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42
ICCS 2012
cytometry. We identified specific signatures in Alzheimer
patients compared to age-matched non-Alzheimer patients.
The frequency of the T cell subsets identified correlated
to circulating levels of inflammatory mediators. Diabetic
and frail elderly also display distinct profiles assessed
by flow cytometry. The role of chronic infections such as
cytomegalovirus in such events will be discussed.
P22
FREQUENCY OF COLOR COMPENSATION IN CLINICAL
USE
Kim Le, Kim Romang, Melissa Cessna
Intermountain Central Laboratory, Flow Cytometry
Departmant, Intermountain Healthcare
Introduction:
Color compensation is essential in multicolor
flow cytometry. The College of American Pathologists
flow cytometry checklist requires having procedures for
determining color compensation settings, but does not
specify how often the color compensation must be done.
The color compensation procedure can be costly and
time consuming especially if a cytometer uses multiple
fluorochromes and some of them are tandem. Tandem
fluorochromes are sensitive to increased light exposure
and elevated temperature; and therefore require antibody-
specific compensation. We evaluated the stability of spectral
overlaps using the BD FACSCanto II instrument and its
compensation software and determined the frequency
of color compensation performances needed. The BD
FACSCanto II has three lasers and can detect up to
eight fluorochromes.
Materials and Methods:
FITC, PE,
PerCP-Cy5.5, PE-Cy7, APC, APC-H7, V450, and V500
fluorochromes were used for color compensation. PE-Cy7
and APC-H7 were tandem and required eight antibody-
specific compensation tubes. Voltages and spectral overlaps
were computed for mean and standard deviation (SD).
Results:
This study was done from April 28 to May 23, 2012.
All spectral overlaps had small SD less than 2%, even with
tandem fluorochromes. The voltages of all photomultiplier
detectors were very stable with SD less than 4 volts.
Conclusions:
Color compensations remained stable over
time. Daily color compensation performance on the BD
FACSCanto II was not needed due to its daily performance
check to ensure the stability of photomultiplier voltages. We
suggested doing color compensation weekly instead of daily
(per manufacturer’s recommendation) to reduce reagent cost
and processing time.
P23
VALIDATION OF EOSIN 5 MALEIMIDE STAINING FOR
THE DETECTION OF SPHEROCYTES IN HEREDITARY
SPHEROCYTOSIS
Michael A Liew
1
, Roberto Nussenzveig
1
, Archana Agarwal
1,2
,
Carl T Wittwer
1,2
1
ARUP Institute for Clinical & Experimental Pathology,
2
University of Utah School of Medicine
Hereditary spherocytosis (HS) is a common inherited
hemolytic anemia characterized by the presence of spherical
erythrocytes or spherocytes. Spherocytes have a smaller
surface area and can maintain healthy oxygen levels, but
are physically fragile. HS can be diagnosed by clinical
features, peripheral smear examination as well as clinical
laboratory tests including osmotic fragility, ektacytometry or
flow cytometry. Our aim was to validate the flow cytometry
assay using eosin 5 maleimide (EMA) labeled intact red
blood cells that bind to band 3. One microliter of whole
blood was washed in 200μl of phosphate buffered saline
(PBS), and stained using 0.5mg/ml EMA for 60mins at
room temperature. The sample was then washed 3 times
POSTER ABSTRACTS