□ Features

Mixture of six discrete bead populations having distinct fluorescent intensities:

● The lowest intensity represents the autoflorescence signal of cells not expressing the fluorescent protein

(AcGFP1 or mCherry)

● The remaining five peaks are evenly distributed over the remaining scale of the green or red fluorescence

detection channel

Usethese beads to calibrate your flow cytometer prior to analyzing cells thatexpress the AcGFP1 or mCherry fluorescent proteins. The AcGFP Flow CytometerCalibration Beads allow for easy calibration of any flow cytometer with a laserline that excites the green fluorescent proteins AcGFP1 (Aequorea coerulescens GFP) and EGFP. [TheAcGFP Flow Cytometer Calibration Beads work for both AcGFP1 and EGFP becausetheir excitation/emission spectra and brightness are almost identical.] ThemCherry Flow Cytometer Calibration Beads allow for easy calibration of any flowcytometer with a laser line that excites the red fluorescent protein, mCherry. Eachbead suspension contains six distinct populations of beads that vary in thenumber of attached AcGFP1 or mCherry molecules, which gives each population adistinct fluorescence intensity. The Molecular Equivalent of SolubleFluorophore (MESF) value for each peak was determined by correlating thefluorescence intensity of each respective bead population with the amount ofsoluble AcGFP1 or mCherry yielding the same fluorescence intensity. Convertingfluorescence intensity readings to MESF makes it possible to estimate andcompare relative expression levels between cells in the same cell population orin independent samples, and even to compare data from different instruments andexperiments. Using the Flow Cytometer Calibration Beads, you can also determinethe linearity and stability of your flow cytometer’s readouts. The lowestintensity peak represents the autofluorescence signal of cells not expressingthe green or red fluorescent protein; this allows you to measure thefluorescence detection threshold and the background noise level of your flowcytometer. The five remaining peaks are evenly distributed over the remainingscale of the fluorescence detection channel.그림 1. Flowcytometer analysis of AcGFP Flow Cytometer Calibration Beads. 20 μl of the AcGFP Flow Cytometer Calibration Bead suspension wasthorougly resuspended in 1 ml of 1x Flow Cytometer Calibration Beads DilutionBuffer. The bead suspension was then analyzed on a FACS Calibur (BD) flowcytometer using the 488 nm laser line and detecting green fluorescence in theFL-1 channel. 60,000 events were analyzed. The obtained graph shows that thebead suspension contains 6 well-distinguished populations with differentfluorescent intensities.그림2. Flow cytometer analysis of mCherry Flow Cytometer Calibration Beads. 20 μl of the mCherry Flow Cytometer Calibration Bead Suspension wasthoroughly resuspended in 1 ml of 1x Flow Cytometer Calibration Beads DilutionBuffer. The bead suspension was then analyzed on a FACS Diva (BD) flowcytometer using the 561 nm laser excitation line and detecting in the redchannel. 10,000 events were analyzed. This graph shows that the bead suspensioncontains 6 well-distinguished populations with different fluorescent intensities.

​

□ Applications

​