Heterophile Research Articles

Exploration on the Detection Results of 2019 Novel Coronavirus Antibodies in the Real World

Background: In the early stage of COVID-19, the colloidal gold method was the first to be approved by the National Medical Products Administration for clinical use, but the interfering factors of false positive antibody test results are still unclear. Objective: To explore how to analyze and interpret the positive results of COVID-19 antibody test in clinical practice. Methods: A total of 8 678 patients simultaneously submitted for SARS-CoV-2 IgM/IgG antibody and nucleic acid in Peking University International Hospital from March to June 2020 for retrospective analysis were included in the inretrospective analysis. The epidemiological history, clinical manifestations and laboratory indicators(including SARS-CoV-2 IgM/IgG antibody and nucleic acid, rheumatoid factor, complement, immunoglobulin). The colloidal gold method was used to detect the SARS-CoV-2 IgM/IgG antibody, and the positive check of the antibody was performed by the magnetic particle chemiluminescence method. RT-PCR method was adopted to detect SARS-CoV-2 nucleic acid. The heterophilic antibody blocking tube(HBT)was used to process the specimens that were positive by the colloidal gold method, and then the test was performed again. Results: (1)Among the 8 678 patient specimens submitted for examination, 8 677 were negative for SARS-CoV-2 nucleic acid detection, of which 25 cases(0.288%)were positive for SARS-CoV-2 IgM antibody detected by colloidal gold method, SARS-CoV-2 -5 patients(0.058%)with IgG antibody positive, and 0 patients with SARS-CoV-2 IgM and SARS-CoV-2 IgG antibodies at the same time.(2)Thirty patients who tested positive for SARS-CoV-2 antibody by colloidal gold method had no epidemiological history, and at the same time, the results of SARS-CoV-2 nucleic acid test were negative. The possibility of SARS-CoV-2 infection or previous SARS-CoV-2 infection was ruled out. Among them, 10 patients with positive SARS-CoV-2 IgM antibody were still positive after more than 2 times of dynamic monitoring, and the remaining 20 patients were not clinically active monitored.(3)The false positive rate of SARS-CoV-2 IgM antibody detected by colloidal gold method was 0.288%, and the false positive rate of SARS-CoV-2 IgG antibody was 0.058%. After HBT was used to process the SARS-CoV-2 colloidal gold-positive specimens, among the 25 SARS-CoV-2 IgM antibody-positive specimens, except for 1 case which was still positive, the other results became negative;the results of 5 specimens with positive SARS-CoV-2 IgG antibody were still positive.(4)The rheumatoid factor, complement and immunoglobulin levels of 30 SARS-CoV-2 antibody-positive patients were within the normal reference range. Conclusion: The SARS-CoV-2 antibody test results may have false positives. The majority of SARS-CoV-2 IgM antibody positives are caused by heterophilic antibody interference. Those with SARS-CoV-2 IgG antibody positive may also have other potentially unknown interference factors. Heterophile antibodies interfere with colloidal gold methods more than chemiluminescence methods. Therefore, non-suspected patients or confirmed patients should not be tested for SARS-CoV-2 antibody to guide clinical practice, and the interference factors of the experiment must be considered. Copyright © 2021 by the Chinese General Practice.

A Young Child with Subacute Onset of Behavioral Changes.

1. Sarah Pradhan, MD* 2. Jessica Goldstein, MD† 3. Erin Frank, MD* 4. Allayne Stephans, MD* 1. *Department of Pediatric Hospital Medicine, 2. †Department of Pediatric Neurology, Rainbow Babies and Children’s Hospital, Cleveland, OH 1. Address Correspondence to Sarah Pradhan, MD; E-mail: sarah.pradhan6{at}gmail.com A 5-year-old healthy boy presents with a 3-week history of new onset aggression, obsessive-compulsive behavior, and nocturnal enuresis. One month ago he sustained a minor head trauma when he was sliding down the slide at a local playground. He was initially diagnosed with a mild concussion. His symptoms have now progressed and include frequent night waking, hand wringing, and behavioral outbursts. This prompts his pediatrician to refer him to the emergency department for evaluation. He undergoes computed tomography scan of the head, which is unremarkable. Laboratory evaluations include electrolytes, liver enzymes, complete blood cell count, rapid group A streptococcal antigen, thyroids studies, and heterophile antibody test, which are unremarkable. Urine and blood toxicology screens are negative. He is admitted to the inpatient unit. Vital signs are appropriate for age. He has difficulty answering questions and following commands. He is restless and disoriented. He intermittently gets up to run around the room and occasionally bangs his head on the wall. His cranial nerve, motor, reflexes, sensory, coordination, and gait examinations are normal. Brain magnetic resonance imaging is unremarkable. On video electroencephalogram he has bilateral fronto-temporal spikes that correspond with his abnormal movements and subclinical seizures arising from the right and left frontotemporal areas independently (arrows in Fig 1). Fosphenytoin is started, which controls his seizures, but he continues to have abnormal behavioral outbursts. Over the course of the week, he develops dysarthria, oral-facial …

Laboratory interference in the thyroid function test.

Thyroid hormones and thyroid-stimulating hormone (TSH) laboratory tests are commonly used worldwide, and their results have an important influence on decisions about treatment and further diagnostic processes. Any discrepancies between symptoms and laboratory results or between results of different tests should be closely investigated to avoid misdiagnosis and unnecessary treatment. Inconsistencies in hormone tests might be a result of physiological changes in hormonal balance, a disease, drug intake, or laboratory interference. Major factors that interfere with thyroid function tests are: heterophilic antibodies, macro TSH, biotin, thyroid hormones autoantibodies, anti-streptavidin, and anti-ruthenium antibodies. In this paper we discuss the influence of different factors on the procedures of hormonal immunoassays, as well as methods to minimise the risk of false results and misdiagnoses.

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Evaluation of sporadic bovine alkaline phosphatase interference in the Beckman Access unconjugated estriol (uE3) assay affecting maternal serum screening results

ObjectivesBovine alkaline phosphatase (BALP) mediated interference is a potential issue in the Beckman Access unconjugated estriol (uE3) assay. As the uE3 assay is a component of second trimester maternal serum screening characterizing this interference is essential for delivering accurate trisomy 18 and trisomy 21 risks. Design and methodsResidual serum samples (n = 517) were measured by two different lots of uE3 assay. Scavenger BALP (sBALP) was added to all samples to remove potential BALP dependent interference and assessed using both lots of uE3 reagent. ResultsBALP mediated interference was observed in similar frequency in both lots of reagent (~3%), although the patterns of positive and negative interference differed between the lots. Pretreatment with sBALP improved lot-to-lot comparison. The presence of BALP related interference was not related to the concentration of endogenous human alkaline phosphatase. The use of polyethylene glycol and sBALP treatment appeared to mitigate BALP mediated interference equally well, and resulted in concordance in measured uE3 concentrations between reagent lots. Additionally, heterophile antibody interference was observed in two samples affected with BALP interference, and the heterophile antibody interference was resolved by both PEG and heterophile antibody blocking reagent treatment, but not sBALP treatment. While the maternal screen numeric risk for affected samples changed, the risk classification changed from a negative to positive screen in two samples. ConclusionsInterference in the uE3 assay has the potential to affect maternal serum risk calculations in different reagent lots, and pretreatment of samples with scavenger BALP or PEG should be considered in cases of unexplained uE3 concentrations.

An overlooked situation in the interpretation of serum thyroglobulin level in a papillary thyroid cancer patient

Abstract In the present study we report a case of thyroglobulin (TGB) measurement interference in a total thyroidectomized and radio-ablated 61-year old woman with papillary thyroid cancer. We investigated possible interference in the measurement of TGB due to discordant TGB in relation to clinical condition during the follow-up period. Serum TGB was measured with the chemiluminescence method using Beckman Coulter Unicel DxI 800 instrument. To investigate possible interference in TGB measurement serial dilutions, polyethylene glycol precipitation (PEG), treatment with heterophile blocking tube (HBT), rheumatoid factor level determination and retesting of TGB with an alternative method were performed. Serial dilutions of the serum sample revealed linearity but a remarkable decrease in TGB in the patient’s serum samples post PEG and post HBT treatments. Also, TGB results under functional sensitivity level obtained with a different method suggested that TGB interference developed due to heterophile antibody presence in the serum sample. The patient had unnecessarily undergone expensive imaging techniques, and invasive procedures such as lymph node fine needle aspiration biopsy, before the analytical interference was suspected by the clinician. This report illustrates the importance of early communication and close collaboration between clinicians and laboratorians in order to avoid unnecessary clinical intervention.

Falsely markedly elevated 25-hydroxyvitamin D in patients with monoclonal gammopathies

Monoclonal immunoglobulins can cause interference in many laboratory analyses. During a 4month period we observed seven patients with monoclonal disease and falsely extremely elevated 25-hydroxyvitamin D (25(OH)D) results above 160ng/mL (>400nmol/L) measured using an immunoassay from Abbott Diagnostics. Based on these findings, we studied the occurrence of falsely elevated 25(OH)D in samples with paraproteins and investigated possible mechanisms of the observed interference. 25(OH)D was analyzed using the Architect i2000 platform from Abbott Diagnostics and a higher order method, liquid chromatography-mass spectrometry (LC-MS/MS), in serum samples from 50 patients with known monoclonal disease. Patients with falsely elevated 25(OH)D were included in further studies to elucidate the cause of interference. Spuriously elevated results were in addition analyzed on two alternative platforms (Siemens and Roche). Falsely elevated 25(OH)D levels were present in eight patients on the Abbott analyzer and one on the Siemens platform. Results from Roche were comparable with LC-MS/MS. Additional investigations excluded elevated concentrations of rheumatoid factor and heterophilic antibodies as the cause of interference in the Abbott assay. Laboratories should be aware of the risk of falsely elevated 25(OH)D in samples run on the Architect analyzer from patients with monoclonal disease. Highly elevated vitamin D results should be diluted and if the dilution is non-linear, rerun by a different method, preferably LC-MS/MS. In patients with spuriously elevated 25(OH)D without known monoclonal disease, the laboratory should consider requesting protein electrophoresis to exclude paraprotein interference.

Elevated Cardiac Troponin in the Setting of Periodic Symptomatology: A Case Report

Abstract Introduction/Objective Serial cardiac troponin measurement is a sensitive method to identify recent cardiac injury and is a necessary function of the clinical chemistry laboratory. We present a case of out-of-range cardiac troponin I (cTnI) elevation which remained spuriously elevated for at least 30 days. Methods Cardiac troponins were measured on a DxI (Beckman Coulter), Vitros 5600 (Ortho Clinical Diagnostics), iSTAT (Abbott), and Cobas e602 (Roche) according to the manufacturers’ instructions with appropriate controls. Heterophile antibody blocking (Scantibodies Laboratories) was performed according to the package insert. Results A 58-year-old male with a medical history significant for chronic non-ischemic cardiomyopathy with open valvular repair (ejection fraction 15-20%), chronic atrial fibrillation, and Hodgkin’s lymphoma in remission presented to a rural clinic with chest pain and a cTnI of 0.8 ng/mL (reference <0.04 ng/mL). He was transferred to our tertiary center for open revision. The patient’s cTnI (measured on a Beckman Coulter DxI) was above linearity threshold (>71.00 ng/mL) starting day 1 after surgery. The patient self-discharged against medical advice on day 16. On day 21 he experienced new chest pain and dyspnea and was readmitted with a cardiac troponin T (cTnT) of >35 ng/mL (iSTAT; reference <0.08 ng/mL) and cTnI again above threshold. This value remained elevated for the next five days despite abated symptoms. To address the discordant laboratory and physical findings, dilutional cTnI measurements on serum from day 30 demonstrated linearity. Heterophile antibody testing was negative. cTnI on the DxI machine remained >71.00 ng/mL but was separately measured as 3.4 ng/mL (Vitros 5600, reference <0.08 ng/mL). cTnT was found to be 1.05 ng/mL (Cobas e602; reference 0.00 ng/mL). Comparative examination of all methodologies was unrevealing. Conclusion This case demonstrates a spurious elevation of cardiac troponins which remarkably demonstrated linearity on serial dilution. Multiple testing methodologies were necessary to prove fallacy. The cause of the result remains unclear. The clinical pathologist should be aware of possible false positives and investigate unlikely continuous cardiac troponin elevations.

S2737 When the Student Becomes the Patient: A Case of Epstein-Barr Virus-Induced Acute Liver Injury

INTRODUCTION: Viral hepatitis is a diagnosis that can lead to acute liver injury and commonly occurs following an acute Hepatitis A or B infection. Epstein-Barr virus (EBV) is a rare cause of acute viral hepatitis. We present a case of acute liver injury secondary to acute EBV infection. CASE DESCRIPTION/METHODS: A 25-year-old female medical student with no significant past medical history presented with five days of fever, sore throat, nausea, and left upper quadrant abdominal pain while on her Pediatric rotation. She was afebrile, tachycardic to 106 beats/min, and demonstrated tender cervical lymphadenopathy, exudative tonsillar enlargement, and splenomegaly on physical exam. Heterophile antibody was negative three days prior to presentation. Initial studies revealed a leukocytosis of 12,600/mm3 and elevated liver enzymes (AST 421 U/L, ALT 788 U/L, alkaline phosphatase 404 U/L, total bilirubin 1.4 mg/dL, and INR 1.3). CT abdomen revealed hepatosplenomegaly. Hepatic vasculature was patent on doppler ultrasound with an unremarkable gallbladder and biliary tree. Subsequent studies were pertinent for an undetectable serum acetaminophen level, negative Hepatitis A-C serologies, and negative cytomegalovirus PCR. EBV serologies were positive for acute infection including an elevated viral capsid antibody, IgM, IgG, and negative nuclear antigen antibody. The patient was discharged with supportive treatment and followed ten days later in clinic with liver enzymes improved to near normal levels. DISCUSSION: Infectious mononucleosis from EBV infection is associated with development of acute hepatitis in 80% of cases. Diagnosis may be delayed due to high rate of false negative heterophile antibodies early in the disease course. Most cases are subclinical with hepatomegaly occurring in only 14% of cases. The pattern of liver injury is typically hepatocellular. Serum aminotransferases exhibit a transitory rise to three-fold the upper limit of normal, peak in the second or third week of illness, and normalize within thirty days. Treatment is typically supportive, though steroids and antivirals have been used in severe cases. Viral hepatitis is a common cause of acute liver injury, second to drug-induced liver injury, and accounts for 10% of acute liver failure. This case highlights the importance of including uncommon causes of viral hepatitis in the differential for acute liver injury, particularly in patients without traditional risk factors for hepatitis.

Hypeprolactinemia: still an insidious diagnosis

Hyperprolactinemia can have different causes: physiological, pharmacological, and pathological. When investigating the etiology of hyperprolactinemia, clinicians need to be aware of several conditions leading to misdiagnosis. The most popular pitfalls are: acute physical and psychological stress, macroprolactin, hook effect, even though antibodies interferences and biotine use have to be considered. A 52-year-old woman was referred to Endocrinology clinic for oligomenorrhoea and headache. She worked as a butcher. Hormonal evaluation showed very high PRL (305 ng/ml, reference interval: <24 ng/ml) measured with the ECLIA immunoassay analyzer Elecsys 170. The patient’s pituitary MRI was normal and macroprolactin was normal. Hormonal workup showed LH: 71.5 mU/ml (2–10.9 mU/ml), FSH: 111.4 mU/ml (3.9–8.8 mU/ml), Estradiol: 110.7 pg/mL (27–122 pg/ml). Since an interference was suspected, the sample was sent to another laboratory using a different assay. After antibody blocking tubes treatment (Heterophilic Blocking Tube, Scantibodies) PRL was 28.8 ng/ml (reference interval < 29.2 ng/ml). Analytical interference should be suspected when assay results are not consistent with the clinical picture. Endogenous antibodies (EA) include heterophile, human anti-animal, autoimmune and other nonspecific antibodies, and rheumatoid factors, that have structural similarities and can cross-react with the antibodies employed by the immunoassay, causing hyperprolactinemia misdiagnosis. The patient’s job (butcher), led us to suspect the presence of anti-animal antibodies. Clinicians should also carefully investigate the use of supplements. Biotin can falsely increase hormone concentration in competitive assays. Many clinicians are still not informed about these pitfalls that are not mentioned in some recent reviews on PRL measurement.

Immunoglobulin Preparations Can Mislead Clinical Decision-Making in Follow-Up of Differentiated Thyroid Cancer

Objective: Intravenous and subcutaneous immunoglobulins are commonly used for immune substitution or as immune modulators in a variety of inflammatory and autoimmune disorders. Exogenous thyroid-specific thyroglobulin (Tg) antibodies present in the donor plasma may interfere with the interpretation of measurements of Tg autoantibodies (Tg-Abs) in the recipient’s plasma and potentially trigger an immune response in the recipient’s immune cells. Levels of antibodies causing bioassay interferences or those leading to clinically relevant changes in patient outcomes are not known. Tg is used as a biomarker in the long-term surveillance of patients with differentiated thyroid cancer (DTC) following total thyroidectomy and radioactive iodine ablation. However, the presence of Tg-Abs in the circulation interferes with Tg measurements. Assessment of levels of Tg-Abs is thus recommended as a part of standard follow-up of DTC together with Tg testing.Methods: To understand the potential mechanisms and pathophysiologic significance of possible interferences associated with administration immunoglobulin preparations and Tg measurement, we overview the current knowledge on interactions between Tg autoimmunity and immunoglobulin preparations and illustrate diagnostic challenges and perspectives for follow-up of patients with DTC treated with exogenous immunoglobulins.Results: In patients with DTC treated with immunoglobulin preparations, monitoring of thyroid cancer using Tg and Tg-Abs is challenging due to possible analytical interferences through passive transfer of exogenous antibodies from immunoglobulin preparations.Conclusion: Analytical interferences must be suspected when a discrepancy exists between clinical examination and diagnostic tests. Collaboration between endocrinologists, biologists, and pharmacologists is fundamental to avoid misdiagnosis and unnecessary medical or radiologic procedures.Abbreviations: CT = computed tomography; DTC = differentiated thyroid cancer; FNAB = fine-needle aspiration biopsy; HAb = heterophile antibody; IMA = immunometric assay; IVIg = intravenous immunoglobulin; RAI = radioactive iodine; RIA = radioimmunoassay; SCIg = subcutaneous immunoglobulin; Tg = thyroglobulin; Tg-Ab = thyroglobulin autoantibody; Tg-MS = thyroglobulin mass spectrometry; TPO-Ab = thyroid peroxidase autoantibody; TSHR-Ab = thyrotropin receptor autoantibody

Clinical laboratory investigation of a patient with an extremely high D-dimer level: A case report

BACKGROUNDD-dimer, a soluble degradation product of cross-linked fibrin, is commonly used as an important marker for the diagnosis of disseminated intravascular coagulation and differential diagnosis of thrombosis. Herein, we present a geriatric case with an unusually elevated D-dimer level.CASE SUMMARYAn 82-year-old woman, admitted to the ward with a diagnosis of chronic heart failure, was noted to have a remarkably elevated D-dimer level, beyond the qualified range (> 100 mg/L), utilizing the Innovating D-dimer for Sysmex CS-5100 System™. However, no evidence, including clinical symptoms, radiographic evidence of thromboembolic disease, and parallel fibrinogen degradation product values, suggested that this patient was at high risk of thrombopenia. To confirm the discrepancy, a series of approaches including sample dilution, re-analysis via alternative methods, and sample treatment with blockage of specific heterophilic antibodies were performed. A remarkable disappearance of the elevated D-dimer values was observed in the samples after they were subjected to these approaches (4.49, 9.42, 9.06, and 12.58 mg/L, respectively). This confirmed the presence of heterophilic antibodies in this case. In addition, a reduction in cardiac output due to the presence of cardiac failure could also be responsible for the existence of a hypercoagulable state in this case.CONCLUSIONIn conclusion, the presence of heterophilic antibodies should be considered when an elevated D-dimer value is not in conformity with the clinical evidence, and a viral infection should be considered when interference by a heterophilic antibody exists.

Pre-existing canine anti-IgG antibodies: implications for immunotherapy, immunogenicity testing and immunoassay analysis

One of the most enigmatic features of humoral immunity is the prevalent presence of circulating autoantibodies against IgG. These autoantibodies consist of several subsets, including rheumatoid factors, anti-Fab/anti-F(ab′)2-autoantibodies, and anti-idiotypic antibodies. Anti-IgG autoantibodies can impair the safety and efficacy of therapeutic antibodies and interfere with immunogenicity tests in clinical trials. They can also cross-react with allospecific IgG, presenting as heterophilic antibodies that interfere with diagnostic immunoassays. Owing to these factors, recent years have seen a resurgent interest in anti-IgG autoantibodies, but their underlying clinical significance, as well as biological roles and origins, remain opaque. Increased knowledge about canine anti-IgG autoantibodies could facilitate the development of canine immunotherapies and help in understanding and counteracting immunoassay interference. This study investigated the clinical significance and interconnection of heterophilic antibodies, anti-Fab, and anti-F(ab′)2-autoantibodies in dogs. We performed a 2-year prospective follow-up of dogs with heterophilic antibodies and analyzed serum for anti-Fab and anti-F(ab′)2-autoantibodies. Canine heterophilic antibodies can persist for at least 2 years in serum. A widespread occurrence of anti-Fab and anti-F(ab′)2-autoantibodies was found, with reactivity to cryptic epitopes in the IgG hinge region and sporadic cross-reactivity with mouse IgG. Canine anti-Fab and anti-F(ab′)2-autoantibodies are thus potential sources of clinical immunogenicity and immunoassay interference.

Glycosylation may influence sex hormone-binding globulin measurements

A discordance between sex hormone-binding globulin (SHBG) measurements by 2-site ELISAs was investigated using pairings of various “in house” SHBG antibodies together with a concordant control. The 2-site monoclonal ELISAs used the same base coat (11F11) and discordance was observed with one top coat monoclonal antibody (7H9) and also when a polyclonal SHBG antibody was paired with the basecoat antibody (11F11). Sialidase treatment of the discordant sample and purified SHBG revealed increased levels using 7H9 whereas there was no change in SHBG in the concordant sample. Conversely, following sialidase treatment, the discordant sample showed no change in SHBG measured using the other monoclonal antibody pairings whereas the SHBG levels in the concordant sample declined following sialidase using the same monoclonal antibody pairings. This implicated glycosylation as a factor in antibody recognition and synthetic peptides spanning the two N-linked and one O-linked glycosylation regions showed that SHBG recognition by monoclonal antibody 7H9 could be disrupted by a peptide spanning the O-linked glycosylation site. Hence rather than immunoassay discordance being attributed to heterophile antibodies or other circulating antibodies here it can be likely attributed to glycosylation affecting antibody recognition and hence the measurement of SHBG.

MON-481 Remarkable Euthyroid Hyperthyroxinemia Mistaken for Thyrotoxicosis

CLINICAL CASEA 46 year old caucasian female with past medical history of menorrhagia was referred from primary care for evaluation of thyrotoxicosis. Thyroid function was assessed in the context of menometrorrhagia. She did not have any history of thyroid disorder or abnormal thyroid function tests. Per outside records, recent labs demonstrated TSH 0.88 uIU/mL (0.36-3.74), Free T4 > 8.00 ng/dL (0.76-1.46), Free T3 2.9 pg/mL (2.18-3.98). All other labs were within normal limits. Thyroid ultrasound revealed normal parenchyma and volume. She did not take any medications or supplements including biotin. She denied heat intolerance, anxiety, palpitations, dyspnea, tremors, hyperdefecation, or change in hair, skin, or mood. No epiphora, diplopia, or eye irritation was reported. Her father had been diagnosed with hyperthyroidism, mother with hypothyroidism.Repeat labs at our visit revealed normal TSH of 1.05 uIU/mL (0.358- 3.74), normal Free T3 2.58 pg/mL (2.18- 3.98), normal Total T3 136 ng/dL (80-200), elevated Free T4 >8.00 ng/dL (0.76-1.46) and elevated Total T4 11.6 ug/dL (4.5-10.5). These lab values were not consistent with patient’s euthyroid clinical status, prompting assessment of Free T4 by dialysis, normal at 1.5 ng/dL (0.9-2.2) and T3 uptake, high at 40% (24-39%). This picture was consistent with Familial Dysalbuminemic Hyperthyroxenemia (FDH). The decision was made not to treat the patient with anti-thyroid medications and to perform a confirmatory genetic testing to test for mutations in the ALB (albumin) gene.DISCUSSIONThe free T4 assay used by our institution is performed on the Siemens Dimension Vista platform using a two-step chemiluminescent immunoassay. While in theory two-step assays should not yield abnormal results in FDH, several two-step assays are known to yield falsely high results in patients with FDH (1, 2, 3).Other potential etiologies for discordant Free T4 levels include thyroid hormone autoantibodies, heterophile antibodies, biotin use, and anti-streptavidin antibodies (3).CONCLUSIONRecognition of laboratory error in the workup of thyroid disease is essential. Clinicians must ensure thyroid function labs are consistent with each other and with the patient’s presentation. In such cases misdiagnosis of hyperthyroidism or thyroid hormone resistance may lead to unnecessary testing and inappropriate treatment (3).

MON-462 Rebel Nodule or Reluctant Gland? the Challenge Presented by Heterophile Antibodies

Introduction:Interpretation of thyroid function tests (TFTs) in background of non-specific symptoms concerning for hypothyroidism, has become challenging with assay interference. Heterophile antibody interferences are both test and laboratory platform dependent. We present a case of a toxic nodule erroneously diagnosed as isolated central hypothyroidism due to heterophile antibody interference.Case:A 38 year old lady was referred to Endocrinology for possible central hypothyroidism with symptoms of progressive fatigue, 40lb unintentional weight loss, poor appetite, nausea, intermittent diarrhea/constipation, occasional cold intolerance and intermittent lightheadedness. TFTs suggested low TSH 0.175 mcIU/ml (N 0.35-3.8 mcIU/ml) and low FT4 0.74 ng/dl (N 0.8-1.8 ng/dl). Pituitary hormone evaluation revealed normal ACTH, AM cortisol, IGF-1, prolactin and FSH. A pituitary MRI was normal.Two months later, she had worsening fatigue, anxiety with palpitations and tremors, and 14lb weight gain. TFTs again confirmed low TSH (0.575 mcIU/ml) and lower FT4 (0.70 ng/dl). Empirical weight-based levothyroxine (LT4) supplementation was initiated for central hypothyroidism. Six weeks later, TFTs showed hyperthyroidism (TSH <0.005 mcIU/ml, FT4 1.57 ng/dl). The patient endorsed worsening lightheadedness and palpitations on LT4, which resolved on LT4 discontinuation. One month later, TFTs were abnormal again (TSH 0.047 mcIU/ml, FT4 0.74 ng/dl).Due to persistent discordance in symptoms and biochemical tests, TFTs were evaluated for heterophile antibody interference. While HAMA-treated TSH remained low (0.04 mIU/l), FT4 by equilibrium dialysis (FT4 ED) was normal (1.1 ng/dl, regular assay 0.74 ng/dl), suggesting subclinical hyperthyroidism. A thyroid uptake scan confirmed an autonomous toxic right thyroid nodule with suppressed remaining gland. Patient ultimately underwent 15 mCi RAI ablation of the nodule. TFTs done 1 month post-RAI suggested normal TSH (1.2 mcIU/ml) and normal FT4 ED (1 ng/dl). Previous symptoms have since resolved.Conclusion:Current methods of TFTs are generally reliable in diagnosing and monitoring thyroid disease. Rarely, medications, supplements, and endogenous antibodies can bind to TSH, T4 or lab reagent, resulting in inaccurate values. This has become increasingly common with use of high dose biotin. TSH with HAMA/heterophile antibodies and FT4 ED are more accurate forms of diagnostic testing.When approaching patients with symptoms not consistent with typical hypo- or hyperthyroidism and are not responding as expected to therapy, it is important to consider more accurate testing to rule out assay errors.

SUN-493 A “Curve Ball” in the Management of an Infertile Hyperthyroid Patient

Introduction: We present the case of a young female patient referred for hyperthyroidism with persistently detectable TSH despite elevated free thyroxine (FT4) levels which, interestingly, failed to normalize following anti-thyroidal treatment. Further testing elucidated the underlying causation for detectable TSH in presence of hyperthyroxinemia to be due to interfering substances in the thyroid function assays. Case: A 35-year-old female is evaluated for hyperthyroidism discovered in preparation for embryo transfer. History includes transient hyperthyroidism two years ago while undergoing in vitro fertilization. Prior workup included negative antibodies and nuclear uptake scan revealing a right-sided autonomously functioning thyroid nodule with 24-hour uptake of 40.7%. She was briefly treated with PTU in first trimester and lost to follow-up. While once again undergoing fertility treatment with clomiphene, the patient developed tremor and heat intolerance. She was found to have TSH 0.3 (0.45-4.12 mU/L) and analog FT4 of 6.4 (0.8-1.8 ng/dL). She denied use of OTC supplements. PTU 50 mg twice daily was initiated with minimal improvement with TSH 0.7 mU/L, FT4 3.7 ng/dL. On evaluation, slight tremor and mild thyroid enlargement were noted. Our differential expanded to include TSH-secreting pituitary adenoma, thyroid hormone resistance (no family history), and assay interference. Both MRI pituitary and an alpha-subunit level (0.36) were normal. Symptoms improved gradually with repeat TSH 1.79 mU/L, FT4 2.4 ng/DL by standard analog methods. We suspected ‘assay interference’ for which FT4 via direct equilibrium dialysis was obtained and was indeed normal at 0.76 ng/dL. Further lab testing verified interference in both in the standard TSH and FT4 assays presumed secondary to heterophile antibodies. Discussion: Interpretation of thyroid studies discordant with the clinical picture or incongruent with each other requires understanding of thyroid physiology and the intricacies of commonly utilized assays. The differential of elevated thyroxine levels with detectable/normal TSH rests between thyroid hormone resistance syndromes, TSH secreting pituitary tumors and interfering substances in the assay. Most commercially available TSH assays are based on a ‘sandwich’ assay which is notoriously interfered with by heterophile antibodies and excess biotin. Determination of FT4 is also challenging as the assay must detect very low concentrations of free hormone relative to excess of protein-bound analyte. When in question, it is important consider utilization of laboratory expertise and retesting by alternative assays.

Recent Advances in Diagnostic Approaches for Epstein-Barr Virus.

Epstein–Barr virus (EBV) is the causative agent of many diseases including infectious mononucleosis (IM), and it is associated with different subtypes of lymphoma, sarcoma and carcinoma such as Hodgkin’s lymphoma, non-Hodgkin’s lymphoma, nasopharyngeal carcinoma, and gastric carcinoma. With the advent of improved laboratory tests for EBV, a timelier and accurate diagnosis could be made to aid better prognosis and effective treatment. For histopathological lesions, the in situ hybridization (ISH) of EBV-encoded RNA (EBER) in biopsy tissues remains the gold standard for detecting EBV. Methods such as the heterophile antibody test, immunofluorescence assays, enzyme immunoassays, Western blot, and polymerase chain reaction (PCR) are also employed in the detection of EBV in different types of samples. The determination of EBV viral load using PCR, however, is gaining more prominence in the diagnosis of EBV-associated diseases. Given the challenge of false positive/negative results that are sometimes experienced during the detection of EBV, variability in results from different laboratories, and the impact of factors such as sample type and the immunological status of patients from whom samples are collected, the need to critically examine these present methods is invaluable. This review thus presents current advances in the detection of EBV, detailing the advantages and disadvantages of the various techniques. In addition, fundamental virological concepts are highlighted to enhance the greater understanding, the proper application, and the interpretation of EBV tests.

FIFTH GENERATION TROPONIN ASSAY IS SUBJECT TO ANTIBODY INTERFERENCE: FIRST REPORTED CASE

Both the 5th generation, or high-sensitivity, Troponin T (hs-TnT) and 4th generation Troponin T (4G-TnT) assays are electrochemiluminescent immunoassays. The hs-TnT assay utilizes a chimeric mouse-human antibody whereas the 4G-TnT assay is solely murine. Thus, heterophilic antibody interference to

Non-coronarogenic causes of increased cardiac troponins in clinical practice

Cardiospecific isoforms of troponins are the most sensitive and specific biomarkers for the diagnosis of myocardial infarction. However, though elevated troponin levels indicate myocardial damage, they do not determine the cause and mechanism of the damage. With the new highly sensitive methods, very minor damages of the heart muscle can be detected. Myocardial damage can occur in many non-coronarogenic diseases. In this review, we discuss the mechanisms of elevation, the diagnostic value of cardiac troponins in the renal failure, tachyarrhythmias, endocarditis, myocarditis, pericarditis, sepsis, neurogenic pathologies (stroke), pulmonary embolism. In addition, we pay attention to the main reasons for a false-positive increase of the concentration of cardiac troponins: heterophilic antibodies, rheumatoid factor, alkaline phosphatase, cross-reactions with skeletal muscle troponins.