RESUMEN
Protein carbamylation, a nonenzymatic posttranslational modification promoted during uremia, is linked to a poor prognosis. In the present study, carbamylation of serum albumin was assayed using the symmetry factor on a capillary electrophoresis instrument (Helena V8). The symmetry factor has been defined as the distance from the center line of the peak to the back slope, divided by the distance from the center line of the peak to the front slope, with all measurements made at 10% of the maximum peak height. Serum albumin, creatinine, and urea concentrations were assayed using routine methods, whereas uremic toxins were determined using HPLC. In vitro carbamylation induced a marked albumin peak asymmetry. Reference values for the albumin symmetry factor were 0.69-0.92. In kidney patients, albumin peak asymmetry corresponded to the chronic kidney disease stage (p < 0.0001). The symmetry factor correlated well with serum urea (r = -0.5595, p < 0.0001) and creatinine (r = -0.5986, p < 0.0001) concentrations. Several protein-bound uremic toxins showed a significant negative correlation with the symmetry factor. Morphology of the albumin fraction was not affected by presence of glycated albumin and protein-bound antibiotics. In conclusion, the presented method provides a simple, practical way for monitoring protein carbamylation.
Asunto(s)
Carbamatos/sangre , Electroforesis Capilar/métodos , Albúmina Sérica/análisis , Adulto , Anciano , Carbamatos/química , Nefropatías Diabéticas , Femenino , Humanos , Masculino , Persona de Mediana Edad , Insuficiencia Renal Crónica/sangre , Albúmina Sérica/química , Urea/sangreRESUMEN
INTRODUCTION: Molecular diagnostic systems for point-of-care (POC) testing are nowadays routinely used and are part of many labs. Although often intended for bedside use outside of the microbiology lab, there is still room for expansion. AREAS COVERED: This review discusses the two techniques that are currently the most widespread, real-time polymerase-chain reaction (RT-PCR) and loop-mediated isothermal amplification (LAMP). An overview is provided of the various manufacturers and products as well as the evidence and current use in clinical practice. The article further sheds light on some newer techniques, such as CRISPR-based diagnostics and lab-on-a-chip, which are still in development. EXPERT OPINION: With many new platforms and techniques still in the pipeline and their potential currently not yet fully exploited, we expect the use of molecular POC testing to increase in the years to come. However, even when used in hospital - in lab, the main advantages of the tests being fast and easy to perform already provide significant benefits in terms of patient outcome.
Asunto(s)
Enfermedad Crítica , Pruebas en el Punto de Atención , Humanos , Técnicas de Amplificación de Ácido Nucleico/métodos , Técnicas de Diagnóstico Molecular/métodos , Reacción en Cadena en Tiempo Real de la Polimerasa , Sistemas de Atención de Punto , Sensibilidad y EspecificidadRESUMEN
INTRODUCTION: Accurate quantification of the BCR::ABL1 fusion gene in whole blood is pivotal for the clinical management of chronic myeloid leukemia (CML) patients. The fusion protein encoded by BCR::ABL1 can vary in size, depending on the BCR and/or ABL1 gene breakpoint. The vast majority of CML patients have a p210 BCR::ABL1 fusion gene (M-BCR), which can be attributed to the presence of either e14a2 (b3a2) or e13a2 (b2a2) mRNA transcript junctions. METHODS: Twenty-five CML samples were analyzed in two different ISO15189-accredited centers that both use an Europe Against Cancer-based quantitative polymerase chain reaction (qPCR) protocol. Reanalysis of the sample set with transcript-specific standard curves and digital droplet PCR (ddPCR) were performed. RESULTS: qPCR quantification revealed a significant (up to 1 log) difference specifically for the e13a2 transcript variant in contrast to e14a2 transcripts (Hodges-Lehman 4.29; p < 0.001). Reanalysis of the sample set with transcript-specific standard curves abolishes the initial transcript-specific difference (Hodges-Lehman 0.003; p = 0.8192). Comparison of transcript-specific qPCR results of both centers with ddPCR, an absolute quantification method, showed a statically significant association, especially in the lower range, indicating the clinical utility of transcript-specific or absolute quantification methods. CONCLUSION: Our data show that differences between transcript-specific quantification might exist between centers, leading to potential clinical impact on the follow-up of CML patients. The use of transcript-specific standard curves for qPCR quantification, or absolute quantification, can significantly reduce these differences. Specific attention should be applied to the interpretation of quantification differences of CML patients that switch between diagnostic centers.
RESUMEN
INTRODUCTION: Biotin supplementation (mainly OTC preparations) has gained popularity. There are concerns about biotin interference in immunoassays and potential misdiagnosis, especially since the discovery of high dose therapy in MS. This review summarizes the dangers of biotin usage and possible countermeasures. METHODS: Immunoassays design determines whether positive or negative analytical errors may occur. Techniques using biotinylated reagent and biotin binding proteins may generate errors. In sandwich immunoassays, biotin causes lowered results. Competitive immunoassays are more vulnerable: biotin usage causes false increased results. The interference is platform dependent. Parameters vary in their susceptibility: a combination of false positives and negatives mimicking a coherent profile is dangerous, e.g. combining falsely lowered TSH with falsely elevated FT4/FT3 mimicking hyperthyreosis. Other susceptible parameters are thyroglobulin, DHEA-S, estradiol, testosterone, ferritin, progesterone, Vitamin D, Vitamin B12, PSA, PTH, LH, FSH, Troponins I and T, Pro-BNP. Digoxin and PSA may also be affected. Tumor markers and ß-hCG are robust. Inserts of serological markers of HIV, hepatitis B and C warn for biotin interference. RESULTS: Manufacturers have made assays less vulnerable for biotin interference. In doubtful cases, it is helpful to determine testosterone in females and estrogen in males. Both are elevated if biotin interference is present. Biotin supplementation should be discontinued. However, this is impossible in MS patients needing biotin, as interrupting this medication is discouraged. CONCLUSIONS: Solutions to overcome this interference are: informing patients prior to analysis (avoiding peak biotin values when sampling), choice of appropriate immunoassays, and use of biotin removing steps prior to analysis.
Asunto(s)
Biotina , Técnicas de Laboratorio Clínico , Errores Diagnósticos , Inmunoensayo , Reacciones Falso Positivas , Femenino , Humanos , MasculinoRESUMEN
We report on sample IS/17575 since it generated highly divergent results in the Belgian SARS-CoV-2 serology external quality assessment scheme. Sample IS/17575 was serum originating from a 30 years old male patient. 124 diagnostic laboratories analysed this sample. A total of 168 results was returned (including 5 doubles). Overall, 38 were positive. All tests against S1 were positive except the Euroimmun IgG ELISA and the Ortho clinical Diagnostics VITROS IgG CLIA. All tests against S1/S2 (Liaison, Diasorin) resulted in a signal above cutoff. Assays against RBD, mostly generate a negative result. An exception are the Wantai SARS-CoV-2 ELISA's. All tests targeting N protein were negative. The survey shows, when >6 months post-infection, assays targeting at least S1, and preferably S1 combined with S2, are the most sensitive. This finding accentuates the necessity of external quality assessment schedules and importance of antigenic composition of serologic SARS-CoV-2 assays.
Asunto(s)
Antígenos Virales/inmunología , Prueba Serológica para COVID-19/métodos , COVID-19/diagnóstico , Proteínas de la Nucleocápside de Coronavirus/inmunología , SARS-CoV-2/inmunología , Glicoproteína de la Espiga del Coronavirus/inmunología , Adulto , Anticuerpos Antivirales/inmunología , Bélgica , Pruebas Diagnósticas de Rutina , Ensayo de Inmunoadsorción Enzimática , Humanos , Inmunoglobulina A/inmunología , Inmunoglobulina G/inmunología , Inmunoglobulina M/inmunología , Masculino , Fosfoproteínas/inmunología , Sensibilidad y EspecificidadRESUMEN
Being a member of the IL-17 family, comprising six structurally related ligands, IL-17A is a cytokine, produced by multiple cell types, such as CD4+αß T cells, γδ T cells, natural killer cells, neutrophils, macrophages, dendritic cells, lymphoid tissue inducer cells, mast cells and plasma cells. IL-17A participates in tissue inflammation by inducing the expression of chemokines, proinflammatory cytokines and matrix metalloproteases. Besides its role in host defence against infectious diseases, IL-17A is involved in different autoimmune and inflammatory diseases. In this review, we will highlight the role of IL-17A in the pathogenesis of acute and chronic kidney diseases. Due to its pleiotropic character, IL-17A is involved in the development of atherosclerosis, hypertension, diabetic nephropathy, ischaemia-reperfusion injury, fibrosis, glomerulonephritis, nephrotic syndrome, minimal change disease and acute renal allograft rejection. In addition, inhibition of IL-17A may be a promising therapeutic target to prevent end-stage renal disease.