Department of Laboratory Medicine
University of Washington
 
Clinical assay interference and limitations

Methodology: Abbott AxSYM Total b-hCG immunoassay (1)

LIMITATIONS OF THE PROCEDURE

This assay is capable of detecting whole molecule (intact) hCG as well as free b-hCG subunits.

For diagnostic purposes, hCG results should be used in conjunction with other data; e.g., symptoms, results of other tests, clinical impressions, etc. Ectopic pregnancy cannot be distinguished from normal pregnancy by hCG measurements alone (2,3).

Detection of very low levels of hCG does not rule out pregnancy (4). Low levels of hCG can occur in apparently healthy, nonpregnant subjects (5,6). Because hCG values double approximately every 48 hours in a normal pregnancy (7), patients with very low levels of hCG should be resampled and retested after 48 hours.

Post menopausal specimens may elicit weak positive results due to low hCG levels unrelated to pregnancy. With a weak positive result, it is good laboratory practice to resample and retest after 48 hours, or to test with an alternate hCG method.

Because of the high degree of sensitivity of the assay, specimens tested as positive during initial days after conception may later be negative due to natural termination of the pregnancy. Natural termination occurs in 22% of clinically unrecognized pregnancies and 31% of pregnancies overall (8). It is good laboratory practice to resample and retest weak positive results after an additional 48 hours.

Infrequently, hCG levels may appear consistently elevated and could be due to, but not limited to, the presence of the following (4,9,10,11,12,13):

- heterophilic antibodies
- nonspecific protein binding
- altered forms of hCG
- hCG-like substances
- trophoblastic or nontrophoblastic neoplasms.

If the hCG level is inconsistent with clinical evidence, results should be confirmed by an alternate hCG method. This may include the qualitative hCG testing of urine (11,12).

Elevated hCG levels have been associated with some abnormal physiological states (e.g., trophoblastic and nontrophoblastic neoplasms [14,15]) and should not be used in the diagnosis of these abnormal states that are not related to pregnancy.

Specimens from patients who have received preparations of Mouse Monoclonal Antibodies for diagnosis or therapy may contain Human Anti-Mouse Antibodies (HAMA). Such specimens may demonstrate either falsely elevated or falsely depressed results when tested with assay kits which employ Mouse Monoclonal Antibodies (16,17). These specimens should not be tested with the AxSYM Total b-hCG assay.

Heterophilic antibodies in human serum can react with reagent immunoglobulins, interfering with in vitro immunoassays (4,10). Patients routinely exposed to animals or to animal serum products, can be prone to this interference and anomalous values may be observed. Additional information may be required for diagnosis.

Red blood cells may interfere with AxSYM Total b-hCG assay. Remove red blood cells from specimens prior to use. Red blood cell interference may cause depressed results.

Weeks Post LMP (Last Menstrual Period)	Approximate HCG Range (mIU/mL)
3-4 weeks	9 ­ 130
4-5 weeks	75 ­ 2,600
5-6 Weeks	850 ­ 20,800
6-7 Weeks	4,000 ­ 100,200
7-12 Weeks	11,500 ­ 289,000
12-16 Weeks	18,300 ­ 137,000
16-29 Weeks (2nd Trimester)	1,400 ­ 53,000
29-41 Weeks (3rd Trimester)	940 ­ 60,000

This information was originally enclosed as a package insert for the Abbott AxSYM Total b-hCG test (February 2000), Abbott Laboratories Diagnostics division, Abbott Park, IL 60064

Bibliography

1. Felig P, Baxter JD, Broadus AE, Frohman LA, eds. Endocrinology and Metabolism (2nd Ed.). New York: McGraw-Hill Book Co., 1987:253.
2. Kadar N, DeVore G, Romero R. Discriminatory hCG Zone: Its Use in the Sonographic Evaluation for Ectopic Pregnancy. Obstet Gynecol 1981;58:156-61.
3. Kadar N, Caldwell BV, Romero R. A Method of Screening for Ectopic Pregnancy and Its Indications. Obstet Gynecol 1981;58:162-6.
4. Hussa RO. The Clinical Marker hCG. Westport, CT: Praeger Publishers, 1987:137-50.
5. Alfthan H, Haglund C, Dabek J, Stenman U-H. Concentrations of Human Choriogonadotropin, its b-subunit, and the Core Fragment of the b-subunit in Serum and Urine of Men and Non-pregnant Women. Clin Chem 1992;38:1981-7.
6. Borkowski A, Muquardt C. Human Chorionic Gonadotropin in the Plasma of Normal, Nonpregnant Subjects. N Engl J Med 1979;301:298-302.
7. Braunstein GD. HCG Testing: Volume II. Answers to Frequently Asked Questions About hCG Testing. Monograph, 97-9325, 1991. Abbott diagnostics, Educational Services. Abbott Park, IL.
8. Wilcox AJ, Weinberg CR, O'Connor JF, Baird DD, Schlatterer JP, Canfield RE, Armstrong EG, Nisula BC. Incidence of Early Loss of Pregnancy. N Engl J Med 1988;319:189-94.
9. Hussa RO, Rinke ML, Schweitzer PG. Discordant Human Chorionic Gonadotropin Results: Causes and Solutions. Obstet Gynecol 1985;65:211-9.
10. Boscato LM, Stuart MC. Heterophilic Antibodies: A Problem for All Immunoassays. Clin Chem 1988;34:27-33.
11. Cole LA. Phantom hCG and Phantom Choriocarcinoma. Gynecol Oncol 1998;71:325-9.
12. Braunstein GD. HCG Testing: Volume III Manual. An Update to Molecular Forms of hCG. Monograph, 98-0380, 1999. Abbott Diagnostics. Educational Services. Abbott Park, IL.
13. Mishalani SH, Seliktar J, Braunstein GD. Four Rapid Serum-Urine Combination Assays of Choriogonadotropin (hCG) Compared and Assessed for Their Utility in Quantitative Determinations of hCG. Clin Chem 40/10, 1944-1949 (1994).
14. Braunstein GD, Vaitukaitis JL, Carbone PP, Ross GT. Ectopic Production of Human Chorionic Gonadotropin by Neoplasms. Ann Intern Med 1973;78:39-45.
15. Hussa RO. Clinical Utility of Human Chorionic Gonadotropin and µ-Subunit Measurements. Obstet Gynecol 1982;60:1-12.
16. Primus FJ, Kelly EA, Hansen HJ, Goldenberg DM. "Sandwich"-Type Immunoassay of Carcinoembryonic Antigen in Patients Receiving Murine Monoclonal Antibodies for Diagnosis and Therapy. Clin Chem 1988;34:261-4.
17. Schroff RW, Foon KA, Beatty SM, Oldham RK, Morgan Jr AC. Human Anti-Murine Immunoglobulin Responses in Patients Receiving Monoclonal Antibody Therapy. Cancer Res 1985;45:879-85.

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