Fluorescence measurements were taken using a Becton Dickinson (BD) special order cytometer with a 514-nm laser excitation fluorescence that is cut off at 525 nm prior to photomultiplier tube collection (BD, Franklin Lakes, NJ). Events were annotated, subset to singlet yeast using the FlowTime R package (https://github.com/wrightrc/flowTime ). A total of 10,000 to 20,000 events above a 400,000 FSC-H threshold (to exclude debris) were collected for each sample and data exported as FCS 3.0 files for processing using the flowCore R software package and custom R scripts (65 (link), 66 (link)). Data from at least two independent replicates were combined and plotted in R (https://ggplot2.tidyverse.org/ ).
Special order cytometer
Manufactured by BD
The BD special order cytometer is a flow cytometry instrument designed for the analysis of cells and particles in liquid samples. It is capable of simultaneously measuring and analyzing multiple parameters of individual cells or particles passing through a laser beam. The core function of this cytometer is to provide quantitative data on the physical and fluorescent characteristics of the measured samples.
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Lab products found in correlation
3 protocols using special order cytometer
1
Yeast Cytometry: Fluorescence Measurement
2
Fluorescence-Based Yeast Cell Analysis
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Fluorescence measurements were taken using a Becton Dickinson (BD) special order cytometer with a 514 nm laser exciting fluorescence that is cut off at 525 nm prior to photomultiplier tube collection (BD, Franklin Lakes, NJ). Events were annotated, subset to singlet yeast using the FlowTime R package (Wright et al., 2019 ). A total of 10,000–20,000 events above a 400,000 FSC-H threshold (to exclude debris) were collected for each sample and data exported as FCS 3.0 files for processing using the flowCore R software package and custom R scripts (Supplementary file 1 ; Havens et al., 2012 (link); Pierre-Jerome et al., 2017 (link)). Data from at least two independent replicates were combined and plotted in R (ggplots2).
3
Fluorescence-Based Yeast Strain Validation
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Fluorescence measurements were taken using a Becton Dickinson (BD) special order cytometer with a 514 nm laser exciting fluorescence that is cut off at 525 nm prior to photomultiplier tube collection (BD, Franklin Lakes, NJ). For validation of R-SGA strains by flow cytometry fluorescence measurements were taken using a Sony SA3800 spectral cell analyzer with 488 nm, 405 nm & 561 nm lasers exciting fluorescence (Sony Biotechnology, San Jose, CA). Events were annotated, subset to singlet yeast using the FlowTime R package105 (link). A total of 10,000–20,000 events above a 400,000 FSC-H threshold (to exclude debris) were collected for each sample and data exported as FCS 3.0 files for processing using the flowCore R software package and custom R scripts106 (link),107 (link) (available in Github: https://github.com/achillobator/TPL-H1_Mechanism ). Data from at least two independent replicates were combined and plotted in R (ggplots2).
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