38
ICCS 2011
P2
Acoustic Cytometry for Rare Event Detection of
PNH Cells
Jolene A Bradford
1
, Mike Suter
2
1
Flow Cytometry Systems, Life Technologies, Eugene,
OR, USA,
2
PeaceHealth Laboratories, Springfeld,
OR, USA
Rare event analysis is an area of broad interest in
pathology. An area of recent interest is detection of
small populations of neutrophils in the blood which
lack surface expression of markers due to absence
of a glycophosphatidylinositol (GPI)-anchor protein.
In classic paroxysmal nocturnal hemoglobinuria
(PNH), a clone of hematopoeitic stem cells arises
which expands. However, small PNH-like neutrophil
populations have been detected in the blood of up
to 15% of patients who have myelodysplasia (MDS)
and 24% of patients with aplastic anemia. These
patients do not progress to classic PNH, and do not
have the clinical sequelae of hemolytic anemia or
thrombosis of PNH A recent advance in cytometry
instrumentation utilizes acoustic focusing, in which
acoustic energy is used to precisely align cells for
more sensitive detection, and sample acquisition time
can be reduced by 10-fold or more. The detection limit
with acoustic focusing cytometry is in the range of
0.1-0.01% of nucleated cells, accumulating 1 million
total events over a 5 minute interval per sample.
We present an approach for the detection of PNH-
like neutrophils using acoustic cytometry. Utilizing a
panel of fuorescent reagents including CD45, CD15,
CD16, CD24 and fuorescent aerolysin (FLAER)
surface markers, a gating strategy is employed for
high-sensitivity analysis of neutrophils using lineage
markers. Results of both normal and abnormal patient
samples are shown. Acoustic cytometry provides
the ability to detect PNH neutrophils by acquiring
statistically signifcant cells for rare event detection.
POSTER ABSTRACTS