National Veterinary Institute Sweden

Tanya von Rosen (Visiting researcher)

    Activity: Visiting an external institutionVisiting another research institution

    Description

    EPIZONE Short term mission

    The main focus of the visit to SVA was a thorough introduction to the use and application of the Luminex/BioPlex microsphere (bead) assay based technology. As described earlier in the “Plan of activities” the principle of the assay is similar to a capture sandwich immunoassay, but instead of coating the well in a plate, beads are being coated, which allows for the simultaneous analysis of multiple proteins, peptides, and nucleic acids in a single well of a 96-well microtiter plate. As the expectations to the end output of the short term mission is “a novel serological multiplex assay for detection and differentiation of antibodies against CSFV” the introductorily training was based on an already developed microsphere-based immunoassay for the detection of BVDV antibodies that was recently developed by a group of researchers from SVA/SLU (Xia et al., 2010), including Sandor Belak, Hongyan Xia and Liu Lihong. The laboratory training was supervised by Hongyan Xia and Liu Lihong. More specific did the introductorily training include intensive training in each of the various steps integrated in building up the BVDV microsphere-based immunoassay: - coupling of capture molecules to magnetic beads, different beads and capture molecules was used - optimization and confirmation of the effectiveness of the coupling, it was here additionally investigated whether it makes a difference for the optimal coating concentration whether magnetic/non-magnetic beads are used - running of the established BVDV microsphere-based immunoassay with beads I had coupled with capture molecules, known positive and negative clinical samples were used for verification - optimization of an assay – what parameters to adjust: samples (diluted/undiluted), secondary Ab (what concentration), incubation time, volumes etc. - how to get the most optimal performance and reproducible results (the minimum bead count needed for statistically significant analyses and optimal bead regions to use in a multiplexed assay) - validation of results – how to determine whether irregular assay results are due to hardware, software, or assay artifacts The most important achievements from the short-term research stay at SVA have been: - that I have gained routine in coupling magnetic beads with capture molecules - that I have gained routine in optimization and confirmation of the effectiveness of the coupling - that I have become familiar in using the Luminex machine system as well as building up assays, which on my own hand would have taken months - that I have got protocols to bring home which I can easily modify for own purposes - that I have got a network of scientist that I know I can always contact in case I need help solving problems regarding the Luminex/Bioplex system.
    Period24 Jan 201111 Feb 2011
    VisitingNational Veterinary Institute