The use of black plates will result in lower raw signals, because the black color of the plate can quench the light. The use of white plates will result in higher raw signals, because the light is reflected maximally by the white color of the plate. Because of this, time-resolved fluorescence assays can be run in either black or white plates. This allows background autofluorescence to fade before you begin collecting emission signal from assays involving a long half-life fluorophore. Because of the longer half-life of the fluorescent signal, you can set up your instrument to incorporate a “lag time” or “delay time” between the time the fluorophore is excited, and the time you begin reading the emission signal (time-resolved mode). Examples of such fluorophores include Europium chelates and cryptates, Samarium chelates, and Terbium chelates and cryptates. Time-resolved fluorescence assays involve fluorophores that have longer half-lives (µsec – msec, for microplate-based assays). Time-resolved fluorescence assays (TRF, TR-FRET) We recommend running assays utilizing short half-life fluorophores in black plates to reduce background autofluorescence. These fluorophores have relatively-short half-lives (< µsec, for microplate-based assays). General fluorescence assays include traditional fluorophores such as fluorescein, cyanine 3, cyanine 5, green fluorescent protein (GFP), rhodamine, Texas Red, coumarin, and other fluorophores. General fluorescence assays (fluorescence intensity, fluorescence polarization, FRET) Time-resolved fluorescence assays, which use longer half-life fluorophores, can use either white or black plates (read below for more information). For this reason, black plates are typically recommended for fluorescence assays that use short half-life fluorophores. For example, higher excitation wavelengths (above 650 nm) usually cause less autofluorescence than wavelengths in the UV/Vis range.īecause white plates reflect light and black plates tend to quench light, background fluorescence will be higher in white plates as opposed to black plates. The severity of background autofluorescence can vary based on the excitation wavelength being used in a particular assay. Autofluorescence is triggered by the same excitation light used to excite the fluorophore in the fluorescence assay. Many components of assays buffers and biological samples can autofluoresce. white plates ( Information taken from PerkinElmer)Īutofluorescence is fluorescence resulting from substances other than the fluorophore-of-interest, and can negatively affect an assay by increasing background signal. POLARstar Omega Platereader (BMG Labtech)īlack vs.The VBI Equipment Resource has one additional platereader in the building. black plates is included below from the PerkinElmer website. Fluorescence readings use filter wheels with excitation and emission filters at specific wavelengths.There are many other manufacturers of these plates and so you should look for the best product to suit your needs. These are more expensive because they are sterile which may not be necessary for your application. An example of these plates are Costar white PS microtiter plates (Cat# 3917). Please be sure to use plates suitable for luminescence assays. It should be noted we do not have injectors on the Synergy. The readings are detected from the top of the plate. Luminescence readings use a Photomultiplier Tube (PMT).The Synergy HT can be used for fluorescence, luminescence and absorbance readings (see user manual at bottom). However, if you do not have a reservation and someone comes to use the instrument and they do have a reservation, then you must remove your sample and give them immediate access. The Log sheet must be filled out legibly and in full.Ī reservation is not required for this instrument. BIO-TEK Synergy HT Microtiter Platereader Location: U3202 MRB3 Reservations Optional in VU iLab websiteīilling is determined by number of scans written on the log sheet located near the instrument.
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