Diagnostic Dilemma: A Case of Undetectable TSH

Introduction: Under steady-state conditions, measurement of TSH is accepted as the best assessment of thyroid function. The widely used TSH chemiluminometric assays have very low limits of detection and can help distinguish between the various causes of subnormal TSH. However, when evaluating a patient with abnormal thyroid tests but without thyroid symptoms, an appraisal of the test should be considered. Clinical Case: A 63-year-old South Asian man was referred to endocrinology for evaluation of a non-detectable TSH (<0.01 µIU/mL) that was reproduced on repeat testing, both using Siemens ADVIA Centaur TSH3-UL immunoassay. The patient was clinically euthyroid and denied taking biotin supplements. Testing of thyroid hormone showed normal values for free T3, total T3, free T4, and total T4. Additional labs included normal studies for free thyroxine by equilibrium dialysis, thyroid stimulating immunoglobulin, and heterophilic antibodies. Thyroid uptake and scan showed uniform uptake of 5.1% and 15.1% at 2-hours and 24-hours, respectively, with no dominant nodules. Hypothalamic-pituitary hormonal testing and MRI pituitary were both normal as well. When TSH testing was repeated on a separate platform, Roche’s eCLIA immunoassay, detectable values were obtained (TSH 6.48 µIU/mL). Conclusions: Testing of serum TSH by commercially available immunoassays is based on the sandwich method in which one antibody binds to the β-subunit of TSH and the other to the α-β interface. Most assays use monoclonal antibody pairs to achieve high selectivity. Immunoassay tests are prone to interferences, particularly by way of altering the measurable concentration of the analyte or by altering antibody binding (1). In this case, the presence of detectable TSH depended on the platform by which it was measured. This finding suggests a TSH-β variant with impaired immunoreactivity but functionally normal bioactivity. Such a mutation has been previously reported to occur five times more frequently among South Asian individuals than the general population (2). Genetic testing was offered to the patient to confirm this suspicion but was declined. It is incumbent on the clinician to reconcile a test result that is discordant with the clinical presentation. Having a fundamental understanding of the principles of the testing platform can assist in identifying potential sources of error. Failing to recognize a possible interference can lead to unnecessary healthcare expenditures, misdiagnosis and inappropriate management, potentially at a cost to the patient’s wellbeing. When faced with an undetectable TSH with otherwise normal thyroid hormones and unremarkable clinical picture, it is best to repeat the TSH test using a different available platform. References: (1)Favresse J et al. Endocr Rev. 2018;39(5):830-850(2)Pappa T et al. Thyroid. 2015 Aug;25(8):869-76

Factors determining the metal ion binding ability and selectivity of hydroxamate based compounds

: There has been a long tradition for a broad spectrum of applications of both natural and synthetic hydroxamic acids and derivatives. Even nowadays, a huge number of newly designed representatives (from different monohydroxamate-based compounds to siderophore conjugates) are intended to develop potential drug candidates with desired activities. Since these compounds are effective metal-chelating agents their biological roles and actions as well as their various applications e.g. in the medicinal practice are all in direct correlation with their metal complexation. Consequently, the knowledge of the stoichiometry and binding modes of metal complexes with hydroxamic acid based ligands, their thermodynamic parameters, speciation profiles in solution is crucial for scientists working at any of the above-mentioned fields. This review, in addition to presenting a few factors, which might affect the metal binding capabilities of these organic ligands, displays and summarizes the different parameters typically used to give the stoichiometry/composition and stability of the species formed in a solution equilibrium system in measurable concentration. Discussion of the possibilities for quantitative comparison of metal binding effectivity and selectivity of various hydroxamic acids with each other by using solution equilibrium data is also in the focus of this publication.

Expanding the range of determination of elements on the Grand-AAS atomic absorption spectrometer using several of their absorption lines

One limitation of atomic absorption spectrometry is the narrow range of measurable concentration (1–2 orders of magnitude). In simultaneous multi-element analysis, this may require multiple dilutions of the sample to determine several elements with different concentrations in the sample. Possible ways to expand the range are to linearize the calibration curve by correcting the integral of the absorption signal or to use absorption values on the line wing as an analytical signal and plot several graphs along one line at different distances from its center. Both methods have their drawbacks. We propose another method for expanding the measurable concentration range by using less sensitive absorption lines of elements. A number of elements are identified that have a sufficient number of lines with different sensitivities. The proposed method is compared with the method of linearization of the calibration graph and the calculation of the absorption signal on the line wing. Using as an example Co and Ni, which have sufficiently rich absorption spectra, we have shown the possibility of expanding the measurable concentration range by using several absorption lines: for cobalt, the range is expanded to six orders of magnitude, and for nickel, to five orders of magnitude. Calibration curves are plotted in the concentration ranges 0.24–250.000 μg/L for cobalt and 1.9–250.000 μg/L for nickel. The calibration error is lower than that of the linearization method: 5% against 25 % for cobalt and 4% against 24% for nickel. Thus, the proposed method can be used for simultaneous multielement determination in a wide range of concentrations without diluting the sample.

Dexmedetomidine and ketamine simultaneous administration in tigers (Panthera tigris): pharmacokinetics and clinical effects

BackgroundThe study determines the pharmacokinetic profiles of dexmedetomidine (DEX), ketamine (KET) and its active metabolite, norketamine (NORKET), after simultaneous administration. Moreover, the study evaluates the sedative effects of this protocol, its influence on the main physiological variables and the occurrence of adverse effects.MethodsEighteen captive tigers were initially administered with a mixture of DEX (10 µg/kg) and KET (2 mg/kg) by remote intramuscular injection. In case of individual and specific needs, the protocol was modified and tigers could receive general anaesthesia, propofol or additional doses of DEX and KET.ResultsBased on the immobilisation protocol, nine animals were assigned to the standard protocol group and the other nine to the non-standard protocol group. Higher area under the first moment curve (AUMC0-last) and longer mean residence time (MRT0-last) (P<0.05) were observed in the non-standard protocol group for DEX, KET and NORKET, and higher area under the concentration-time curve from administration to the last measurable concentration (AUC0-last) only for KET. The KET metabolisation rate was similar (P=0.296) between groups. No differences between groups were detected in terms of stages of sedation and recoveries. All physiological variables remained within normality ranges during the whole observation period. During the hospitalisation period, no severe adverse reactions and signs of resedation were observed.ConclusionThe simultaneous administration of 10 µg/kg of DEX and 2 mg/kg of KET can be considered an effective protocol for chemical immobilisation of captive tigers, along with dosage adjusments or when other drugs are needed.

Trichoderma and sulphoethyl glucan reduce maize root rot infestation and fusaric acid content

Roots of maize seedlings (cv. Pavla) infested by Fusarium verticillioides (10⁵/ml) were cultivated on Murashige-Skoog medium (MSM, Sigma, USA) containing CaCl₂,IAA and kinetin. Simultaneously, a strain of the antagonistic fungus Trichoderma sp. and a sulphoethyl glucan (SEG) isolated from the cell walls of Saccharomyces cerevisiae, were added. Two evaluations (on 7 and 14 days) were done. Productivity parameters of leaves and roots (fwt, dwt, and length), disease severity index (DSI) and fusaric acid (FA) concentration were evaluated. Both Trichoderma sp. and SEG increased productivity parameters of plants in infested variants and maintained it on the level of control plants during 14&nbsp;days of experiment. Trichoderma reduced the DSI, while SEG increased it. DSI correlated with FA concentration. After seven days of cultivation concentration of FA was lower in all infected variants cultivated concomitantly with agents, compared with the one without them. After 14 days of cultivation both agents reduced the concentration of FA up to 50% to the non-measurable concentration in variant with Trichoderma. In variant with positive control, where FA was added to SEG, its concentration decreased up to 30%.

PIXE ANALYSIS OF BLOOD SAMPLES OF ORTHODONTIC PATIENTS TO DETECT Ni POISONING

The present study was carried out with the aim to determine if, orthodontic patients accumulate measurable concentration of Ni in blood or not, since the recent evidences shows the allergenic actions of Ni in various forms and orthodontic appliances have been reported to produce Ni allergy. In our experiment, the blood samples were taken before the insertion of appliance and at an interval of 6 months over a total time period of 18 months (four sets) from the Oral Health Department of Post Graduate Institute of Medical Education & Research (PGIMER), Chandigarh, India. In all the four sets of blood samples the common trace elements were detected viz. S , K , Ca , Cr , Fe , Cu , Zn and Br . Our result shows the complete absence of Ni in the blood.

Enzyme immunoassay of thyroxin-binding globulin in dried blood samples on filter paper.

A double-antibody enzyme immunoassay was developed for determination of thyroxin-binding globulin in dried blood samples on filter paper. The measurable concentration range of thyroxin-binding globulin in two 3-mm blood discs was 3.3 to 52 mg/L equivalent of serum (i.e., equivalent to the concentrations in known serum standards). Thyroxin-binding globulin in dried blood samples on filter paper was stable for at least four weeks when kept dry at -20 degrees C, 4 degrees C, or room temperature. The mean coefficients of variation were 6.6% (within assay) and 5.9% (between assays). The concentrations of thyroxin-binding globulin in dried blood samples determined by this method correlated well with those in serum determined by radioimmunoassay (r = 0.95) and by enzyme immunoassay (r = 0.96). This method is applicable for detecting cases of thyroxin-binding globulin deficiency and avoids the false-positive results for neonatal hypothyroidism obtained by measuring thyroxin.

Gas-Chromatographic Measurement of Codeine and Norcodeine in Human Plasma

A gas-chromatographic method is described for determination of codeine and norcodeine in human plasma. The method is specific, sensitive, and precise. It was developed for use in bioavailability studies of therapeutic doses of codeine sulfate. After ingestion of a 60-mg codeine sulfate tablet, mean peak codeine concentration in plasma was 107 µg/liter at 1.0 hour. No measurable concentration of norcodeine was found in the plasma by this method.

Radioimmunoassay of Plasma Testosterone, with Use of Polyethylene Glycol to Separate Antibody-Bound and Free Hormone

We have developed a reliable radioimmunoassay for testosterone in plasma, polyethylene glycol ("Carbowax 6000") being used to separate antibody-bound and free hormone. Testosterone is separated from interfering steroids, notably dihydrotestosterone, by liquid—liquid partition chromatography on infusorial earth (Celite). The assay is sensitive (9 pg for standards), precise, and accurate. The lowest measurable concentration of testosterone is 350 ng/liter for plasma from men and 70 ng/liter for plasma from women. Intraand inter-assay coefficients of variation were 6.9% and 9.7%, respectively, for plasma from men, and 9.6% and 11.8%, respectively, for plasma from women. Our method for separating antibody-bound and free hormone is practical and convenient and may be generally applicable to all radioimmunoassays of steroid hormones in plasma.

KINETIC STUDIES OF PROTON TRANSFER FROM PHENOLS TO TRINITROBENZYL ANION

The rates of reaction of various phenols with the 2,4,6-trinitrobenzyl anion in the solvent ethanol have been determined by a spectrophotometric method. The rate constants at −40 °C are related to the dissociation constants of the phenols in water at 25 °C and the value of α in the Brönsted relation is 0.84 ± 0.07; α drops to 0.44 ± 0.05 if the results for the substituted acetic acids (1) are included. The rate constants for the phenols are also correlated by the Hammett relationship, the ρ value at −40 °C being 1.82 ± 0.2. The activation energies range from 9.4 to 10.9 kcal/mole.The rate of reaction of trinitrobenzyl anion with 3-methylphenol at −30 °C is reduced by a factor of 12 if the phenol is deuterated at the OH group and the solvent is deuteroethanol. The large isotope effect confirms that the rate-determining step involves proton transfer from the OH group of the phenol. Substitution of lithium or potassium cations for the sodium cation does not affect the rate constant at −10 °C.In the reaction with 3-methylphenol, a measurable concentration of trinitrobenzyl anion remains at equilibrium and the equilibrium constant for the reaction is 1.3 ± 0.2 at 25 °C. The heat and entropy changes are approximately −6.5 kcal/mole and −21 e.u./mole respectively.