RESUMO
BACKGROUND: Guidelines for sweat chloride testing endorse a minimum sweat rate for reporting results. Bilateral sweat collection is recommended, but if both sites fail to meet the minimum rate (quantity not sufficient, QNS), the test should be repeated. In this study, we examine the correlation between sweat rate and sweat chloride concentration ([Cl-]), assess the accuracy of specimens collected at suboptimal rates, and investigate the use of pooled bilateral specimens for chloride measurement. METHODS: Pearson correlation was employed to analyze the relationship between sweat rate and chloride concentration, [Cl-], in 674 macroduct collections. Weighted kappa was evaluated to determine cystic fibrosis (CF) diagnostic classification concordance for 18 tests with paired arms above vs below the minimum sweat rate. Deming regression was applied to compare [Cl-] from pooled bilateral specimens vs neat specimens in 27 collections with residual volume available after clinical testing. RESULTS: Pearson correlation of sweat rate vs [Cl-] was minimal (r = -0.0735) across specimens with varying rates and [Cl-]. There was substantial agreement in CF diagnostic classification between arms for bilateral collections with discordant sweat rates. Regression analysis of [Cl-] in pooled vs nonpooled specimens revealed a slope of 0.984 and an intercept of 0.796. CONCLUSIONS: Negligible correlation of sweat rate and [Cl-] suggests the minimum sweat rate for macroduct collectors may be overly stringent. Reporting of [Cl-] in specimens with ≥10 µL (rate ≥0.3 µL/min) may reduce QNS rates without compromising diagnostic accuracy. Preliminary data suggests pooling of bilateral collections may be a feasible option to achieve the required volume for testing.
Assuntos
Fibrose Cística , Suor , Humanos , Suor/química , Cloretos , Fibrose Cística/diagnóstico , NonoxinolAssuntos
Aminoglicosídeos/efeitos adversos , Anticorpos Monoclonais Humanizados/efeitos adversos , Antineoplásicos Imunológicos/efeitos adversos , Hemólise/efeitos dos fármacos , Leucemia Mieloide Aguda/sangue , Aminoglicosídeos/uso terapêutico , Anticorpos Monoclonais Humanizados/uso terapêutico , Antineoplásicos Imunológicos/uso terapêutico , Criança , Evolução Fatal , Gemtuzumab , Haptoglobinas/análise , Testes Hematológicos , Hemoglobinas/análise , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Masculino , Cuidados PaliativosRESUMO
PURPOSE: Effectively identifying the proteins present in the cellular secretome is complicated due to the presence of cellular protein leakage and serum protein supplements in culture media. A metabolic labeling and click chemistry capture method is described that facilitates the detection of lower abundance glycoproteins in the secretome, even in the presence of serum. EXPERIMENTAL DESIGN: Two stromal cell lines were incubated with tetraacetylated sugar-azide analogs for 48 h in serum-free and low-serum conditions. Sugar-azide labeled glycoproteins were covalently linked to alkyne-beads, followed by on-bead trypsin digestion and MS/MS. The resulting glycoproteins were compared between media conditions, cell lines, and azide-sugar labels. RESULTS: Alkyne-bead capture of sugar-azide modified glycoproteins in stromal cell culture media significantly improved the detection of lower abundance secreted glycoproteins compared to standard serum-free secretome preparations. Over 100 secreted glycoproteins were detected in each stromal cell line and significantly enriched relative to a standard secretome preparation. CONCLUSION AND CLINICAL RELEVANCE: Sugar-azide metabolic labeling is an effective way to enrich for secreted glycoproteins present in cell line secretomes, even in culture media supplemented with serum. The method has utility for identifying secreted stromal proteins associated with cancer progression and the epithelial-to-mesenchymal transition.
Assuntos
Azidas/química , Carboidratos/química , Glicoproteínas/metabolismo , Células Estromais/metabolismo , Alcinos/química , Linhagem Celular Tumoral , Cromatografia Líquida de Alta Pressão , Química Click , Transição Epitelial-Mesenquimal , Glicoproteínas/química , Humanos , Espectrometria de Massas em TandemRESUMO
The forensic applications of DNA-based human identity laboratory testing are often underappreciated. Molecular biology has seen an exponential improvement in the accuracy and statistical power provided by identity testing in the past decade. This technology, dependent upon an individual's unique DNA sequence, has cemented the use of DNA technology in the forensic laboratory. This paper will discuss the state of modern DNA-based identity testing, describe the technology used to perform this testing, and describe its use as it relates to forensic applications. We will also compare individual technologies, including polymerase chain reaction (PCR) and Southern Blotting, that are used to detect the molecular differences that make all individuals unique. An increasing reliance on DNA-based identity testing dictates that healthcare providers develop an understanding of the background, techniques, and guiding principles of this important forensic tool.