Laboratory Pitfalls Research Articles

Macro-FSH is a rare cause of inappropriately high FSH concentrations.

Assessment of hormone concentrations can be subjected to laboratory pitfalls. Macro-hormones are hormone-autoantibody complexes which are cleared slowly from circulation and cause a false elevation in hormones' concentrations. Macro-prolactin and macro-thyroid-stimulating hormone (TSH) are most frequently encountered while macro-follicle-stimulating hormone (FSH) has been rarely reported. We describe the case of a 30-year-old woman who had a gynaecological consultation due to failure in achieving pregnancy after 8 months of unprotected intercourse. She had regular menses, did not complain of climacteric symptoms and her medical history was unremarkable. Antral follicle count and anti-mullerian hormone concentrations were normal, and regular ovulation was documented. Unexpectedly, high early follicular phase FSH concentrations were confirmed on two occasions (57 and 51 IU/L), raising the suspicion of primary ovarian insufficiency. After excluding Turner's syndrome and autoimmune oophoritis, a laboratory artifact was hypothesized. Following polyethylene glycol precipitation, FSH levels dropped from 41.1 IU/L to 6.54 IU/L (recovery 16%) and the presence of macro-FSH was concluded. Laboratory interference can lead to misdiagnosis and unnecessary treatments. A laboratory artifact should be suspected when inconsistency exists between clinical presentation and laboratory results. Only five other cases of macro-FSH have been reported to date. Although macro-hormones generally have low biological activity and do not require treatment, the role of anti-FSH antibodies has been hypothesized in primary ovarian insufficiency and in vitro fertilization failure. Hormone quantification is a cornerstone in the diagnostic workup of endocrine disorders, but it can be subjected to laboratory interferences which can lead to unnecessary investigations and inappropriate treatments. A laboratory artifact should be suspected when a discrepancy is observed between clinical presentation and laboratory results, when extremely unusual analyte concentrations are observed and when inconsistent results are obtained by different analytical methods. Macro-hormones are hormone-autoantibody complexes which are cleared slowly from circulation and cause a false elevation in hormone concentrations. Macro-prolactin and macro-TSH are most frequently encountered, while macro-FSH has been rarely reported. Macro-hormones can be detected by polyethylene glycol precipitation, gel filtration chromatography, or by using protein G or protein A columns. Although macro-hormones generally have low biological activity and do not require treatment, the role of anti-FSH antibodies has been hypothesized in primary ovarian insufficiency and in vitro fertilization failure.

P-39 Fictitious hyperestrogenism in perimenopause: Difficulties in diagnosis by laboratory pitfalls

Abstract Introduction Laboratory errors related to interfering substances can cause significant morbidity related to unnecessary investigations and interventions and patient anxiety. Estradiol (E2) is quantified mainly with competitive immunoassays. The prevalence of analytical interference can be up to 6% and is due to the impairment of the interaction between test antibody and analyte, causing falsely elevated levels. Nevertheless, interference due to heterophile antibody is extremely rare. Clinical Case A 51-year-old woman, normal weight, nulliparous, irrelevant previous medical history, menarche at 12 years old and regular menstrual cycles, was referred to the Endocrinology Department due to elevated E2 level. She was followed 4 years before in another institution with no formal diagnosis complaining of severe vasomotor symptoms, vaginal dryness and menometrorrhagias. She was on combined oral contraceptives for 10 years as a method of contraception, which she stopped two years ago. The first two E2 random values were 2362 and 3121 pmol/L, respectively when she had 47 and 49 years old. During our evaluation in January 2021, at the age of 51 years old, in the 4th day of her menstrual cycle, the laboratory evaluation revealed: FSH 9.56 mUI/mL (2.5-10.2), LH 5.43 mUI/mL (1.9-12.5), Prolactin 18.12 ng/mL (2.8-29.2), E2 3530.51 pmol/L (71.6-529.2), 17-Hydroxiprogesterone 0.52 ng/mL (0.32-1.47), DHEA-SO4 75.9 ug/dL (35-430), 4-Androstenedione 1 ng/mL (0.3-3.3) and SHBG 137 nmol/L (18-144). Transvaginal pelvic ultrasonography performed 15 days after menstrual period revealed 2 subserous leiomyomas, a 12 mm endometrial line and a right ovary with a pre-ovulatory follicle. CT scan of adrenal glands was unremarkable and a sellar MRI revealed a 4mm area of hypocaptation in the left region of the adenohypophysis. During this period the patient was treated with escitalopram 10 mg od, with vasomotor symptoms improvement. Two years later, after a 6-month period of amenorrhea, she repeated the analytical evaluation that revealed raised gonadotropins levels (FSH 102.12 mUI/mL, LH 25.37 mUI/mL), low progesterone 0.91 mUI/mL (< 4.45) but maintained a largely elevated estradiol level (1128.01 pmol/L) with undetectable free Estriol (< 0.10 ng/mL). This pattern motivated laboratorial review of blood samples by mass spectrometry and not by chemiluminescence as before. The value was substantially lower (447 pmol/L) hypothesizing the presence of heterophilic antibodies. Conclusion Although false E2 elevation due to analytical interference is rare, it must be suspected when results are discordant with the clinical picture and there is no evidence of neoplastic disease. Here, we report a case of falsely elevated E2 that led to prolonged investigation and major impact on a female's quality of life during the challenging period of perimenopause.

Extremely high testosterone level in a woman without virilization signs. Is it only laboratory pitfalls?

Searchable abstracts of presentations at key conferences in endocrinology ISSN 1470-3947 (print) | ISSN 1479-6848 (online)

Improving Science by Overcoming Laboratory Pitfalls With Hormone Measurements.

Despite all the effort taken, there is often surprisingly little attention paid to the hormone analyses involved in research studies. Thinking carefully about the quality of the hormone measurements in these studies is, however, of major importance, as this attention to methods may prevent false conclusions and inappropriate follow-up studies. We discuss issues regarding hormone measurements that one should consider, ideally prior to starting, or otherwise, as they arise during a scientific study: quality of the technique, expertise, matrices, timing and storage conditions, freeze-thaw cycles, lot-to-lot and day-to-day variation, analyses per batch or sample-wise, singlicate or duplicate measurements, combining methods, and standardization. This article and the examples mentioned herein aim to clarify the need to pay attention to the hormone analyses, and to help in making decisions. In addition, these examples help editors and reviewers of scientific journals to pay attention to the methods section in the submitted manuscripts and ask the right critical questions when needed.

Diagnostic Pitfalls at Admission in Patients with Acute Superior Mesenteric Artery Occlusion

BackgroundAcute superior mesenteric artery (SMA) occlusion leads to acute intestinal ischemia and is associated with high mortality. Early diagnosis is often missed, and confounding factors leading to diagnostic delays need to be highlighted. ObjectivesTo identify potential diagnostic laboratory pitfalls at admission in patients with acute SMA occlusion. MethodsFifty-five patients with acute SMA occlusion were identified from the in-hospital register during a 4-year period, 2005–2009. ResultsThe median age was 76 years; 78% were women. The occlusion was embolic in 53% and thrombotic in 47% of patients. At admission, troponin I was above the clinical decision level (> 0.06 μg/L) for acute ischemic myocardial injury in 9/19 (47%) patients with embolic occlusion. Elevated pancreas amylase and normal plasma lactate were found in 12/45 and 13/27, respectively. A troponin I (TnI) above the clinical decision level was associated with a high frequency of referrals from the general surgeon to a specialist in internal medicine (p = 0.011) or a cardiologist (p = 0.024). The diagnosis was established after computed tomography angiography in 98% of the patients. The overall in-hospital mortality rate was 33%. Attempting intestinal revascularization (n = 43; p < 0.001), with a 95% frequency rate of completion control of the vascular procedure, was associated with a higher survival rate, whereas referral to the cardiologist was associated with a higher mortality rate (p = 0.018). ConclusionElevated TnI was common in acute SMA occlusion, and referral to the cardiologist was found to be associated with adverse outcome. Elevated pancreas amylase and normal plasma lactate values are also potential pitfalls at admission in patients with acute SMA occlusion.

Seroprevalence of IgG antibodies to SARS-coronavirus in asymptomatic or subclinical population groups.

We systematically reviewed the current understanding of human population immunity against SARS-CoV in different groups, settings and geography. Our meta-analysis, which included all identified studies except those on wild animal handlers, yielded an overall seroprevalence of 0.10% [95% confidence interval (CI) 0.02-0.18]. Health-care workers and others who had close contact with SARS patients had a slightly higher degree of seroconversion (0.23%, 95% CI 0.02-0.45) compared to healthy blood donors, others from the general community or non-SARS patients recruited from the health-care setting (0.16%, 95% CI 0-0.37). When analysed by the two broad classes of testing procedures, it is clear that serial confirmatory test protocols resulted in a much lower estimate (0.050%, 95% CI 0-0.15) than single test protocols (0.20%, 95% CI 0.06-0.34). Potential epidemiological and laboratory pitfalls are also discussed as they may give rise to false or inconsistent results in measuring the seroprevalence of IgG antibodies to SARS-CoV.

Rhodococcus equi Brain Infection in an Immunocompetent Patient: A Case Report and Review of the Literature

Rhodococcus spp. are opportunistic pleomorphic aerobic gram-positive bacteria that usually cause infection in the context of hosts with cellular immune deficiency. Initially described as a common zoonotic pathogen in foals, infection in humans has been more commonly reported since the emergence of the AIDS epidemics. Pulmonary cavitary lesions are, by far, the most common clinical manifestation, but virtually any organ may be affected, usually because hematogenous spread of this bacterium. Clinically significant infection in immunocompetent hosts is a rare event, especially when the site of involvement is extrapulmonary. Primary brain Rhodococcal abscesses are very unusual and, to the best of our knowledge, they have never been documented in immunocompetent individuals in the absence of direct inoculation of the organism. We report a peculiar case of Rhodococcus spp. causing a primary brain abscess in an immunocompetent man. The clinician should be aware of potential laboratory pitfalls in the identification of Rhodococcus spp., as its diphtherioid morphology may prompt discarding the organisms as a contaminant. The importance of identifying the causative organism is paramount to adequately select antimicrobial therapy, particularly in the clinical setting of central nervous system (CNS) infection, where antibiotics are not only selected according to their bactericidal properties but also taking in consideration the blood-brain barrier permeability of the drug. Therefore, the optimal therapeutic approach of Rhodococcus spp. brain infections relies on surgical drainage plus relatively long-term multi-drug antimicrobial treatment, preferably based on drug sensitivity analysis. This article reviews the clinical and pathologic features of Rhodococcus spp. infection as an emerging opportunistic pathogenic organism in immunocompromised and immunocompetent individuals.

False-positive polymerase chain reaction results for human papillomavirus in lichen planus: Potential laboratory pitfalls of this procedure

Background: The polymerase chain reaction (PCR) is a common tool in laboratory evaluation of tissue samples. It has been particularly useful in testing for nucleic acids of infectious organisms implicated in the pathogenesis of cutaneous diseases. Objective: Our purpose was to evaluate archival biopsy material (paraffin-embedded) of lichen planus for human papillomavirus (HPV) DNA with PCR and in situ hybridization (ISH). Methods: Tissue sections were obtained from 13 specimens of lichen planus and 10 control tissues. Standard methods for PCR and ISH were used to evaluate the presence of HPV DNA. Results: Initial PCR results demonstrated HPV DNA in 11 of 13 specimens and in none of the control tissue. However, testing for specific HPV types revealed nucleic acid contamination. ISH was negative in all specimens. Conclusion: Accurate evaluation of tissue with PCR is difficult because of the procedure's profound sensitivity. Positive results reported in the literature should be viewed with caution. Potential causes for false-positive and false-negative results should be considered.

Laboratory pitfalls that contribute to embrasure clasp failure

Use of an embrasure clasp demands attention to proper clinical preparation and to laboratory procedures as well. This article outlined pitfalls that occur in the laboratory, and appropriate solutions were offered.