Interday validation of of developed quantatation method
Ontology highlight
ABSTRACT:
ORGANISM(S): Fecal Bacteria
TISSUE(S): Feces
PROVIDER: ST001498 | MetabolomicsWorkbench |
REPOSITORIES: MetabolomicsWorkbench
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Project description:Glycomic-based approaches to discover potential biomarkers have shown great promise in their ability to distinguish between healthy and diseased individuals; these methods can identify when aberrant glycosylation is significant, but they cannot practically be adapted into widely implemented diagnostic assays because they are too complex, expensive, and low-throughput. We have developed a new strategy that addresses challenges associated with sample preparation, sample throughput, instrumentation needs, and data analysis to transfer the valuable knowledge provided by protein glycosylation into a clinical environment. Notably, the detection limits of the assay are in the single-digit picomole range. Proof of principle is demonstrated by quantifying the changes in the sialic acid content in fetuin. As the sialic acid content in proteins varies in a number of disease states, this example demonstrates the utility of the method for biomarker analysis. Furthermore, the developed method can be adapted to other biologically important saccharides, affording a broad array of quantitative glycomic analyses that are accessible in a high-throughput, plate-reader format. These studies enable glycomic-based biomarker discovery efforts to transition through the difficult landscape of developing a potential biomarker into a clinical assay.
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Project description:BackgroundDetecting SARS-CoV-2 using a simple real time molecular assay will be helpful for the mitigation efforts in low / middle income countries during the pandemic. We have developed and validated a rapid and simple real time loop mediated isothermal amplification assay (LAMP) for screening of SARS-CoV-2 infection in known infected and non-infected individuals.MethodsSix sets of primers were designed targeting the N-gene of the SARS-CoV-2 (Accession ID MN994468). LAMP reactions were performed using Warm Start 2X Master Mix and real-time PCR machine at 65 °C for 60 cycles with 15 s for each cycle. Results were read by visualizing turbidity under ultraviolet light and real time fluorescence detection through FAM channel of the real time PCR machine. We tested a total of 320 including 240 SARS CoV-2 positive (Ct values <40) and 80 SARS CoV-2 negative samples as tested by a real time RT-PCR using the newly developed LAMP assay.ResultsA total of 206 out of 240 SARS CoV-2 positive samples were tested positive by the newly developed LAMP assay with a sensitivity of 86%. All 80 SARS CoV-2 negative samples were tested negative by the newly developed LAMP assay with a specificity of 100%.ConclusionThe newly developed real time LAMP assay has a sensitivity of 86% and specificity of 100% compared to the real time RT-PCR for the detection of SARS CoV-2. The new assay will be useful to screen large number of samples if adopted to minimize the time and cost.
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Project description:BackgroundRisk perception is a multidimensional phenomenon that describes the individual's judgment of the likelihood of experiencing something unpleasant. Risk perception helps to understand how rheumatoid arthritis patients perceive disease-related-risks. We developed and validated a risk perception questionnaire for Spanish speaking rheumatoid arthritis patients.MethodsThe questionnaire development and validation was performed in 3 steps, using respective convenience samples. Step-1 included the conceptual model construction, 20 patient's interviews to identify components from the conceptual model-dimensions and 11 healthcare provider´s consultations who identified RA related manifestations/complications (network and frequencies analysis). Step-2 consisted of item generation and reduction and questionnaire feasibility (n = 100). Step-3 consisted of the questionnaire psychometric validation (n = 270), which included content, face, construct (exploratory factor analysis) and criterion validity (logistic regression analysis) and consistency and stability (Cronbach's α and test-retest).ResultsSamples were representative of typical RA outpatients. Initial conceptual model included 7 dimensions, 3 for probability and 1 each, for responsibility, prevention, control and for severity (Step-1). The final version was considered feasible by the patients and included 27 items (Step-2). A five-factor model was most appropriated and resulted in 68.8% of the variance explained: Cronbach's α = 0.90, intraclass-correlation-coefficient = 0.93 (95% CI = 0.90-0.95). A positive relation between number of external criteria from the charts and risk perception was found; all items had ≥80% agreement from experts; patients agreed about item´s semantic clarity (89%) and format adequacy (97%), (Step-3).ConclusionsThe risk perception questionnaire was valid and reliable to evaluate risk perception construct in RA outpatients; it can be incorporated to routine care and clinical research, and guide interventions to improve patient's health behaviors.
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