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1.
Endocrinol. diabetes nutr. (Ed. impr.) ; 67(1): 43-52, ene. 2020. tab
Artigo em Espanhol | IBECS | ID: ibc-186146

RESUMO

Antecedentes y objetivos: El alelo de riesgo (G) de la variante rs10830963 en el gen del receptor de melatonina 1 B (MTNR1B) se relaciona con la obesidad. En este trabajo evaluamos el efecto de este SNP sobre los factores de riesgo cardiovascular y la pérdida de peso secundaria a 2 dietas hipocalóricas. Métodos: Trescientos sesenta y un sujetos obesos fueron asignados aleatoriamente durante 3 meses (dieta M: dieta hipocalórica alta en grasas monoinsaturadas vs. dieta P: dieta hipocalórica alta en grasas poliinsaturadas). Se midieron los parámetros antropométricos, glucemia en ayunas, proteína C reactiva, concentración de insulina, resistencia a la insulina (HOMA-IR), perfil de lípidos y los niveles de adipocitoquinas. Se evaluó el genotipo del polimorfismo del gen MTNR1B (rs10830963). Resultados: Todos los parámetros antropométricos, la presión arterial sistólica y los niveles de leptina disminuyeron en todos los sujetos después de ambas dietas. Esta mejora de los parámetros antropométricos fue mayor en los no portadores del alelo G que en los portadores del alelo G. Tras la intervención con dieta M (CC vs. CG + GG), el colesterol total (delta: -10,4 ± 2,1 mg/dl vs. -6,4 ± 1,2 mg/dl: p < 0,05), colesterol LDL (delta: -7,1 ± 0,9 mg/dl vs. –2,8 ± 0,8 mg/dl: p < 0,05), insulina (delta: -3,0 ± 0.8 UI/l vs. -2,0 ± 1,0 UI/l: p < 0,05) y HOMA IR (delta: -3,4 ± 1,0 unidades vs. -2,9 ± 0,9 unidades: p < 0,05) mejoraron en los no portadores del alelo G. Tras la dieta P, en el grupo de sujetos sin alelo G, los niveles de insulina (delta: -2,9 ± 1,0 UI/l vs. -0,6 ± 0,2 UI/l: p < 0,05) y HOMA-IR (delta [CC vs. CG + GG]: -0,8 ± 0,2 unidades vs. -0,4 ± 0,3 unidades: p < 0,05) también disminuyeron. Conclusiones: Nuestro estudio detectó una relación de la variante rs10830963 de MTNR1B con la pérdida de peso corporal y la modificación de la resistencia a la insulina inducida por 2 dietas hipocalóricas diferentes. Solo la dieta hipocalórica enriquecida en grasa monoinsaturada y los no portadores del alelo G mostraron un efecto significativo sobre las lipoproteínas


Background & aims: The risk allele (G) of rs10830963 in the melatonin receptor 1 B (MTNR1B) gene presents an association with obesity. We study the effect of this SNP on cardiovascular risk factors and weight loss secondary to 2hypocaloric diets. Methods: 361 obese subjects were randomly allocated during 3 months (Diet M - high monounsaturated fat hypocaloric diet vs. Diet P – high polyunsaturated fat hypocaloric diet). Anthropometric parameters, fasting blood glucose, C-reactive protein (CRP), insulin concentration, insulin resistance (HOMA-IR), lipid profile and adipocytokines levels were measured. Genotype of MTNR1B gene polymorphism (rs10830963) was evaluated. Results: All anthropometric parameters, systolic blood pressure and leptin levels decreased in all subjects after both diets. This improvement of anthropometric parameters was higher in non G allele carriers than G allele carriers. After dietary intervention with Diet M, (CC vs. CG + GG); total cholesterol (delta: -10.4 ± 2.1 mg/dl vs. -6.4 ± 1.2 mg/dl: P <.05), LDL-cholesterol (delta:-7.1 ± 0.9 mg/dl vs. -2.8 ± 0.8 mg/dl: P <.05), insulin (delta:-3.0 ± 0.8 UI/L vs. -2.0 ± 1.0 UI/L: P<.05) and HOMA-IR (delta:-3.4 ± 1.0 units vs. -2.9 ± 0.9 units: P<.05) improved in no G allele carriers. After Diet P, in the group of subjects without G allele CC, insulin levels (delta: -2.9 ± 1.0 UI/L vs. -0.6 ± 0.2 UI/L: P <.05) and HOMA-IR (delta (CC vs. CG + GG): -0.8 ± 0.2 units vs. -0.4 ± 0.3 units: P <.05) decreased, too. Conclusions: Our study detected a relationship of rs10830963 MTNR1B SNP with body weight loss and insulin resistance modification induced by 2 different hypocaloric. Only monounsaturated enriched hypocaloric diet and in no-G allele carriers showed a significant effect on lipoproteins


Assuntos
Humanos , Adolescente , Adulto Jovem , Adulto , Pessoa de Meia-Idade , Idoso , Polimorfismo Genético , Gorduras na Dieta/uso terapêutico , Resistência à Insulina , Perda de Peso/efeitos dos fármacos , Obesidade/dietoterapia , Fatores de Risco , Gorduras na Dieta/metabolismo , Doenças Cardiovasculares , Antropometria , Glicemia , Composição Corporal , Relação Cintura-Quadril , Radioimunoensaio/métodos
2.
J S Afr Vet Assoc ; 90(0): e1-e6, 2019 Oct 22.
Artigo em Inglês | MEDLINE | ID: mdl-31714111

RESUMO

The Coat-A-Count® radioimmunoassay has been long and widely used to determine the concentration of progesterone in serum or plasma of bitches (progRIA), but was discontinued in 2014. The Immulite® 1000 LKPG1 chemiluminescence immunoassay has gained prominence since 2003 to determine the concentration of progesterone in serum of bitches, but the assay changed in 2012 (Immulite® 1000 LKPW1). This study assessed the feasibility of using Immulite® 1000 LKPW1 (progImm) to estimate the time of clinically relevant events during oestrus and compared progRIA and progImm 2 and 3 days after the first or only day of the luteinizing hormone surge (LH1). ProgImm first exceeded 5.1 nmol/L on the same day that progRIA first exceeded 6 nmol/L, a proxy for the occurrence of the LH surge, or the day before in 28 of 31 (90%) of oestrous periods. ProgImm first exceeded 13.6 nmol/L on the same day that progRIA first exceeded 16 nmol/L (a proxy for the day of ovulation) or the day before in 34 of 35 (97%) oestrous periods. ProgImm first exceeded 5.4 nmol/L on LH1 or the day before in 24 of 25 (95%) of oestrous periods. The median of progImm 2 days after LH1 was 1.2 nmol/L lower than the 10.7 nmol/L of progRIA (p = 0.001). The mean of progImm 3 days after LH1 was 2.2 nmol/L lower than the 19.0 nmol/L of progRIA (p 0.001). In conclusion, the days on which progImm first exceeded 5.1 nmol/L, 13.6 nmol/L and 5.4 nmol/L effectively estimate the days on which progRIA reached 6 nmol/L or 16 nmol/L or LH1.


Assuntos
Cães/sangue , Medições Luminescentes/veterinária , Detecção da Ovulação/veterinária , Progesterona/sangue , Radioimunoensaio/veterinária , Animais , Tomada de Decisão Clínica , Estro/sangue , Feminino , Medições Luminescentes/métodos , Detecção da Ovulação/métodos , Radioimunoensaio/métodos , Reprodução/fisiologia
3.
Vet Immunol Immunopathol ; 215: 109904, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31420068

RESUMO

During immune activation, CD25 is expressed by T cells, and its soluble form (sCD25) is released into the extracellular matrix and the bloodstream. In humans, serum sCD25 concentrations are used as a surrogate marker for autoimmune diseases, malignancies, and transplant rejection. However, a canine-specific assay for the measurement of sCD25 in dog serum has not previously been described. Therefore, the aims of this study were to develop and analytically validate a radioimmunoassay to measure sCD25 in canine serum, to establish a reference interval for canine sCD25, and to test the clinical utility of this assay with serum samples for dogs with various diseases. A competitive radioimmunoassay (RIA) was developed and analytically validated. Analytical validation consisted of lower limit of detection (LLOD), dilutional parallelism, spiking recovery, and intra- and inter-assay variability using pooled surplus canine serum samples. A reference interval was established in healthy dogs and serum samples from dogs with various types of neoplasia, IBD, liver disease, suspected pancreatitis, or suspected small intestinal disease and serum samples with an increased C-reactive protein concentration (CRP) were analyzed to test the clinical utility of the assay. LLOD was calculated to be 0.5 ng/mL. The mean (±SD) observed-to-expected ratio (O/E) for serial dilutions was 101.7 ±â€¯14.0%, and the mean (± SD) O/E for spiking recovery was 93.2 ±â€¯4.2%. Coefficients of variation (CVs) for intra-assay variability were ≤12.5% (mean ±â€¯SD: 7.5 ±â€¯4.2%), and inter-assay CVs were ≤15.7% (mean ±â€¯SD: 11 ±â€¯4.4%). A reference interval (RI) for canine sCD25 of 1.2-4.2 ng/mL was established from a population of 112 clinically healthy dogs. Dogs with neoplasia and dogs with suspected small intestinal disease had decreased concentrations of serum sCD25 when compared to healthy dogs (p < 0.0001, respectively). However, the majority of clinical samples used in this study were within the reference interval. Median concentrations of serum sCD25 were 1.9 ng/mL for healthy dogs. Dogs with cancer, IBD, liver disease, suspected pancreatitis, or suspected small intestinal disease, as well as sera with an increased serum CRP concentration, had median serum sCD25 concentrations of 1.6 ng/mL, 2.1 ng/mL, 2.2 ng/mL, 1.7 ng/mL, 1.5 ng/mL, and 1.8 ng/mL, respectively. Thus, the RIA described here is linear, accurate, precise, and reproducible for measuring sCD25 in canine serum. However, this assay shows little clinical utility of sCD25 as a biomarker for dogs with inflammatory, autoimmune, and/or neoplastic conditions.


Assuntos
Doenças do Cão/sangue , Cães/sangue , Subunidade alfa de Receptor de Interleucina-2/sangue , Radioimunoensaio/veterinária , Animais , Doenças do Cão/imunologia , Ensaio de Imunoadsorção Enzimática/veterinária , Humanos , Radioimunoensaio/métodos , Valores de Referência , Sensibilidade e Especificidade
4.
J Pineal Res ; 67(1): e12572, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30919486

RESUMO

There has been increased interest in the measurement of melatonin in plasma and saliva recently either as a marker of circadian phase or to understand the physiological role of melatonin. For both situations, there is a need for a specific assay for melatonin that is sensitive enough to detect low concentrations (<2 pg/mL). Since the mid-1970s, there have been many assays developed to measure melatonin in blood and saliva. Radioimmunoassays and ELISA have predominated because of their relative simplicity and high throughput. In this review, I show that the early radioimmunoassays while providing valuable information about nocturnal melatonin levels in humans, generally produced inaccurate basal (daytime) levels. Mass spectrometry assays, however, have provided us with the target values that immunoassays need to achieve, that is, daytime plasma melatonin levels <1 pg/mL. There are now many contemporary commercial assays available utilising both RIA and ELISA technologies, but not all achieve the standards set by the mass spectrometry assays. The performance of these assays is reviewed. I conclude with recommendations on issues researchers need to consider when conducting melatonin studies, including the importance of time of day of collection, validation of assays, the potential causes of poor assay specificity at low levels, the advantages/disadvantages of using saliva vs plasma and extraction assays vs direct assays, kit manufacturers responsibilities and the reporting requirements when publishing melatonin studies.


Assuntos
Ritmo Circadiano , Melatonina , Saliva/metabolismo , Animais , Biomarcadores/análise , Biomarcadores/metabolismo , Ensaio de Imunoadsorção Enzimática/história , Ensaio de Imunoadsorção Enzimática/métodos , História do Século XX , História do Século XXI , Humanos , Melatonina/análise , Melatonina/história , Melatonina/metabolismo , Radioimunoensaio/história , Radioimunoensaio/métodos
5.
Anal Biochem ; 570: 51-55, 2019 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-30771337

RESUMO

Scintillation proximity assay (SPA) is a type of radioimmunoassay (RIA). We apply ultrasound enhancement to the general SPA. All assay procedures, including the antibody coating and radiolabeled antigen binding are achieved by simply mixing then standing for 5 min in an ultrasound chamber. No additional incubation time is required. To further demonstrate the capability of the UE-SPA, a quantitative measurement of CD55 in various grades of colon tumors was assessed on human tissue slides. The results showed a significant correlation between CD55 expression and tumorigenesis. In conclusion, we confirmed that UE-SPA is a reliable, rapid and alternative to RIA.


Assuntos
Antígenos CD55/análise , Radioimunoensaio/métodos , Anticorpos Monoclonais/imunologia , Antígenos CD55/imunologia , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Humanos , Sonicação
6.
J Diabetes Investig ; 10(3): 685-689, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30307126

RESUMO

Anti-glutamic acid decarboxylase antibody (GADA) is an important islet cell-associated autoantibody for the diagnosis of autoimmune type 1 diabetes mellitus. In Japan, the GADA assay kit was recently changed from radioimmunoassay (RIA) to enzyme-linked immunosorbent assay (ELISA). Thereafter, a mismatched measurement between the two tests became apparent in clinical situations. The present study aimed to clarify the actual extent of mismatch between the two measurements on a larger-scale real-world clinical practice. In this cross-sectional non-local/non-hospital-based study, we collected anonymized data on GADA levels of 598 participants, who were simultaneously measured with GADA-RIA and GADA enzyme-linked immunosorbent assay tests. We found that 34% of the GADA-RIA-positive participants showed negative results in the GADA enzyme-linked immunosorbent assay test; the mismatch was predominantly observed in participants with relatively low GADA-RIA levels (<32 U/mL). This considerable mismatch might lead to physicians' confusion in diagnosing type 1 diabetes mellitus.


Assuntos
Autoanticorpos/sangue , Diabetes Mellitus Tipo 1/diagnóstico , Ensaio de Imunoadsorção Enzimática/métodos , Glutamato Descarboxilase/imunologia , Radioimunoensaio/métodos , Autoanticorpos/imunologia , Estudos Transversais , Diabetes Mellitus Tipo 1/sangue , Diabetes Mellitus Tipo 1/imunologia , Diagnóstico Diferencial , Seguimentos , Humanos , Prognóstico , Inquéritos e Questionários
7.
J Diabetes Investig ; 10(4): 990-996, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30582775

RESUMO

AIM/INTRODUCTION: Autoantibodies to the 65 kDa isoform of glutamic acid decarboxylase (GADA) are a valuable diagnostic and predictive marker for type 1 diabetes. Recently, it has been reported that a significant proportion of sera in the commercial RSR radioimmunoassay (RIA) that have tested positive for GADA have then turned negative in RSR enzyme-linked immunosorbent assay (ELISA) tests in patients with type 1 diabetes. The present study aimed to investigate whether the GADA result discrepancies between RSR-RIA and RSR-ELISA are related to autoantibody affinity. METHODS: GADA affinity was measured by a competitive binding experiment using unlabeled recombinant human GAD65 in 12 discordant samples (5 RIA[+]/ELISA[-] and 7 RIA[-]/ELISA[+] sera). Furthermore, the effect of the initial incubation time on the GADA positivity was also examined using the ELISA test. RESULTS: GADA affinities were >1010  L/mol in two of five RIA(+)/ELISA(-) and all of seven RIA(-)/ELISA(+) sera. After an initial incubation time longer than the recommended 1 h, the GADA titer in three of five RIA(+)/ELISA(-) sera and all RIA(-)/ELISA(+) sera increased 1.6- to 100-fold. However, the titer in 12 GADA-negative sera from healthy controls remained unchanged after the longer incubation. The increment ratio of GADA titer was positively correlated with GADA affinity (r = 0.991, P < 0.001). CONCLUSIONS: The RSR-RIA test identifies both high- and low-affinity GADA, whereas the RSR-ELISA test identifies only high-affinity GADA. A longer initial incubation time in the RSR-ELISA test increases the sensitivity of GADA with the same specificity in patients with type 1 diabetes.


Assuntos
Afinidade de Anticorpos , Autoanticorpos/sangue , Autoantígenos/imunologia , Diabetes Mellitus Tipo 1/diagnóstico , Ensaio de Imunoadsorção Enzimática/métodos , Glutamato Descarboxilase/imunologia , Radioimunoensaio/métodos , Adulto , Idoso , Idoso de 80 Anos ou mais , Autoanticorpos/imunologia , Diabetes Mellitus Tipo 1/sangue , Diabetes Mellitus Tipo 1/imunologia , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/diagnóstico , Diabetes Mellitus Tipo 2/imunologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Prognóstico , Curva ROC
8.
J Med Primatol ; 47(6): 402-411, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30187920

RESUMO

BACKGROUND: The objective of this study was to develop and analytically validate a radioimmunoassay (RIA) for the measurement of alpha1 -proteinase inhibitor (α1 -PI) concentrations in serum and feces from the common marmoset. METHODS: Serum samples (n = 30) and 3-day fecal samples (n = 30) were obtained from healthy marmosets. An RIA was established and validated by determination of sensitivity, working range, dilutional parallelism, spiking recovery, and intra- and interassay variability. A reference interval for mα1 -PI in serum and feces was established. RESULTS: Sensitivity and upper limit of the working range were 0.75 and 100.62 µg/L, respectively. Observed-to-expected (O/E) ratios for serial dilutions ranged from 89.9% to 123.0% (mean ±SD: 106.0 ± 11.5%) for 8 serum samples, and from 90.6% to 132.7% (mean ±SD: 107.6 ± 19.2%) for 4 fecal samples. O/E ratios for spiking recovery ranged from 97.6% to 104.4% (mean ±SD: 101.3 ± 3%) for 4 serum samples, and from 97.5% to 101.4% (mean ±SD: 99.2 ± 1.8%) for 4 fecal samples and 3 different spiking concentrations. Coefficients of variation (CV) for intra-assay variability for 8 serum samples ranged from 1.7% to 10.6% and 2.2% to 5.1% in the 8 fecal samples. The interassay CV for eight serum samples ranged from 1.3% to 9.9%, and from 1.0% to 6.7% in the 8 fecal samples. The reference interval in serum was determined to be 1047-1484 µg/L. The reference interval in serum was determined to be 1047-1484 µg/L. The reference interval for the 3-day mean fecal concentration, and 3-day maximum fecal concentration were determined to be 32.4-124.4 µg/g and 39.1-158.7 µg/g of feces, respectively. CONCLUSION: The developed assay is sensitive, linear, accurate, precise, and reproducible. Further studies are needed to determine the clinical utility of this assay.


Assuntos
Callithrix/metabolismo , Fezes/química , Radioimunoensaio/métodos , alfa 1-Antitripsina/metabolismo , Animais , Valores de Referência , Reprodutibilidade dos Testes , alfa 1-Antitripsina/sangue
9.
Diabetes Res Clin Pract ; 144: 260-269, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30213772

RESUMO

AIMS: We investigated the glucagon levels in patients with heart failure (HF), using long oral glucose tolerance test (OGTT). METHODS: In this prospective observational study, we enrolled 30 undiagnosed diabetes patients (age 69 ±â€¯10 years, 70% males, HbA1c 43 mmol/mol). A 4-h OGTT was performed. Glucose, insulin, and glucagon (radioimmunoassay [RIA] and sandwich ELISA [S-W] methods) were evaluated during 4-h. We compared glucagon levels between HF and non-HF patients. RESULTS: There were 11 HF and 19 non-HF patients. In patients with HF, glucagon (S-W) during 4-h was lower than in patients without HF, with no significant difference. The area under the curve (AUC) of glucagon (RIA) during 4-h was significantly lower among HF patients. Moreover, in patients with reduced left ventricular ejection fraction (LVEF) (<40%), AUC glucagon (RIA) was significantly lower than in patients with non-reduced EF (≥40%). However, there was no difference in glucagon values between the high E/e' (≥13.0) and low E/e' (<13.0) groups. CONCLUSIONS: Although glucagon (S-W) showed no significant difference in patients with and without HF, especially reduced LVEF, glucagon (RIA) secretion was significantly lower in HF patients than in patients without HF. It is suggested that low glucagon secretion might be correlated with low EF.


Assuntos
Diabetes Mellitus/fisiopatologia , Glucagon/metabolismo , Insuficiência Cardíaca/fisiopatologia , Radioimunoensaio/métodos , Disfunção Ventricular Esquerda/epidemiologia , Idoso , Feminino , Teste de Tolerância a Glucose , Humanos , Incidência , Japão/epidemiologia , Masculino , Estudos Prospectivos , Disfunção Ventricular Esquerda/metabolismo
10.
J Clin Endocrinol Metab ; 103(11): 3965-3973, 2018 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-30137438

RESUMO

Context: Current threshold values for primary aldosteronism (PA) diagnostic testing are based on measuring aldosterone (PAC) using immunoassays. Quantification of PAC by liquid chromatography-tandem mass spectrometry (LC-MS/MS) yields lower values. Objective: To compare aldosterone measurement by radioimmunoassay (RIA) with LC-MS/MS and evaluate performances of proposed LC-MS/MS-specific cutoffs for PA screening and confirmatory testing. Patients and Intervention: Forty-one patients underwent aldosterone/renin ratio (ARR) testing to screen for, and fludrocortisone suppression testing (FST) to confirm or exclude, PA. Renin (DRC) was measured by chemiluminescent immunoassay. Results: Median serum PACLC-MS/MS was 27.8% lower (P < 0.05) than plasma PACRIA in 164 pairs of FST samples. A positive correlation (Spearman coefficient, 0.894, P < 0.01; Pearson r coefficient, 0.861, P < 0.01) was observed between the two assays. Thirty-seven patients showed consistent FST diagnoses (29 positive, 8 negative), whereas four showed inconsistent FSTs by the two assays. Good agreement (κ coefficient, 0.736; P < 0.01) was observed between the current FST diagnostic PACRIA cutoff of 165 pmol/L and the proposed PACLC-MS/MS cutoff of 133 pmol/L. Among 37 patients with consistent FST results, no differences were observed in sensitivity (89.7% vs 93.1%) or specificity (87.5% vs 87.5%) for PA screening between the current ARR cutoff of 70 pmol/mU (PACRIA/DRC) and the proposed cutoff of 55 pmol/mU (PACLC-MS/MS/DRC). Conclusions: Adjustment of the current cutoffs for PA diagnostic testing is necessary if PAC is measured by LC-MS/MS. Our preliminary results suggest that the proposed LC-MS/MS cutoffs for ARR and FST perform as well as current RIA cutoffs.


Assuntos
Aldosterona/sangue , Hiperaldosteronismo/diagnóstico , Hipertensão/etiologia , Programas de Rastreamento/normas , Espectrometria de Massas em Tandem/normas , Adulto , Idoso , Cromatografia Líquida de Alta Pressão/métodos , Cromatografia Líquida de Alta Pressão/normas , Técnicas de Diagnóstico Endócrino/normas , Feminino , Fludrocortisona/administração & dosagem , Humanos , Hiperaldosteronismo/sangue , Hiperaldosteronismo/complicações , Hipertensão/sangue , Masculino , Programas de Rastreamento/métodos , Pessoa de Meia-Idade , Estudos Prospectivos , Radioimunoensaio/métodos , Radioimunoensaio/normas , Renina/sangue , Sensibilidade e Especificidade , Espectrometria de Massas em Tandem/métodos
11.
Acta Haematol ; 140(1): 10-17, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30007985

RESUMO

BACKGROUND: Direct antiglobulin test (DAT)-negative warm autoimmune hemolytic anemia (AIHA) is mainly caused by three mechanisms: red blood cell (RBC)-bound immunoglobulin (Ig)G below the detection limit of routine DAT; RBC-bound IgA or IgM; or low-affinity autoantibodies. Although most cases of DAT-negative AIHA are thought to be caused by RBC-bound IgG, and combinatory serological analyses are recommended, the relative ratios of each mechanism have not been clarified. METHODS: Two groups of patients with undiagnosed hemolytic anemia and negative conventional tube method-DAT (TM-DAT) were investigated using anti-IgA and anti-IgM sera, or column agglutination method-DAT (CM-DAT), respectively, in addition to radioimmunological quantitation of RBC-bound IgG. RESULTS: Three of 73 patients with DAT-negative AIHA showed positive RBC-bound IgA and normal amounts of RBC-bound IgG. Another group of 3 patients were RBC-bound IgM-positive, but only one of these showed normal amounts of RBC-bound IgG. In another group of patients with DAT-negative AIHA, 4 of the 20 showed positive CM-DAT and negative CM-DAT after washing RBCs. Three of these patients had normal amounts of RBC-bound IgG. Five patients with positive CM-DAT both before and after washing RBCs had high amounts of RBC-bound IgG. CONCLUSION: Relative ratios of patients with DAT-negative AIHA resulting from RBC-bound IgG, RBC-bound IgA, RBC-bound IgM, and low-affinity IgG were estimated as 80, 4, 1 and 15%, respectively. A new classification and diagnostic algorithm for DAT-negative AIHA were proposed.


Assuntos
Algoritmos , Anemia Hemolítica Autoimune/diagnóstico , Autoanticorpos/sangue , Teste de Coombs/métodos , Idoso , Criança , Eritrócitos/imunologia , Eritrócitos/metabolismo , Feminino , Humanos , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Imunoglobulina M/sangue , Laboratórios , Masculino , Pessoa de Meia-Idade , Radioimunoensaio/métodos
12.
Reprod Domest Anim ; 53(6): 1483-1490, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30058255

RESUMO

This is the first time that PAG determination using two different antisera raised against PAG molecules purified from both caprine (RIA-706) and bubaline placentas (RIA-860) is reported in water buffalo. Ninety-eight buffalo cows, belonging to a buffalo herd subjected to a synchronization and artificial insemination (AI) programme, were enrolled in this study. Blood samples were taken on days 0 (AI), 23, 25, 28, 30 and 45. Pregnancy was confirmed by ultrasonography on days 28 and 45. The blood of 20 buffaloes that had calved was tested every five days from the day of calving until day 50 postcalving. Differences in PAG concentrations were observed between pregnant and nonpregnant buffaloes starting from day 23 post AI using both RIA-706 and RIA-860 (p < 0.001). However, estimated mean concentrations of PAG measured by RIA-706 were higher than RIA-860 (p < 0.001) and Bland-Altman analysis showed biases ranged from 0.0 ng/ml at day 23 to 0.79 ng/ml at day 28 post AI. Moreover, RIA-706 showed greater sensitivity and accuracy both at 23 and 25 days of pregnancy. RIA-706 and RIA-860 decreased below 1 ng/ml from 40 and 30 days postpartum, respectively, suggesting that PAG are better recognized by the antisera raised against the caprine PAG in the postpartum period also. This is essential when using PAG as an appropriate marker of early pregnancy after postpartum for detecting new pregnancies. The results of this study show that the ability of RIA systems to recognize early PAG could be improved using antisera raised against PAG molecules isolated from caprine placenta.


Assuntos
Búfalos/sangue , Glicoproteínas/sangue , Proteínas da Gravidez/sangue , Radioimunoensaio/veterinária , Animais , Feminino , Cabras/imunologia , Soros Imunes , Inseminação Artificial/veterinária , Placenta/imunologia , Período Pós-Parto/sangue , Gravidez , Radioimunoensaio/métodos , Sensibilidade e Especificidade
13.
Biomed Chromatogr ; 32(11): e4323, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-29920712

RESUMO

Insulin is an effective therapeutic for diabetes, and the level of insulin in vivo is directly related to the health of diabetic patients. Traditionally, the concentrations of insulin in vivo are determined by the radioimmunoassay (RIA) method. In this study, we developed an LC-MS/MS method for the quantification of human insulin in dog plasma and directly compared the RIA and LC-MS/MS methods. Our LC-MS/MS method exhibited superior accuracy, efficiency and cost-effective for the pharmacokinetic (PK) assessment of human insulin. The LC-MS/MS method can quantitate human insulin and canine insulin simultaneously without cross-reactivity, making the analysis more efficient. The LLOQ of our LC-MS/MS method was 38.5 pg/mL, which was necessary to fully describe the PK profiles of endogenous and exogenous insulin in vivo. The direct comparison of PK data obtained from the two methods demonstrated that LC-MS/MS could be an alternative to the RIA method and should be widely used for the quantification of insulin drugs, especially in preclinical studies.


Assuntos
Cromatografia Líquida/métodos , Insulina Regular Humana/sangue , Insulina Regular Humana/farmacocinética , Radioimunoensaio/métodos , Espectrometria de Massas em Tandem/métodos , Animais , Cães , Feminino , Humanos , Limite de Detecção , Modelos Lineares , Masculino , Reprodutibilidade dos Testes
14.
Medicine (Baltimore) ; 97(26): e11032, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29952942

RESUMO

Vitamin D deficiency has become one of the most prevalent health problems in modern society. However, there has been no study that has reported the trend of vitamin D status in Asia. We performed an observational study to investigate the trend of vitamin D status in South Korea based on a representative national database acquired from the Korea National Health and Nutrition Examination Surveys (KNHANES) conducted from 2008 to 2014. A total of 39,759 patients were included in the final analyses. Serum 25-hydroxyvitamin D (25 (OH)D) levels were measured by radioimmunoassay. The overall mean serum level of 25 (OH)D was 45.7 nmol/L in males and 40.9 nmol/L in females in KNHANES 2008 to 2014. There was a significant trend toward lower serum 25 (OH)D levels from 2008 to 2014 in males by -1.2 (95% confidence interval [CI] -1.5 to -0.9) nmol/L per year and in female by -0.7 (95% CI -0.9 to -0.4) nmol/L per year. The overall mean serum level of 25 (OH)D in 2008 was 53.0 nmol/L in males and 45.7 nmol/L in females. It decreased to 43.2 nmol/L in males and 39.2 nmol/L in females in 2014. Vitamin D deficiency, defined as the serum 25 (OH)D level of <50 nmol/L, was found in 65.7% of males and 76.7% of females in overall population. A significant increasing trend of vitamin D deficiency was also observed. The prevalence of vitamin D deficiency in 2008 was 51.8% in males and 68.2% in females, but rose to 75.2% and 82.5%, respectively, in 2014. The present study demonstrated that vitamin D status in South Koreans is still deteriorating. More extensive and proactive measures are needed to improve vitamin D status in South Korea.


Assuntos
Deficiência de Vitamina D/sangue , Deficiência de Vitamina D/epidemiologia , Vitamina D/análogos & derivados , Vitamina D/sangue , Adolescente , Adulto , Idoso , Ásia/epidemiologia , Criança , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Inquéritos Nutricionais/métodos , Prevalência , Radioimunoensaio/métodos , República da Coreia/epidemiologia , Estações do Ano , Adulto Jovem
15.
Clin Biochem ; 58: 118-121, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-29709501

RESUMO

OBJECTIVES: We report a case of discordant total and free testosterone values in a patient with hypogonadism and juvenile hypophosphatasia after he initiated treatment with asfotase alfa, recombinant tissue non-specific alkaline phosphatase. METHODS: Total testosterone was evaluated using immunoassay pre and post initiation of therapy with asfotase alfa, and free testosterone was evaluated using radioimmunoassay and LC-MS/MS while on asfotase alfa therapy. RESULTS: Total testosterone measured by immunoassay was normal prior to therapy with asfotase alfa, and was low post initiation of therapy. During the same time frame, free testosterone measured using RAI and total testosterone measured using LC-MS/MS were normal on asfotase alfa therapy. This suggests assay interference with the total testosterone immunoassay. CONCLUSION: When laboratory results are discordant or do not match the clinical impression, the possibility of assay interference should be considered. Alternative laboratory methods free of the interference should be selected to evaluate these patients. HUMAN GENES DISCUSSED IN THE PAPER: ALPL gene, Approved name: Alkaline phosphatase, liver/bone/kidney, Synonym: Tissue non-specific alkaline phosphatase (TNSAP).


Assuntos
Fosfatase Alcalina/administração & dosagem , Fosfatase Alcalina/efeitos adversos , Hipofosfatasia/sangue , Hipofosfatasia/tratamento farmacológico , Imunoglobulina G/administração & dosagem , Imunoglobulina G/efeitos adversos , Proteínas Recombinantes de Fusão/administração & dosagem , Proteínas Recombinantes de Fusão/efeitos adversos , Testosterona/sangue , Adulto , Cromatografia Líquida/métodos , Humanos , Masculino , Espectrometria de Massas/métodos , Radioimunoensaio/métodos
16.
Lab Med ; 49(3): 259-267, 2018 Jul 05.
Artigo em Inglês | MEDLINE | ID: mdl-29608696

RESUMO

Background: Adrenal incidentalomas (AIs) are present in 4% of adults. As many as 30% may secrete cortisol autonomously in the absence of specific signs of overt hypercortisolism, in a phenomenon called subclinical hypercortisolism (SH). Diagnosis of SH is established by serum cortisol resistance to dexamethasone suppression. Methods: We compared serum cortisol concentrations, as determined by radioimmunoassay (RIA) and liquid chromatography/tandem mass spectronomy (LC/MS-MS), in 73 patients with AI group (52 with unilateral AI) and 34 control subjects in 3 scenarios: basal; after 1-mg dexamethasone suppression; and after 0.25-mg stimulation with cosyntropin, a synthetic derivative of adrenocorticotropic hormone (ACTH). To bolster evidence for the diagnosis of SH, we also measured salivary cortisol levels at 11 PM and after DST, as well as plasma ACTH and serum dehydroepiandrosterone sulfate (DHEA-S) levels. Results: We observed significant positive correlation (r = 0.9345, P <.001) for all 318 pairs of serum cortisol values, as measured by both methods. Conclusions: Serum cortisol concentrations in patients with AI and in control subjects were very similar, as measured by RIA and LC/MS-MS.


Assuntos
Análise Química do Sangue , Cromatografia Líquida/métodos , Hidrocortisona/sangue , Espectrometria de Massas/métodos , Radioimunoensaio/métodos , Neoplasias das Glândulas Suprarrenais/sangue , Adulto , Idoso , Idoso de 80 Anos ou mais , Análise Química do Sangue/métodos , Análise Química do Sangue/normas , Estudos de Casos e Controles , Feminino , Voluntários Saudáveis , Humanos , Modelos Lineares , Masculino , Pessoa de Meia-Idade , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Adulto Jovem
17.
Clin Exp Immunol ; 192(3): 348-365, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29431871

RESUMO

We examined the assay formats used to detect anti-drug antibodies (ADA) in clinical studies of the anti-tumour necrosis factor (TNF) monoclonal antibodies adalimumab and infliximab in chronic inflammatory disease and their potential impact on pharmacokinetic and clinical outcomes. Using findings of a recent systematic literature review of the immunogenicity of 11 biological/biosimilar agents, we conducted an ancillary qualitative review of a subset of randomized controlled trials and observational studies of the monoclonal antibodies against anti-TNF factor adalimumab and infliximab. Among studies of adalimumab and infliximab, the immunoassay method used to detect antibodies was reported in 91 of 111 (82%) and 154 of 206 (75%) adalimumab and infliximab studies, respectively. In most adalimumab and infliximab studies, an enzyme-linked immunosorbent assay or radioimmunoassay was used [85 of 91 (93%) and 134 of 154 (87%), respectively]. ADA incidence varied widely among assays and inflammatory diseases (adalimumab, 0-87%; infliximab, 0-79%). Pharmacokinetic and clinical outcomes were only reported for ADA-positive patients in 38 of 91 (42%) and 61 of 154 (40%) adalimumab and infliximab studies, respectively. Regardless of assay format or biological used, ADA formation was associated with lower serum concentrations, reduced efficacy and elevated rates of infusion-related reactions. Consistent with previous recommendations to improve interpretation of immunogenicity data for biologicals, greater consistency in reporting of assay methods and clinical consequences of ADA formation may prove useful. Additional standardization in immunogenicity testing and reporting, application of modern, robust assays that satisfy current regulatory expectations and implementation of international standards for marketed products may help to improve our understanding of the impact of immunogenicity to biologics.


Assuntos
Adalimumab/imunologia , Anticorpos/imunologia , Antirreumáticos/imunologia , Ensaio de Imunoadsorção Enzimática/métodos , Infliximab/imunologia , Radioimunoensaio/métodos , Adalimumab/uso terapêutico , Antirreumáticos/uso terapêutico , Humanos , Infliximab/uso terapêutico , Fator de Necrose Tumoral alfa/biossíntese , Fator de Necrose Tumoral alfa/imunologia
18.
Clin Lab ; 64(1): 69-75, 2018 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-29479885

RESUMO

BACKGROUND: Recent reports have described inherent problems with androgen immunoassays compared with mass spectrometry analyses. In this study, a new liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed according to CLSI rules. The developed method was compared with two immunoassay methods, the enzyme-linked immunosorbent assay (ELISA) and radioimmunoassay (RIA). METHODS: After liquid-liquid extraction, a Shimadzu Prominence LC unit coupled to an ABSCIEX API 3200 mass spectrometer with atmospheric pressure chemical ionization was used to quantify serum androstenedione levels. Serum androstenedione results taken from tandem mass spectrometry were compared with the immunoassays. RESULTS: The androstenedione assay was linear up to 50 ng/mL. Lower limit of quantitation and lower limit of detection were 0.195 ng/mL and 0.097 ng/mL, respectively. This method was not affected by matrix effect and other steroid hormone interferences. In this study, the obtained recovery was 69 - 99%, carryover value was determined as 0.026 ng/mL. According to the results of an interference study, androstenedione bias % did not exceed the limit of allowable bias % and 88.7% recovery was acquired for androstenedione. In the LC-MS/MS and ELISA comparison study, the slope value was found as 18.412, intercept -22.87, and r2 value as 0.1033. In the LC-MS/MS and RIA comparison study, slope value was found as 1.085, intercept 0.4541, and r2 value as 0.3712. In the RIA and ELISA comparison study, slope value was found as 9.57, intercept -15.5, and r2 value as 0.19. CONCLUSIONS: The LC-MS/MS provides agreement with the results of radioimmunoassay but not with ELISA. This method offers better selectivity compared to immunoassay systems.


Assuntos
Androstenodiona/sangue , Cromatografia Líquida/métodos , Imunoensaio/métodos , Espectrometria de Massas em Tandem/métodos , Calibragem , Ensaio de Imunoadsorção Enzimática/métodos , Feminino , Humanos , Masculino , Radioimunoensaio/métodos , Reprodutibilidade dos Testes
19.
J Exp Biol ; 221(Pt 3)2018 02 08.
Artigo em Inglês | MEDLINE | ID: mdl-29439063

RESUMO

Neuropeptides are one of the most diverse classes of signaling molecules and have attracted great interest over the years owing to their roles in regulation of a wide range of physiological processes. However, there are unique challenges associated with neuropeptide studies stemming from the highly variable molecular sizes of the peptides, low in vivo concentrations, high degree of structural diversity and large number of isoforms. As a result, much effort has been focused on developing new techniques for studying neuropeptides, as well as novel applications directed towards learning more about these endogenous peptides. The areas of importance for neuropeptide studies include structure, localization within tissues, interaction with their receptors, including ion channels, and physiological function. Here, we discuss these aspects and the associated techniques, focusing on technologies that have demonstrated potential in advancing the field in recent years. Most identification and structural information has been gained by mass spectrometry, either alone or with confirmations from other techniques, such as nuclear magnetic resonance spectroscopy and other spectroscopic tools. While mass spectrometry and bioinformatic tools have proven to be the most powerful for large-scale analyses, they still rely heavily on complementary methods for confirmation. Localization within tissues, for example, can be probed by mass spectrometry imaging, immunohistochemistry and radioimmunoassays. Functional information has been gained primarily from behavioral studies coupled with tissue-specific assays, electrophysiology, mass spectrometry and optogenetic tools. Concerning the receptors for neuropeptides, the discovery of ion channels that are directly gated by neuropeptides opens up the possibility of developing a new generation of tools for neuroscience, which could be used to monitor neuropeptide release or to specifically change the membrane potential of neurons. It is expected that future neuropeptide research will involve the integration of complementary bioanalytical technologies and functional assays.


Assuntos
Invertebrados/fisiologia , Neuropeptídeos/fisiologia , Vertebrados/fisiologia , Animais , Biologia Computacional/métodos , Imuno-Histoquímica/métodos , Invertebrados/genética , Espectrometria de Massas/métodos , Optogenética/métodos , Radioimunoensaio/métodos , Vertebrados/genética
20.
Braz J Otorhinolaryngol ; 84(4): 448-452, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-28625809

RESUMO

INTRODUCTION: In patients with papillary thyroid carcinoma who have negative serum thyroglobulin after initial therapy, the risk of structural disease is higher among those with elevated antithyroglobulin antibodies compared to patients without antithyroglobulin antibodies. Other studies suggest that the presence of chronic lymphocytic thyroiditis is associated with a lower risk of persistence/recurrence of papillary thyroid carcinoma. OBJECTIVE: This prospective study evaluated the influence of chronic lymphocytic thyroiditis on the risk of persistence and recurrence of papillary thyroid carcinoma in patients with negative thyroglobulin but elevated antithyroglobulin antibodies after initial therapy. METHODS: This was a prospective study. Patients with clinical examination showing no anomalies, basal Tg<1ng/mL, and elevated antithyroglobulin antibodies 8-12 months after ablation were selected. The patients were divided into two groups: Group A, with chronic lymphocytic thyroiditis on histology; Group B, without histological chronic lymphocytic thyroiditis. RESULTS: The time of follow-up ranged from 60 to 140 months. Persistent disease was detected in 3 patients of Group A (6.6%) and in 6 of Group B (8.8%) (p=1.0). During follow-up, recurrences were diagnosed in 2 patients of Group A (4.7%) and in 5 of Group B (8%) (p=0.7). Considering both persistent and recurrent disease, structural disease was detected in 5 patients of Group A (11.1%) and in 11 of Group B (16.1%) (p=0.58). There was no case of death related to the disease. CONCLUSION: Our results do not support the hypothesis that chronic lymphocytic thyroiditis is associated with a lower risk of persistent or recurrent disease, at least in patients with persistently elevated antithyroglobulin antibodies after initial therapy for papillary thyroid carcinoma.


Assuntos
Autoanticorpos/sangue , Carcinoma Papilar/etiologia , Carcinoma Papilar/cirurgia , Doença de Hashimoto/complicações , Neoplasias da Glândula Tireoide/etiologia , Neoplasias da Glândula Tireoide/cirurgia , Adolescente , Adulto , Idoso , Carcinoma Papilar/sangue , Feminino , Doença de Hashimoto/sangue , Humanos , Medições Luminescentes/métodos , Masculino , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/etiologia , Estudos Prospectivos , Radioimunoensaio/métodos , Medição de Risco , Fatores de Risco , Estatísticas não Paramétricas , Neoplasias da Glândula Tireoide/sangue , Tireoidectomia/métodos , Adulto Jovem
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