Takara AcGFP Flow Cytometer Calibration Beads \20 Assays

The AcGFP Flow Cytometer Calibration Beads allow for easy calibration of any flow cytometer with a 488 nm laser line that excites the green fluorescent proteins AcGFP1 (Aequorea coerulescensGFP) and EGFP. The excitation/emission spectrum and brightness of AcGFP are almost identical to those of EGFP. The beads consist of a mixture of six distinct populations that vary in the number of attached AcGFP1 molecules, giving each population a distinct fluorescent signature. The value for the corresponding Molecular Equivalent of Soluble Fluorophore (MESF) per peak was determined by correlating the fluorescence intensity of each respective bead population with the amount of soluble AcGFP1 yielding the same fluorescence intensity. The lowest intensity represents the autofluorescence signal of cells not expressing green fluorescent protein, while the five remaining peaks are evenly distributed over the remaining scale of the green fluorescence detection channel.

Flow cytometer analysis of AcGFP Flow Cytometer Calibration Beads. 20 µl of the AcGFP Flow Cytometer Calibration Bead suspension was thorougly resuspended in 1 mL of 1X Flow Cytometer Calibration Beads Dilution Buffer. The bead suspension was then analyzed on a FACS Calibur (BD) flow cytometer using the 488 nm laser line and detecting green fluorescence in the FL-1 channel. 60,000 events were analyzed. The obtained graph shows that the bead suspension contains six well-distinguished populations with different fluorescent intensities.