology Division (T.G.A., M.C.Z.M., A.M., J.R.M.M.), Department of Biochemistry, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04044-020 São Paulo, SP, Brazil.
authors.forename: Ilda S
authors.initials: IS
authors.lastname: Kunii
authors.affiliation: Thyroid Disease Center and Laboratory of Molecular and Translational Endocrinology (T.G.A., T.S.K., J.H.Y., I.S.K., S.C.L., C.P.C., M.R.D.d.S., R.M.B.M., J.G.H.V., J.R.M.M.), Division of Endocrinology, Department of Medicine, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04039-032 São Paulo, SP, Brazil; and Molecular Biology Division (T.G.A., M.C.Z.M., A.M., J.R.M.M.), Department of Biochemistry, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04044-020 São Paulo, SP, Brazil.
authors.forename: Susan C
authors.initials: SC
authors.lastname: Lindsey
authors.affiliation: Thyroid Disease Center and Laboratory of Molecular and Translational Endocrinology (T.G.A., T.S.K., J.H.Y., I.S.K., S.C.L., C.P.C., M.R.D.d.S., R.M.B.M., J.G.H.V., J.R.M.M.), Division of Endocrinology, Department of Medicine, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04039-032 São Paulo, SP, Brazil; and Molecular Biology Division (T.G.A., M.C.Z.M., A.M., J.R.M.M.), Department of Biochemistry, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04044-020 São Paulo, SP, Brazil.
authors.forename: Cléber P
authors.initials: CP
authors.lastname: Camacho
authors.affiliation: Thyroid Disease Center and Laboratory of Molecular and Translational Endocrinology (T.G.A., T.S.K., J.H.Y., I.S.K., S.C.L., C.P.C., M.R.D.d.S., R.M.B.M., J.G.H.V., J.R.M.M.), Division of Endocrinology, Department of Medicine, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04039-032 São Paulo, SP, Brazil; and Molecular Biology Division (T.G.A., M.C.Z.M., A.M., J.R.M.M.), Department of Biochemistry, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04044-020 São Paulo, SP, Brazil.
authors.forename: Magnus R
authors.initials: MR
authors.lastname: Dias da Silva
authors.affiliation: Thyroid Disease Center and Laboratory of Molecular and Translational Endocrinology (T.G.A., T.S.K., J.H.Y., I.S.K., S.C.L., C.P.C., M.R.D.d.S., R.M.B.M., J.G.H.V., J.R.M.M.), Division of Endocrinology, Department of Medicine, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04039-032 São Paulo, SP, Brazil; and Molecular Biology Division (T.G.A., M.C.Z.M., A.M., J.R.M.M.), Department of Biochemistry, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04044-020 São Paulo, SP, Brazil.
authors.forename: Rui M B
authors.initials: RM
authors.lastname: Maciel
authors.affiliation: Thyroid Disease Center and Laboratory of Molecular and Translational Endocrinology (T.G.A., T.S.K., J.H.Y., I.S.K., S.C.L., C.P.C., M.R.D.d.S., R.M.B.M., J.G.H.V., J.R.M.M.), Division of Endocrinology, Department of Medicine, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04039-032 São Paulo, SP, Brazil; and Molecular Biology Division (T.G.A., M.C.Z.M., A.M., J.R.M.M.), Department of Biochemistry, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04044-020 São Paulo, SP, Brazil.
authors.forename: José Gilberto H
authors.initials: JG
authors.lastname: Vieira
authors.affiliation: Thyroid Disease Center and Laboratory of Molecular and Translational Endocrinology (T.G.A., T.S.K., J.H.Y., I.S.K., S.C.L., C.P.C., M.R.D.d.S., R.M.B.M., J.G.H.V., J.R.M.M.), Division of Endocrinology, Department of Medicine, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04039-032 São Paulo, SP, Brazil; and Molecular Biology Division (T.G.A., M.C.Z.M., A.M., J.R.M.M.), Department of Biochemistry, Escola Paulista de Medicina, Universidade Federal de São Paulo, 04044-020 São Paulo, SP, Brazil.
authors.forename: João Roberto M
authors.initials: JR
authors.lastname: Martins
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D002116:Calcitonin / Q000097:blood*; D018278:Carcinoma, Neuroendocrine / Q000097:blood* / Q000235:genetics / Q000601:surgery; D002851:Chromatography, High Pressure Liquid; D005189:False Positive Reactions; D005260:Female; D006044:Goiter, Nodular / Q000097:blood; D006801:Humans; D007118:Immunoassay; D007453:Iodide Peroxidase / Q000097:blood; D008297:Male; D008875:Middle Aged; D009154:Mutation / Q000235:genetics; D009362:Neoplasm Metastasis; D011498:Protein Precursors; D051096:Proto-Oncogene Proteins c-ret / Q000097:blood / Q000235:genetics; D013954:Thyroglobulin / Q000032:analysis; D013964:Thyroid Neoplasms / Q000097:blood* / Q000235:genetics / Q000601:surgery; D013965:Thyroidectomy; D055815:Young Adult
chemical_list: D011498:Protein Precursors; D002116:Calcitonin; D013954:Thyroglobulin; D007453:Iodide Peroxidase; D051096:Proto-Oncogene Proteins c-ret; C099282:RET protein, human