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Modified Annexin V/Propidium Iodide Apoptosis Assay For Accurate Assessment of Cell Death
Published on: April 24, 2011
Low propidium iodide intensity in flow cytometric white blood cell counting as a marker of cell destruction?
Thomas Wagner1, Sylvia E Guber, Maria-Luise Stubenrauch
1Department of Blood Group Serology and Transfusion Medicine, Medical University of Graz, Graz, Austria. thomas.wagner@meduni-graz.at
Background:
Residual white blood cells (WBC) in filtered blood products were investigated with flow cytometry. Frequently two distinct populations with different propidium iodide (PI) intensities can be found. The aim of this study was to specify a population with low PI intensity and discuss it as a marker of ongoing cell destruction and their possible impact on cytomegalovirus safety.
Study Design And Methods:
Buffy coat-depleted red blood cells were filtered with an in-line filtration set (LCR5, MacoPharma) after 4 hours (LCR5/4 hr) and 16 hours (LCR5/16 hr) of storage, and whole blood was filtered with a whole-blood filtration set (LST1, MacoPharma [LST1/4 hr]). The population with low PI intensity was sorted with a flow cytometer and prepared for transmission electron microscopy.
Results:
The absolute count obtained in the low-PI-intensity area before filtration was significantly different comparing LCR5/4 hr (11.5 x 10(6) +/- 6.84 x 10(6) and 0.12 x 10(6) +/- 0.1 x 10(6)/unit) and LCR5/16 hr (69.3 +/- 42.12 and 0.06 +/- 0.05; p < 0.002). By use of LST1/4 hr no difference was found compared to LCR5/4 hr after filtration (0.12 +/- 0.09 vs. 0.12 +/- 0.1), but a significant difference was found when comparing the results before filtration (1.25 +/- 0.41 vs. 11.5 +/- 6.84; p < 0.02). Electron microscopy revealed that the sorted population consisted of predominantly cell and nuclear fragments.
Conclusions:
Events found in the low-PI-intensity area are not WBCs but partially degraded DNA coming from ongoing cell destruction during extended storage. Our results provide evidence that the absolute count of events found in the low-PI-intensity area can be used as a semiquantitative marker of WBC destruction.
