Heterophilic Antibody Interference Research Articles

False high testosterone of unknown reason in a clinically inconspicuous female

Abstract Background This report investigates an unusual total testosterone result in a middle-aged female. Case presentation A 40-year-old female patient was found to have high serum total testosterone of 30.82 nmol/L on a Beckman Coulter UniCel® DxI800 automatic chemiluminescence immunoanalyzer without appropriate clinical appearance. Heterophile antibody interference was considered for elevated testosterone and investigated; reanalysis of the original serum sample and liquid chromatography with tandem mass spectrometry (LC-MS/MS) were performed. Reanalysis of the original serum sample using Roche® and Siemens® immunoassays both gave a normal total testosterone level. Beckman Coulter® also excluded the possibility of interference by heterophile antibodies. Finally, LC-MS/MS showed that the total testosterone level was in the normal range. Conclusions This report highlights the importance of ruling out the interfering factors for false high values of the analyte in laboratories.

Heterophilic interference in specimens yielding false-reactive results on the Abbott 4th generation ARCHITECT HIV Ag/Ab Combo assay

BackgroundFalse-reactivity in HIV-negative specimens has been detected in HIV fourth-generation antigen/antibody or ‘combo’ assays which are able to detect both anti-HIV-1/HIV-2 antibodies and HIV-1 antigen. ObjectivesWe sought to characterize these specimens and determine the effect of heterophilic interference. Study designSpecimens previously testing as false-reactive on the Abbott ARCHITECT HIV Ag/Ab combo assay and re-tested on a different (Siemens ADVIA Centaur HIV Ag/Ab) assay. A subset of these specimens were also pre-treated with heterophilic blocking agents and re-tested on the Abbott assay. ResultsHere we report that 95% (252/264) of clinical specimens that were repeatedly reactive on the Abbott ARCHITECT HIV Ag/Ab combo assay (S/Co range, 0.94–678) were negative when re-tested on a different fourth generation HIV combo assay (Siemens ADVIA Centaur HIV Ag/Ab). All 264 samples were subsequently confirmed to be HIV negative. On a small subset (57) of specimens with available volume, pre-treatment with two different reagents (HBT; Heterophilic Blocking Tube, NABT; Non-Specific Blocking Tube) designed to block heterophilic antibody interference either eliminated (HBT) or reduced (NABT) the false reactivity when re-tested on the ARCHITECT HIV Ag/Ab combo assay. ConclusionsOur results suggest that the Abbott ARCHITECT HIV Ag/Ab combo assay can be prone to heterophilic antibody interference.

45 Human Chorionic Gonadotropin (hCG) Heterophile Interference Investigations: Experience from a Referral Laboratory

hCG quantification by immunoassay is useful in the evaluation of pregnancy and monitoring of gestational trophoblastic diseases and tumors such a germ cell tumors. Heterophile antibody (HA) interference can cause falsely elevated hCG leading to incorrect diagnosis and treatments options. When results are not consistent with the clinical picture, HA interference investigation may be requested by the physician. The purpose of this study was to (1) determine the frequency of HA interference among cases where the physician requested hCG assay interference investigation and (2) evaluate the effectiveness of commercially available blocking reagents to detect HA interferences. Physician requests for hCG HA investigation from 2008 to 2016 were reviewed (n = 112). hCG was measured using the Beckman Coulter UniCel-DxI800 as a primary method and the Siemens Immulite 2000 as the secondary method from 2008 to 2010 (n = 45). From 2014 to 2016, a Roche Diagnostics Cobas e601 was used as the primary method, and Beckman as the secondary method (n = 67). HA investigation included measurement by two immunoassays before and after treatment of samples with heterophile blocking reagent (HBR) nonmurine solution (Beckman assay) and heterophile blocking tubes (HBT) (Roche assay; Scantibodies Laboratory). The absolute difference and the percent difference between untreated and treated results were calculated. Serial dilutions were also performed (primary method) and percent recovery calculated. From the 112 physician requests, five cases of HA interference were identified. The HA frequency was 6.7% (n = 3) for the Beckman assay and 3.0% (n = 2) for the Roche assay. The range of hCG concentrations on all the evaluated samples was 0.1-2,797.0 IU/L (mean 71.1 IU/L; median: 10.2 IU/L). The presence of HA was detected using HBR/HBT reagents in three cases with a percent difference from the untreated sample of –80%, –51%, and –64% from the initial value of 13, 5.5, and 9 IU/L, respectively. The two cases not detected by HBR/HBT reagents were detected due to discrepant results with the secondary method (50.44 vs 1.74 IU/L and 56.6 vs <1.0 IU/L) and nonlinear serial dilution (recovery outside 80%-120%). HA-negative cases showed an absolute difference of ±0.9 IU/L at <10 IU/L and percent difference ≤+/–20% from the untreated at concentrations >10 IU/L for the Beckman assay; and an absolute difference of ±1.2 U/L at <10 IU/L and a ≤+/–10% difference from the untreated at concentrations >10 IU/L for the Roche assay. Frequency of HA interference was similar between the Beckman and the Roche assays (6.7 vs 3.0%, P = .357). The HBR/HBT blocking reagents failed to detect 40% of HA interference cases and should not be solely used for these investigations. Multiple strategies including the use of serial dilutions and measurement using an alternative platform are critical to identify the presence of HA.

Establishing reference intervals for hCG in postmenopausal women

BackgroundPlasma concentrations of human chorionic gonadotropin (hCG) have been shown to increase with age due to pituitary secretion. We previously recommended that an hCG cutoff of 14.0IU/L be used for women ≥55years of age. However, it remains unknown whether concentrations >14.0IU/L can be expected in women with advanced age. Our objectives were to establish plasma hCG reference intervals and correlate follicle stimulating hormone (FSH) and hCG concentrations in postmenopausal females ≥55years. MethodsResidual plasma samples from 798 women ≥55years were utilized with 303, 269, and 226 samples belonging to the age groups 55–69, 70–84, and ≥85years, respectively. FSH and hCG were measured using the Abbott ARCHITECT. All positive hCG samples (hCG ≥5IU/L) were analyzed for potential heterophile antibody interference and 3 were excluded. Electronic medical records were reviewed and patients with malignancy were excluded. Results8% (56/666) of women age≥55years had plasma hCG ≥5IU/L. There were 19, 16, and 21 patients with hCG ≥5IU/L in the age groups 55–69, 70–84, and ≥85years, respectively. The highest hCG concentrations observed in each age group were: 55–69years maximum=11.7IU/L and 97.5th percentile=9.6IU/L; 70–84years maximum=18.09IU/L, 97.5th percentile=6.2IU/L; ≥85years maximum=11.1IU/L and 97.5th percentile=10.0IU/L, and the overall 97.5th percentile=8.5IU/L for all women ≥55years of age. Neither hCG nor FSH concentrations continued to increase with age in women ≥55years. ConclusionsThe prevalence of positive hCG in women ≥55years is 8%. This study confirms our previously recommended cutoff of 14IU/L for women ≥55years of age. In women ≥55years of age, FSH concentrations do not predict hCG concentrations.

Effect of heterophilic antibody interference in ACTH immunoassay to Subclinical Cushing's Syndrome screening: A pilot study

Effect of heterophilic antibody interference in ACTH immunoassay to Subclinical Cushing's Syndrome screening: A pilot study

Heterophilic antibody interference affecting multiple hormone assays: Is it due to rheumatoid factor?

ABSTRACTAssay interference with heterophilic antibodies has been well described in literature. Rheumatoid factor is known to cause similar interference leading to falsely elevated hormone levels when measured by immunometric methods like enzyme-linked immunosorbent assay (ELISA) or multiplex immunoasays (MIA). We report a case of a 60-year-old male patient with a history of rheumatoid arthritis referred to our endocrine clinic for investigation of hypogonadism and was found to have high serum levels of LH, FSH, SHBG, Prolactin, HCG and TSH. We suspected assay interference and further tests were performed. We used Heteroblock tubes and PEG precipitation to eliminate the interference and the hormone levels post treatment were in the normal range. We believe the interference was caused by high serum levels of rheumatoid factor. Although he was treated with thyroxine for 3 years, we believe he may have been treated inappropriately as his Free T4 level was always normal despite high TSH due to assay interference. Our case illustrates the phenomenon of heterophilic antibody interference likely due to high levels of rheumatoid factor. It is essential for clinicians and endocrinologists in particular to be aware of this possibility when making treatment decisions in these groups of patients.

Methodological Issues in Cytokine Measurement in Schizophrenia.

There is mounting evidence that inflammation is a major factor in the pathophysiology of schizophrenia. Inflammatory status is commonly ascertained by measuring peripheral cytokine concentrations. An issue concerning research on inflammation and schizophrenia relates to assay methodology. Enzyme-linked immunosorbent assay (ELISA) is the most widely used and the gold standard method used to measure cytokine concentrations. ELISA has a number of limitations. Both ELISA and multiplex are limited by not being able to distinguish between bioactive and inactive molecules and the matrix and heterophilic (auto-) antibody interference. Multiplex assays when combined with gene expression analysis and flow cytometry techniques such as fluorescence-activated cell sorting may be useful to detect abnormalities in specific immune pathways. Peripheral blood mononuclear cells cultures, to evaluate in vitro lipopolysaccharide-induced cytokine production, may be a better technology than measuring cytokines in the serum. The purpose of this paper is to shed light on major methodological issues that need to be addressed in order to advance the study of cytokines in schizophrenia. We make a few recommendations on how to address these issues.

Negative heterophile antibody interference affecting iPTH

Negative heterophile antibody interference affecting iPTH

Persistent Human Chorionic Gonadotropin After Methotrexate Treatment and an Emergency Surgical Procedure for Ectopic Pregnancy.

The case study is a 33-year-old white female with persistently elevated serum human chorionic gonadotropin (hCG) levels following methotrexate treatment and emergency surgery for ectopic pregnancy. At the time of the first methotrexate dose, the serum hCG concentration was 27,995 IU/L. The laboratory was consulted 3.5 months after the surgery, because serum hCG levels had stopped declining and had leveled off to around 80 to 90 IU/L but with negative urine pregnancy tests. Laboratory studies ruled out heterophile antibody interference and hook effect by multiple methods including analysis by different serum hCG assays, treatment with heterophile antibody blocking agents, and dilution studies. Three additional doses of methotrexate over six months were required for serum hCG concentrations to decline to undetectable levels. This case illustrates challenges that may arise with serum hCG measurements in management of ectopic pregnancies. Close collaboration between the laboratory and clinical service is key for optimal patient care.

Decrease in plasma levels of α-synuclein is evident in patients with Parkinson's disease after elimination of heterophilic antibody interference.

There is substantial biochemical, pathological, and genetic evidence that α-synuclein (A-syn) is a principal molecule in the pathogenesis of Parkinson disease (PD). We previously reported that total A-syn levels in cerebrospinal fluid (CSF), measured with the specific enzyme-linked immunosorbent assay (ELISA) developed by ourselves, were decreased in patients with PD, and suggested the usefulness of A-syn in CSF and plasma as a biomarker for the diagnosis of PD. After our report, a considerable number of studies have investigated the levels A-syn in CSF and in blood, but have reported inconclusive results. Such discrepancies have often been attributed not only to the use of different antibodies in the ELISAs but also to interference from hemolysis. In this study we measured the levels of A-syn in CSF and plasma by using our own sandwich ELISA with or without heterophilic antibody (HA) inhibitor in 30 patients with PD and 58 age-matched controls. We thereby revealed that HA interfered with ELISA measurements of A-syn and are accordingly considered to be an important confounder in A-syn ELISAs. HA produced falsely exaggerated signals in A-syn ELISAs more prominently in plasma samples than in CSF samples. After elimination of HA interference, it was found that hemolysis did not have a significant effect on the signals obtained using our A-syn ELISA. Furthermore, plasma levels of A-syn were significantly lower in the PD group compared with the control group following elimination of HA interference with an HA inhibitor. Our results demonstrate that HA was a major confounder that should be controlled in A-syn ELISAs, and that plasma A-syn could be a useful biomarker for the diagnosis of PD if adequately quantified following elimination of HA interference.

Interference by Human Anti-mouse Antibodies in Immunoassays: Falsely Elevated Cardiac Troponins Leading to Negative Coronary Angiograms

Cardiac Troponins are an extremely important indicator of myocardial injury and American College of Cardiology (ACC) and American Heart Association (AHA) guidelines include Cardiac Troponins as one of the major diagnostic criteria for myocardial damage. However in some instances the presence of heterophilic antibodies like Human Anti-Mouse Antibodies (HAMA) may interfere with the sandwich assays used for the detection and quantification of cardiac troponins giving false results which can have a major impact on the management of an individual suspected to have myocardial injury. Case Report Alam et al.; ARRB, 6(5): 323-327, 2015; Article no.ARRB.2015.091 324 We present a case of a 56 year old Caucasian male who had negative coronary angiogram 2 years ago presenting with acute chest pain and significantly elevated levels of Cardiac Troponin I (cTnI). He underwent cardiac catheterization which again turned out to be completely normal. His cTnI levels continued to be consistently elevated for months after cardiac catheterization. The reason for elevated cTnI levels was due to high levels of HAMA antibodies that caused heterophilic antibody interference in the assay of cTnI, resulting in falsely elevated levels of cTnI.

Towards improved diagnosis of von Willebrand disease: Comparative evaluations of several automated von Willebrand factor antigen and activity assays

Introductionvon Willebrand disease (VWD) is reportedly the most common bleeding disorder and arises from deficiency and/or defects of von Willebrand factor (VWF). Laboratory diagnosis and typing has important management implications and requires a wide range of tests, including VWF activity and antigen, and involves differential identification of qualitative vs quantitative defects. MethodsWe have assessed several VWF antigen and activity assays (collagen binding [VWF:CB], ristocetin cofactor [VWF:RCo] and the new Siemens INNOVANCE assay [VWF:Ac], employing latex particles and gain of function recombinant glycoprotein Ib to facilitate VWF binding and agglutination without need for ristocetin) using different instrumentation, including the new Sysmex CS-5100, with a large sample test set (n=600). We included retrospective plus prospective study designs, and also evaluated desmopressin responsiveness plus differential sensitivity to high molecular weight VWF. ResultsVWF:Ag and VWF:RCo results from different methods were respectively largely comparable, although some notable differences were evident, including one high false normal VWF:Ag value (105 U/dL) on a type 3 VWD sample, possibly due to heterophile antibody interference in the latex-based CS-5100 methodology. VWF:Ac was largely comparable to VWF:RCo, but VWF:CB showed discrepant findings to both VWF:RCo and VWF:Ac with some patients, most notably patients with type 2M VWD. Conclusions(a) VWF:Ag on different platforms are largely interchangeable, as are VWF:RCo on different platforms, except for occasional (some potentially important) differences, and manufacturer recommended methods may otherwise require some assay optimization; (b) VWF:RCo and VWF:Ac are largely interchangeable, except for occasional differences that may also relate to assay design (differing optimizations); (c) VWF:CB provides an additional activity to supplement VWF:RCo or VWF:Ac activity assays, and is not interchangeable with either.

Practice trends in patients with persistent detectable thyroglobulin and negative diagnostic radioiodine whole body scans: a survey of American Thyroid Association members.

Management of patients with thyroglobulin (Tg)-positive/scan-negative thyroid cancer remains challenging. American Thyroid Association (ATA) guidelines recommend potential use of empiric (131)I therapy and various scanning modalities, but no standard for managing such cases exists. We surveyed ATA members to assess current practice in management of patients with Tg-positive/scan-negative disease. Members participated in a web-based survey of six case scenarios of Tg elevations but iodine scan negativity. A total of 288 ATA members (80% male) participated. Patient age, sex, and basal and stimulated Tg varied between the cases. Respondents were asked their opinion regarding empiric (131)I therapy use, including (131)I dose, use and duration of low-iodine diet, thyroxine withdrawal or recombinant human thyrotropin (rhTSH), and utilization of additional imaging (neck ultrasound (US) or positron emission tomography/computed tomography (PET/CT)) and reconsideration of (131)I therapy. Between 16% and 51% recommended initial use of empiric (131)I for the various scenarios. The majority chose a (131)I dose between 75 and 150 mCi, and 73% employed a low-iodine diet for two or more weeks. Preference between thyroxine withdrawal versus rhTSH was evenly split. More than 98% obtained a neck US if empiric (131)I was not given; 52-89% would proceed to PET/CT if US was negative. Only 44% used rhTSH stimulation in PET scan preparation. (131)I use was more common with stimulated Tg significantly >10 ng/mL. (131)I therapy was slightly more likely with PET-positive (56%) than PET-negative status (45%). Respondents were split regarding empiric (131)I if basal and stimulated Tg increased ≥150% over two years. Providers in North America less commonly utilized (131)I treatment than those from other areas. In the face of possible heterophilic antibody interference in the Tg assay, the majority did not recommend (131)I therapy. Empiric (131)I therapy is still utilized for patients with Tg-positive/scan-negative disease. Neck US is frequently used to further evaluate such cases as (18)FDG-PET/CT, albeit the latter is used somewhat less often. Use of (131)I therapy correlated with the degree of Tg elevation or development of Tg antibodies, and was recommended more commonly with PET-positive than PET-negative status in patients with lower Tg levels. (131)I was less commonly used by providers within North America.

Discrepant serum and urine β-hCG results due to production of β-hCG by a cribriform-morular variant of thyroid papillary carcinoma

BackgroundAlthough patients with medullary thyroid cancer are known to present with paraneoplastic hormone production, this is much less common with papillary thyroid cancer. MethodsWe present a patient with the cribriform morular variant of papillary thyroid cancer in association with familial adenomatous polyposis who developed a positive pregnancy test in the absence of known pregnancy. The patient had developed vaginal bleeding, and her laboratory testing was characterized by elevated serum human chorionic gonadotropin (β-hCG) concentrations, but negative qualitative urine results. After a thorough gynecological evaluation to exclude unexpected normal, ectopic, or molar pregnancy, we pursued an evaluation for other sources of β-hCG production. ResultsWe showed that the elevated serum β-hCG concentrations were not the result of heterophile antibody interferences, and ultimately we proved that her recurrent tumor produced the ectopic β-hCG. This is the first report of β-hCG production by papillary thyroid cancer. Thus, the possibility of ectopic production of β-hCG by papillary thyroid cancer needs to be included in the differential diagnosis of elevated hCG concentration in the absence of pregnancy. ConclusionsThis study of an unusual paraneoplastic syndrome highlights the importance of investigating discrepancies in the clinical laboratory.

A Pseudo-Hyper Triiodothyroninemia Caused by Heterophile Antibodies Interference in Radio Immunoassay

Introduction: Since mouse antibodies are commonly used in diagnostic kits, their presence including auto antibodies/heterophile antibodies in some patients' sera has been known as a potential source of interference and false immunoassay results for a long time. Case Presentation: This is a case report of a 49 year-old woman presenting with nonspecific symptoms suspected to hyperthyroidism. Her total triiodothyronine (T3) test was more than twice of the expected value, exclusively by radio immunoassay (RIA) method. Interferences due to the protein binding disorders was ruled out using radio-labeled T3, binding to the separated binding proteins by electrophoresis. Presence of auto antibody ruled out by a manual (in house) method. In this manual method, anti human IgG and IgM coated wells used for auto antibody stripping. In addition animal serum used as blocking reagent. Precipitating technique by polyethylene glycol (PEG) showed a false-positive result in T3 assay caused by a heterophile IgG class antibody. Discussion: In our review of literatures, so far, this is the first case report of false-positive interference, exclusively in total T3 assay due to Heterophile antibody.

Improvement for identification of heterophile antibody interference and AFP hook effect in immunoassays with multiplex suspension bead array system

BackgroundA variety of immunoassays including multiplex suspension bead array have been developed for tumor marker detections; however, these assays could be compromised in their sensitivity and specificity by well-known heterophile antibody interference and hook effect. MethodsUsing Luminex® multiplex suspension bead arrays, we modified protocols with two newly-developed solutions that can identify heterophile antibody interference and AFP hook effect. Effectiveness of the two solutions was assessed in serum samples from patients. ResultsConcentrations of 9 tumor markers in heterophile antibody positive samples assayed with Solution A, containing murine monoclonal antibodies and mouse serum, were significantly reduced when compared with those false high signals assayed without Solution A (all p<0.01). With incorporation of Solution H (fluorescent beads linked with AFP antigen), a new strategy for identification of AFP hook effect was established, and with this strategy AFP hook effect was identified effectively in serum samples with very high levels of AFP. ConclusionsTwo proprietary solutions improve the identification of heterophile antibody interference and AFP hook effect. With these solutions, multiplex suspension bead arrays provide more reliable testing results in tumor marker detection where complex clinical serum samples are used.

Odd thyroid function tests results: interference versus non-compliance

Despite the advent modern immunoassays, particularly with regards to TSH measurements, clinicians still encounter seemingly inexplicable results of thyroid function tests, for instance raised concentrations of free hormones (free T4 and/or free T3) in the setting of high, or at least not suppressed TSH concentrations. Though cases of TSH-oma or thyroid hormone resistance do exist, they are in fact rare, while some “strange” thyroid function tests results can be commonly explained either by a laboratory error or patient’s non-compliance with L-thyroxine treatment. In the latter case, a patient might deny any compliance problems, while a combination of raised TSH accompanied by an increased free T4 (±free T3) concentration is typically observed. This is usually a result of taking some extra doses of L-thyroxine prior to blood test, in order to catch up for previously missed doses. Confirmation of non-compliance is essential, as it avoids the need for lengthy and costly investigations in order to rule out malabsorption problems, assay interference or genuine cases of thyroid hormone resistance/TSH-oma. There are some strategies that help to demonstrate non-compliance, such as monitoring of thyroid function tests after controlled administration of high dose of L-thyroxine. An example of a case of a non-compliant hypothyroid patient after thyroidectomy for Graves’ disease will be presented, as well as a strategy that helped to elucidate this problem. I will also present the data on the efficacy of once weekly L-thyroxine administration in comparison to daily L-thyroxine administration protocol. Such approach might be sometimes applied in order to deal with some cases of persistent non-compliance with L-thyroxine treatment. Nevertheless, there are cases where abnormal results of thyroid function tests persist, even though patient’s non-compliance has been ruled out. In such cases, assay interference, typically caused by the presence of heterophilic antibodies, must be suspected. These antibodies, usually directed against rodent antigens, are actually quite common, but most modern assays contain blocking antibodies that eliminate this problem in the majority of cases. If, however, the titre of such antibodies is extremely high, this may lead to abnormal test result. It should be mentioned that heterophilic antibody interference is not limited to results of thyroid function tests, but it may affect all other hormonal assays that involve the use of antibodies (e.g. sex steroid assays, etc) as well as other assays, for instance measurements of cardiac troponins. The likelihood of presence of high titre of heterophilic antibodies is increased in the setting of an augmented propensity for generation of autoantibodies, particularly in cases of coexistent rheumatoid disorders (such as rheumatoid arthritis). In cases of suspected assay interference, a clinician must decide in the first instance, whether obtained results fit the clinical picture. I shall endeavour to present a case of suspected heterophilic antibodies interference and I will also describe strategies to deal with such problems. The last, but not least I will also mention recently described cases of assay interference caused by the presence of macro-TSH particles, that may result in abnormal TSH measurements in a manner similar to macroprolactin.

Evaluation of the BRAHMS Kryptor® Thyroglobulin Minirecovery Test in patients with differentiated thyroid carcinoma

The present study was undertaken to evaluate a new sensitive thyroglobulin (Tg) mini-recovery test (Tg-mrec) for the detection of potential interferences in sera from patients with differentiated thyroid carcinoma (DTC) and low Tg levels. 167 DTC patients with serum Tg <2 μg/L were enrolled. Both TgAb and Tg-mrec measurements were performed on the automated Kryptor(®) platform. Serum pretreatment in proprietory blocking tubes was perfomed to screen for heterophile antibody interferences. The concordance rates between tests were evaluated. One case of over-recovery occurred in a patient with discordant Tg results in different immunoassays. The prevalences of a positive TgAb test and a reduced recovery rate were 15%-12%, respectively. Serum TgAb and Tg-mrec tests were both positive in 16 patients, both negative in 138 patients and discordant in 13 patients, respectively. The concordance rate between the Tg-mrec test and the TgAb assay was 92% (Cohens' kappa 0.894; 95% confidence interval: 0.82-0.93, p<0.001). In all, among seven clinically relevant interferences, three were found by TgAb, five by Tg-mrec and six by using both tests. The automated Kryptor(®) Tg-mrec test has a complimentary value to TgAb immunoassay testing in the detection of potential interferences in Tg measurements in patients with DTC and low Tg values (i.e., <2 μg/L). Tg-mrec may detect interferences from TgAb not measured directly by an individual TgAb assay or from other interfering substances, such as heterophile antibodies.

Falsely Elevated Testosterone Due to Heterophile Antibodies

Falsely elevated testosterone is a rare phenomenon that may result from heterophile antibodies. Similar to patients with heterophile antibodies for β-hCG, incorrect management may result in unnecessary testing or therapy. A previously healthy postmenopausal woman presented with a falsely elevated total testosterone level due to interference consistent with heterophile antibodies, with subsequent normal levels detected by liquid chromatography-mass spectrometry. Asymptomatic patients may present with an elevated hormone level due to heterophile antibody interference. Molecular studies for hormone levels can assist greatly when falsely elevated levels are suspected, but these tests are costly and time-intensive.

Identification and elimination of heterophilic antibody interference during antibody pair screening

High background interference during the antibody pair screening process is inevitable. In this study, we found that the high background was associated with heterophilic antibody interference introduced by the application of ascites-derived monoclonal antibodies when conducting large-scale antibody pair screening against different proteins. To eliminate antibody-associated heterophilic antibody interference, both blocking with mouse normal sera and antigen-mediated affinity chromatography were used, resulting in significant improvement in pairing performance and in antibody pair screening efficiency.