Characterization of the hCG variants recognized by different hCG immunoassays: an important step toward standardization of hCG measurements.

Human chorionic gonadotropin (hCG) is a dimeric glycoprotein hormone consisting of noncovalently associated and subunits. It is produced principally by the trophoblastic cells of the placenta but may also be produced by nontrophoblastic tissues including normal pituitary and neoplastic cells. Therefore, qualitative and quantitative measurement of hCG is clinically useful in the diagnosis of normal and abnormal pregnancy and the management of patients with gestational trophoblastic disease (GTD) and other hCG-secreting malignancies. The measurement of hCG is complicated by its molecular heterogeneity in both structure and carbohydrate content. Multiple molecular variants of hCG are present in serum and urine and include intact hCG, nicked hCG (hCGn), free -subunit hCG (hCG ), nicked free -subunit hCG (hCG n), free -subunit hCG (hCG ), and the -core fragment hCG (hCG cf). In addition, hCG is differentially glycosylated in various tissues, resulting in a range of molecular forms from hypoglycosylated to hyperglycosylated. So-called hyperglycosylated hCG (hCG-h) is probably the best known of these glycosylated variants. This molecular heterogeneity has led to numerous problems, including use of a nonstandard nomenclature for hCG variants, absence of purified standards to achieve accurate calibration, and discrepant characterizations of hCG variants that are recognized by different hCG immunoassays. This, in turn, has led to misunderstandings about which hCG immunoassays are appropriately used in different clinical scenarios. For example, although intact hCG is the principal variant in serum throughout most of pregnancy (1 ), hCG-h accounts for a relatively higher proportion of total hCG in the first several weeks of gestation (2 ), and hCG is present in much lower abundance (3 ). Although most trophoblastic tumors produce intact hCG, unusually increased concentrations of the other hCG variants (particularly hCG ) can also be present (4 ). Finally, among germ cell tumors that produce hCG, 20%– 40% produce hCG alone (5 ). In 1994, in an effort to address some of these problems, the IFCC established a Working Group for the Standardization of hCG (6 ). The group was able to prepare and characterize new standards for 6 of the hCG variants described above (7 ). Additionally, they assigned nomenclature and quantified the standards using substance (molar) concentration (8 ). In 2001, the WHO Expert Committee on Biological Standardization approved the 6 standards as the 1st WHO International Reference Reagents (IRRs). Currently, hCG immunoassays are calibrated against the 3rd or 4th WHO International Standards (IS 75/537 or IS 75/589), which are impure mixtures of hCG that were assigned units based on bioactivity (70 g, corresponding to 650 IU). In contrast, the IRRs are highly purified with only IRR 99/642 (hCGn) containing 1.4% of hCG, hCG , and fragments of hCG (7 ). Clinically, the most important IRR is 99/688 (hCG), and it contains no hCGn and only negligible amounts of the other variants (7 ). It was recommended that the IRRs should be used to investigate and characterize the analytical specificity of hCG immunoassays (9 ). In this issue of Clinical Chemistry, Sturgeon et al., on behalf of the IFCC Working Group for Standardization of hCG, report their use of the new IRRs to do exactly that (10 ). As part of the UK National External Quality Assessment Service, Sturgeon et al. sent specimens containing the IRRs to 150 –240 different laboratories over a 6-year period (2001–2007) and determined the analytical specificity of 14 immunometric assays and 2 RIAs for hCG. 1 Department of Pathology and Immunology and 2 Department of Obstetrics and Gynecology, Washington University School of Medicine, St. Louis, MO; 3 Department of Pathology, University of Utah Health Sciences Center, Salt Lake City, UT. * Address correspondence to this author at: Washington University School of Medicine, Department of Pathology and Immunology, Box 8118, 660 S. Euclid, St. Louis, MO 63110. Fax 314-362-1461; e-mail [email protected]. Received May 5, 2009; accepted May 14, 2009. Previously published online at DOI: 10.1373/clinchem.2009.129205 4 Nonstandard abbreviations: hCG, human chorionic gonadotropin; GTD, gestational trophoblastic disease; hCGn, nicked hCG; hCG , free -subunit hCG; hCG n, nicked free -subunit hCG; hCG , free -subunit hCG; hCG cf, -core fragment hCG; hCG-h, hyperglycosylated hCG; IRR, International Reference Reagent; IS, International Standard. Clinical Chemistry 55:8 1447–1449 (2009) Editorial