Batch P9 hCG contains no nicked hCG (5), <0.5% hCGb (DPC Immulite free [beta]-subunit assay on the DPC Immulite Chemiluminescence Automated Immunoassay Platform), no detectable hCGbcf, and 9% hyperglycosylated hCG, the expected proportion of hyperglycosylated hCG for this time of pregnancy (11).
Total hCG and related molecule immunoreactivity was determined in the DPC Immulite hCG assay, hCGb was measured in the DPC Immulite hCGb test, and hyperglycosylated hCG was measured using the Nichols Institute Diagnostics ITA hyperglycosylated hCG-specific test on the Advantage chemiluminescence automated platform (Nichols Institute Diagnostics) (19).
These range from Sigma C6322, which is free of all hCG structural variants (Table 3); to Scripps C0713, which is rich in nicked hCG (15% of total immunoreactivity), hCGb (5%), and urine hCGbcf (16%); to Ferring Choragon; Organon Pregnyl; and Sigma C5297, which are all rich in nicked hCG (12-14% of total immunoreactivity), hCGb (5-14%), and particularly rich in urinary hCGbcf (39-52%).
Aliquots of individual hCG preparation P9 added to serum (850 [micro]g/L), 99/688 added to serum (85 IU/L), the four preparative isoelectric focusing variants of individual hCG preparation P9 added to serum, and the common hCG structural variants added to serum [pure hCGb (230 [micro]g/L by amino acid analysis), pure nicked hCG (313 [micro]g/L), purified hyperglycosylated hCG (84 [micro]g/L), and pure hCGbcf (4.0 [micro]g/L)] were coded and blindly tested in nine commonly used commercial total hCG assays performed in 10 different clinical laboratories.
Wide variation was observed in the detection of hyperglycosylated hCG (at the extremes, 3.3-fold difference in the results among the nine assays), hCGb (at the extremes, 1.7-fold difference among the assays), nicked hCG (at the extremes, 1.5-fold difference among the assays), and hCGbcf (detected by only one of the assays).
On a molar basis, 13% of the immunoreactivity was attributable to hCGbcf and 10% to hCGb. The Hybritech Tandem hCG test did not actually detect hCGb or hCGbcf (16).
We also observed wide variability in the detection of hCGb (230 [micro]g/L) with results ranging from 189 [micro]g/L (Tosoh AIA600) to 323 [micro]g/L (Bayer Centaur).
They vary from ultrapure hCG, free of any hCG structural variants, to preparations that are primarily nicked hCG, hCGb, or hCGbcf.