Fluorescent Plate Reader
The Biomolecular Research Facility has a Molecular Devices SPECTRAmax  Gemini EM fluorescent plate reader. The Gemini plate reader measures fluorescence. It has limited luminescence reading capabilities , but cannot read absorbance.

 

Features of the SPECTRAmax instrument are:

  • dual monochromators (bandwidth 9 nm) allowing any fluorophore to be read within the range of 250 to 850 nm
  • fluorescent spectra capability
  • top or bottom reading of plates for adherent cells
  • well scanning
  • kinetic measurements
  • heating of plates
  • shaking of plate
  • SoftMax Pro 4.7 software for data analysis.
  • claimed sensitivity of 3 fmol/well of FITC with top reading

    • It can perform some time resolved fluorescence measurements but not polarized fluorescence. 

    The instrument can be used as a luminometer for some applications. It does not have the sensitivity of a dedicated luminometer, nor does it have injectors required for flash luminescence assays e.g. Promega Dual Glo Luciferase assays.
    Detection limit: 10 amol/well alkaline phosphatase 200 μL/well (obtained with Emerald IITM reagent from Applied Biosystems).

    Some applications for the instrument are:

    • measuring live and dead cells in the same well, using Promega MultiTox reagents to measure proteinases specific for live and dead cells. This assay takes advantage of the instrument's ability to measure two fluorophores in the same assay and to scan wells to look for clumps of cells.
    • measuring glycogen  content of cultured cells
    • Caspase assay using luminescence
    • quantitation of DNA and RNA with PicoGreen or other dyes
    • cathepsin assay
    • measuring relative cell numbers with Alamar Blue
    • intracellular calcium
    • protein estimation
    • cholesterol assay

    One user is quantitating DNA at 5 ng/mL. 

    The instrument is located in  Jordan Hall room 1074, in the Biomolecular Research Facility.  Users need to contact Dr. John Shannon of the Biomolecular Research Facility at 3-9399 or by email at jds1c@virginia.edu to establish an account on the computer which controls the instrument. Dr. Shannon will offer basic training for the operation of the instrument. Instructions to get started are available.

    Users supply their own plates. For top read fluorescence, all black plates are recommended. For luminescence, white plates which should be kept out of light to reduce light emission from the plastic.

    The instrument is user operated. The current charges for use are $12 for the first 10 minutes, $16/hr for longer periods.