sepharose and Virus-Diseases

There has been an urgent need to quickly screen and isolate patients with viral infections from patients with similar symptoms at point-of-care. In this study, we introduce a new microfluidic method for detection of various viruses using rolling circle amplification (RCA) of pathogens on the surface of thousands of microbeads packed in microchannels. When a targeted pathogen meets the corresponding particular template, the DNAs are rapidly amplified into a specific dumbbell shape through the RCA process, forming a DNA hydrogel and blocking the flow path formed between the beads. Due to the significant increase in reaction surface area, the detection time was shortened to less than 15 min and the detection limit of various pathogens has been reached to 0.1 pM. By injecting the stained liquid, the existence of the target pathogens in a sample fluid can be determined with the naked eye. Furthermore, by integrating multi-channel design, simultaneous phenotyping of various infective pathogens (i.e., Ebola, Middle East respiratory syndrome (MERS), and others) in biological specimens can be performed at a point-of-care.

Topics: Biosensing Techniques; DNA Primers; Fluorescent Dyes; Humans; Hydrogels; Limit of Detection; Microfluidics; Microspheres; Molecular Diagnostic Techniques; Nucleic Acid Amplification Techniques; Sepharose; Time Factors; Virus Diseases; Viruses

1. Serum samples were collected from ten patients hospitalized for acute infections and from a control group of seven normal subjects. Tissue ferritin was obtained by purification of ferritin from normal human liver and from the ferritin standard of a commercially available assay kit. 2. The serum and tissue samples were incubated with concanavalin A--Sepharose, which has the ability to bind normal serum ferritin. 3. Concanavalin A, a plant lectin which binds to glucose, can be coupled to Sepharose particles and by incubation and centrifugation ferritin in normal serum can be absorbed to about 70%. The serum and tissue samples were incubated with concanavalin A--Sepharose and the ferritin content was measured before and after. 4. It was found that ferritin in the serum of patients with acute infections was absorbed to the same extent as in normal serum (about 80%), irrespective of the initial value. Only about 20% of the tissue ferritin was absorbed. 5. It is concluded that the ferritin in serum during infection is of the same glucosylated type as the ferritin normally present in serum, whereas intracellular ferritin is not glycosylated. This indicates that the elevation of serum ferritin during infection is caused by a release along the normal pathways, i.e. an augmented synthesis, not by leakage from damaged cells.

Topics: Absorption; Adult; Bacterial Infections; Concanavalin A; Ferritins; Humans; Liver; Male; Sepharose; Virus Diseases