Interference of recombinant human interferon α2b in human chorionic gonadotropin assays: a case report and clinical analysis

Introduction

Human chorionic gonadotropin (hCG) assays play a critical role in the diagnosis and management of various conditions in gynecology, particularly in confirming pregnancy (1). However, certain medications, including recombinant human interferon α2b (rhIFN-α2b), can interfere with immunoassays, leading to inconsistent results that may affect clinical decisions. In cases in which the assay results are unexpected, investigating the possibility of interference is essential to ensure diagnostic accuracy.

This case report presents a case in which inconsistent hCG results across different assay platforms raised questions about potential interference by rhIFN-α2b. By examining multiple testing platforms and identifying specific assay sensitivities, this report shows the importance of comprehensive assessment in clinical diagnostics to prevent false positives and ensure optimal patient care. We present this case in accordance with the CARE reporting checklist (available at https://acr.amegroups.com/article/view/10.21037/acr-2025-61/rc).

Case presentation

A 24-year-old female presented to the Gynecology Clinic of Maternal and Child Health Hospital of Nanning City on October 13, 2023, complaining of amenorrhea for over 40 days. The initial tests included a urine hCG test using colloidal gold immunochromatography, which was positive, and quantitative immunochromatography, which revealed an hCG level of 124 IU/L. However, her serum β-hCG measured via electrochemiluminescence was <0.200 IU/L, and her progesterone was 29.3 nmol/L. Given the inconsistency between the urine and serum hCG results, a further investigation was initiated. All procedures performed in this study were in accordance with the ethical standards of the institutional research committee and in line with the Helsinki Declaration (and its subsequent amendments). Written informed consent was obtained from the patient for the publication of the case report. A copy of the written consent is available for review by the editorial office of this journal.

Patient history and laboratory findings

The patient had a history of human papillomavirus (HPV) infection; her polymerase chain reaction results indicated that she was positive for HPV-66, but negative for other genotypes. At the time, the patient was receiving rhIFN-α2b, an antiviral and immune-modulating drug that has a prolonged half-life and limited degradation in vivo. We hypothesized that her medication might be interfering with her hCG assay results.

Additional tests revealed normal liver function, with total bilirubin of 5.90 µmol/L, direct bilirubin of 2.5 µmol/L, and alanine aminotransferase (ALT) of 6.3 U/L. A vaginal discharge examination indicated grade-II cleanliness and mild inflammation. Her ultrasound findings (Figure 1) revealed endometrial thickening with a heterogeneous echo pattern, a potential corpus luteum in the right ovary, and pelvic fluid. These findings suggested a non-pregnant status and possible assay interference.

Figure 1 Ultrasound scan showing no pregnancy.

Testing across platforms

To evaluate if there was any assay interference, we tested the patient’s samples across the four chemiluminescence platforms (i.e., Roche, Beckman, Mindray, and Abbott) and using the hCG colloidal gold method (Blue Cross Bio-Pharma Co., Beijing, China) and quantitative immunochromatography (Bioprox, Inc., Shanghai, China). At the same time, we increase rhIFN-α2b (since the drug is a gel, we dilute it 2-fold by HCG-negative serum) to carry out the test. The results are summarized in Table 1.

We know that Roche, Beckman, Mindray, and Abbott detection systems all use mouse monoclonal antibody β-hCG complexes to detect HCG, and they all detect the β subunit of HCG. However, the hCG colloidal gold method and quantitative immunochromatography detected the α subunit and the β subunit of HCG.

The Roche, Beckman, and Mindray platforms yielded negative hCG results, which was consistent with the low serum β-hCG. Conversely, the results for the Abbott platform, hCG colloidal gold method, and quantitative immunochromatography were positive, indicating rhIFN-α2b interference.

Diagnostic evaluation and follow-up

RhIFN-α2b has the effects of broad-spectrum antiviral, inhibition of cell proliferation and improvement of immune function, rhIFN-α2b binds to specific membrane receptors on the surface of human cells, and exerts its cellular activity by inducing certain enzymes, inhibiting cell proliferation, and immunomodulatory activities, such as enhancing the phagocytic activity of macrophages and improving the specific cytotoxicity of lymphocytes to target cells, as well as inhibiting viral replication in virus-infected cells. Clinically, it is widely used in the treatment of viral skin diseases such as cervical erosion, herpes zoster and genital warts caused by viral (HPV, etc.) infection. rhIFN-α2b, on the other hand, exists in the human body in its original form and has a long half-life (peaking at 12 hours and having a half-life of 3 hours). The patient had an HPV infection, for which rhIFN-α2b is a standard treatment. She received 1 mL of rhIFN-α2b gel vaginally every 2 days before bedtime for a 20-day course. The patient was still being treated with rhIFN-α2b the day before the blood draw. Based on the clinical findings and the patient’s treatment history, we concluded that the positive hCG results in the urine and certain assays were likely false positives due to rhIFN-α2b interference. No evidence of pregnancy or hCG-secreting pathology was found.

This case highlights the need to recognize potential assay interference (e.g., rhIFN-α2b interference) in patients undergoing specific treatments. If the hCG results are inconsistent, clinicians should consider using multiple platforms and clinical correlations to avoid false positives. Additionally, assay developers should design reagents to target specific epitopes, reducing the risk of interference. The patient was informed of these findings and advised to follow up if her symptoms recurred. After two courses of rhIFN-α2b treatment, the HPV retest result turned negative.

Discussion

Discussion by the Clinical Laboratory and Gynecology Department of the Maternal and Child Health Hospital of Nanning City

HCG is a highly glycated protein composed of hcgα and hcgβ subunits (1). When rhIFN-α2b exists in the human body in its original form and has a long half-life, the detection interferes with our experiments because the reagent manufacturer may not have good purification and α subunits when preparing antigens and antibodies. This case highlights an uncommon but clinically significant interference in serum β-hCG measurement due to the administration of rhIFN-α2b, which led to falsely elevated β-hCG levels. In clinical settings, β-hCG is a critical biomarker for several conditions, including pregnancy, gestational trophoblastic disease, and certain cancers (2). False-positive β-hCG results can lead to misdiagnosis, unnecessary follow-up testing, and inappropriate clinical interventions. The uniqueness of this case lies in the unexpected source of assay interference (i.e., rhIFN-α2b therapy), which has not been widely documented in the context of hCG testing.

The mechanism behind this interference may be related to the immunomodulatory effects of rhIFN-α2b, which is free of α subunit after rhIFN-α2b enters the body and is present in the human body as an α subunit (3). These antibodies can non-specifically bind to the assay antibodies, causing a falsely elevated hCG reading. In immunoassays, such as those used for β-hCG detection, drugs or heterophile antibodies are known to cause false-positive results by forming bridges between the capture and detection antibodies in an antigen-antibody assay format, even in the absence of the target analyte (4). Drugs or heterophile antibody interference is well recognized in immunoassay diagnostics; however, it is unusual for rhIFN-α2b therapy to be implicated as a triggering factor (5). Thus, this case extends our knowledge of potential interferents in hCG assays and highlights the need for heightened awareness among clinicians and laboratory staff regarding the effects of biological therapies on laboratory tests.

Clinicians need to understand the implications of such interference to avoid diagnostic pitfalls. When treating patients receiving rhIFN-α2b therapy, clinicians should maintain a high level of suspicion if the β-hCG results are unexpectedly elevated without a clear clinical correlation. A falsely elevated hCG level, if misinterpreted, could lead to extensive and unnecessary evaluations, such as imaging for suspected malignancy or inappropriate management in cases involving reproductive health (6). Further, misdiagnosis due to assay interference could have a significant psychological effect on patients, especially if they are falsely informed of a possible pregnancy or malignancy.

Discussion by the Clinical Laboratory Department, Maternal and Child Health Hospital of Nanning City, and the Clinical Laboratory Department, The People’s Hospital of Guangxi Zhuang Autonomous Region

In the course of our clinical immunoassay experiments, chemiluminescence, electrochemiluminescence, colloidal gold method, quantitative immunochromatography and other tests are generally antigen-antibody sandwich reactions, and antigen-antibody reactions will be closely related to animals, patients with autoimmune diseases, taking any monoclonal/polyclonal immunotherapy, specific proteins (even recombinant human technology derivatives), biological or similar drugs may be suspected of immunoassay interference. Immunoassay interference with various antibodies does not always result in false positives. Depending on the type of assay, a sample-specific antibody reagent may produce a false positive or false negative result. In any case, immunoassay interference may occur when the patient’s serum is increased with reagent-specific immunoglobulins (monoclonal/polyclonal immunoglobulins, heterolophyllophylaxis, rheumatic factor, paraproteins, etc.). Different platforms, blockers, serial dilutions, mass spectrometry, and other methods can be used to rule out interferences.

To reduce the risk of misinterpretation, clinicians are advised to confirm elevated β-hCG results using alternative approaches. One option is to conduct a urine hCG test, as heterophile antibodies generally do not affect urinary assays. However, in our case, the patient was still using rhIFN-α2b through the vagina, which largely contaminated the urine specimen. Another approach is to test samples across different assay platforms, as some immunoassay technologies are more robust against heterophile antibody interference. We obtained the same results by simultaneous detection of the patient’s serum and rhIFN-α2b, and we confirmed that rhIFN-α2b interfered with some platform tests. Additionally, the use of serial dilutions or the incorporation of blocking reagents can help reveal non-linear assay behavior suggestive of interference. In cases in which such interference is suspected, laboratory personnel should consider using mass spectrometry as a confirmatory test, as this method is less susceptible to antibody-related interference.

From a laboratory management perspective, this case illustrates the importance of quality control protocols that can identify assay interference in patients undergoing specific therapies. Laboratories should implement reflex testing strategies or alert systems for potential interferences, particularly when dealing with immunoassays susceptible to drugs or heterophile antibodies. Laboratory personnel and clinicians should also be trained to recognize and manage assay interference to enhance diagnostic accuracy. So as well as other biologics, rhIFN-α2b may cause potential immunoassay interference in any test regarding the antibody used in the reagent.

Discussion by the Clinical Laboratory Department and Clinicians

In addition to the immediate diagnostic considerations, the long-term monitoring of patients receiving rhIFN-α2b may be necessary to assess the persistence of interference effects and to monitor the other potential effects of this biologic agent on laboratory diagnostics. This is especially important if patients are undergoing prolonged therapy, as repeated false-positive or fluctuating hCG levels could complicate their ongoing clinical management.

Thus, this case underscores the need to adopt a nuanced approach in interpreting laboratory results in patients receiving biologic therapies, such as rhIFN-α2b. Clinicians should be aware of the potential for assay interference and collaborate closely with laboratory staff to ensure the accurate interpretation of test results. Through this case, we aim to raise awareness of the novel risk posed by rhIFN-α2b for hCG assay interference, and thus prompt further investigation into the prevalence of this effect and potential mitigation strategies. Future studies and case reports on similar occurrences may lead to the development of clearer guidelines for the management of biological agent-induced assay interference.

Conclusions

This case highlights the need for awareness of rhIFN-α2b as a potential source of assay interference. Clinicians and lab professionals should verify unexpected results in treated patients to prevent misdiagnosis and ensure optimal patient care.

Acknowledgments

None.

Footnote

Reporting Checklist: The authors have completed the CARE reporting checklist. Available at https://acr.amegroups.com/article/view/10.21037/acr-2025-61/rc

Peer Review File: Available at https://acr.amegroups.com/article/view/10.21037/acr-2025-61/prf

Funding: None.

Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://acr.amegroups.com/article/view/10.21037/acr-2025-61/coif). The authors have no conflicts of interest to declare.

Ethical Statement: The authors are accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved. All procedures performed in this study were in accordance with the ethical standards of the institutional research committee and in line with the Helsinki Declaration (and its subsequent amendments). Written informed consent was obtained from the patient for the publication of the case report. A copy of the written consent is available for review by the editorial office of this journal.

Open Access Statement: This is an Open Access article distributed in accordance with the Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International License (CC BY-NC-ND 4.0), which permits the non-commercial replication and distribution of the article with the strict proviso that no changes or edits are made and the original work is properly cited (including links to both the formal publication through the relevant DOI and the license). See: https://creativecommons.org/licenses/by-nc-nd/4.0/.

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(English Language Editor: L. Huleatt)