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  Cell Viability

  Cell viability can be assessed by examining parameters such as cell membrane integrity or the activity of cellular enzymes. On a microplate readers, these parameters can be detected using fluorescent reagents. For example, a red cell-impermeant, DNA-binding dye only stains dead or dying cells whose membranes are compromised, while a green live-cell dye only fluoresces when metabolic enzymes are active.

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  DNA/RNA Quantitation

  The absorbance of a DNA sample measured at 260 nm on a spectrophotometer or microplate reader can be used to calculate its concentration. Absorbance quantitation works on samples ranging from about 0.25 ug/mL to about 125 ug/mL in a microplate format. Some instrumentation enables the quantitation of very small sample volumes, as little as 2 uL. When greater sensitivity is required, fluorescence methods allow quantitation of as little as a few picograms of DNA.

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  Western Blot Detection

  Western blotting is among the most common methods employed for the detection and quantitation of specific proteins. In this method, protein sample, e.g. from a cell lysate, is first separated by size on an SDS-PAGE gel. Proteins are then transferred to a nitrocellulose or PVDF membrane, which is probed with antibody specific to the protein of interest. Various techniques are used to detect proteins on western blot membranes including fluorescence and chemiluminescence.

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  HTRF

  HTRF®, an assay technology developed by Cisbio, combines standard FRET (fluorescence resonance energy transfer) with time-resolved measurement of fluorescence, eliminating short-lived background fluorescence. HTRF’s applications include assessment of small phosphorylated peptides, immunoassays for quantifying large glycoproteins, and indirect detection of tagged complexes such as CD28/CD86 via secondary antibodies. These homogeneous assays do not require wash steps and are amenable to small-volume microplate formats used in high-throughput screening.
  (This assay is suitable for SpectraMax® iD5 and SpectraMax® Paradigm readers.)

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  Reporter Gene Assay

  Reporter gene assays enable researchers to determine whether a gene is expressed and at what level. They are used to study signaling pathways, gene regulation, structure of regulatory elements, and much more. A reporter gene vector consists of a promoter or gene of interest, along with a reporter gene such as firefly luciferase, which codes for a protein whose activity can be easily measured. The vector is transfected into mammalian cells, and when the gene of interest is active in the cells, the reporter gene is also expressed and can be measured using appropriate detection reagents.

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Getting to know your new SpectraMax iD3 Multi-Mode Microplate Reader

View a brief overview of the features and benefits of your new SpectraMax iD3 Multi-Mode Microplate Reader.
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