Abstract
The National Institute of Standards and Technology (NIST), the National Institutes of Health (NIH) and other industry stakeholders have been working together to enable fluorescence intensities of flow cytometer calibration beads to be assigned quantitative equivalent reference fluorophore (ERF) values with high accuracy and precision. The ultimate goal of this effort is to accurately quantify the number of antibodies bound to individual living cells. The expansion of this effort to assign ERF values to more than 50 fluorescence channels and particles with diameters ranging from 10 μm down to 80 nm is reported here.
Highlights
A Flow Cytometry Quantitation Consortium has been formed between calibration microsphere/nanosphere manufacturers, National Institutes of Health (NIH), National Institute of Standards and Technology (NIST) and other industry stakeholders [1]to enable fluorescence intensity value assignments of cytometer calibration beads in units of equivalent reference fluorophores (ERF) [2]
ERF-based fluorescence intensities of micrometer-sized calibration beads is improving the accuracy of calibration for Flow cytometry (FCM) fluorescence channels with emission centered from 390 to 850 nm
The purity of reference fluorophores and the concentration of corresponding reference solutions used for ERF assignments has been determined with known uncertainties
Summary
To enable fluorescence intensity value assignments of cytometer calibration beads in units of equivalent reference fluorophores (ERF) [2]. The ultimate goal of ERF value assignments is the quantitation of the number of antibodies bound per cell (ABC) for a particular target antigen of interest, such as CD4 or CD19. Fluorophore solutions of known concentration, such as Standard Reference Material (SRM) 1934 [6], are used to assign fluorescence intensities to the beads in units of ERF. This is done by measuring the fluorescence intensity of both the Materials 2020, 13, 4111; doi:10.3390/ma13184111 www.mdpi.com/journal/materials
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