Calibration Beads To Monitor FACScan Settings And Generate Compensation Profiles

March 02, 2017

Albert Einstein College of Medicine
Clinical Research
Pre-doctoral researcher

Overall

Quality of Results

Ease-of-Optimization

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Company:

BD Biosciences

Product Name:

CaliBRITE 3-color bead kit

Catalog Number:

340486

At our field site, we use a FACScan flow cytometry to quantify CD4 T-cell counts in our patient population. To ensure that the flow cytometry's performance is consistent over the course of each study, we use BD's caliBRITE bead kit. We selected the three color kit (FITC, PE, PerCP) so that we could also use the beads to generate a fluorescence compensation profile/template.

Experimental Design and Results Summary

Application

Flow cytometry.

Starting Material

Bead dilution buffer -- included in the kit.

Protocol Overview

Prepare the test samples by adding 1 drop of the appropriate bead into 1 ml of bead dilution buffer. Data can be acquired immediately -- Adjust voltage settings as necessary to optimize fluorescence. Run an experimental sample in concert with the test (bead) samples, as the optical properties for the beads will likely be different from the cell population of interest. Use the individual test (bead) samples to generate a compensation profile that can be used for future experiments.

Tips

Follow the manufacturer's suggestions EXACTLY. Optimize each protocol before running precious, human samples.

Results Summary

Results are generated following flow cytometric analysis and can be analyzed in FlowJo. We recommend single color and FMO controls to ensure the utmost accuracy and precision during each experiment.

DOI or PMID #

N/A

Additional Notes

N/A

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Summary

The Good

Simple protocol, reliable method, time efficient.

The Bad

Slightly expensive.

The Bottom Line

I highly recommend the 3-color caliBRITE bead kit for 1. quality control and evaluation of instrument performance and 2. generation of a compensation profile that can be applied to each FACS experiment. The beads are easy to use and provide an excellent quality control measure.

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