Comparison of two single-platform ISHAGE-based CD34 enumeration protocols on BD FACSCalibur and FACSCanto flow cytometers.
OBJECTIVE
Enumeration of viable CD34(+) cells provides critical information for the bone marrow (BM) transplant physician. The single-platform ISHAGE protocol is the most reliable method currently available to quantitate accurately this important subset of cells. Previous studies have shown that 5 CD34(+) cells/microL blood predicts the collection of at least 0.5x10(6) CD34(+) cells/kg patient weight. From the apheresis product, infusion of 2.5x10(6) viable CD34(+) cells (measured pre-cryopreservation)/kg patient weight will reliably permit engraftment of the hematopoietic system (as measured by the time to 20000 platelets/microL) by day 12-14 post-infusion.


METHODS
We compared the CD34(+) cell numbers derived from Flow Count-based Stem-Kit; (Beckman Coulter) and Trucount tube-based stem cell enumeration (SCE) kit (BD Biosciences) ISHAGE templates on BD FACSCalibur and BD FACSCanto cytometers on 12 granulocyte-colony-stimulating factor (G-CSF)-mobilized peripheral blood (MPB) and 10 peripheral blood stem cell (PBSC) samples.


RESULTS
Comparison of results showed that there was no statistical difference between samples run with Stem-Kit on the FACSCalibur versus SCE kit-based assays on either the FACSCalibur or FACSCanto. Mean results for the Stem-Kit/Calibur combination were 137, for SCE kit/Calibur 140 and for SCE kit/Canto 137 cells/microL. Pair-wise comparison of data based on rank order showed no statistically significant difference and all correlation coefficients had an R(2)>0.98.


CONCLUSIONS
The two kits generated very similar data on a range of fresh samples regardless of instrument platform. These results confirm and extend the utility of the single-platform ISHAGE protocols with a variety of reagent kits and instrument platforms.
Sutherland
D Robert
DR
Clinical Flow Cytometry Laboratory, University Health Network, Toronto, Canada. rob.sutherland@utoronto.ca
Nayyar
Rakesh
R
Acton
Erica
E
Giftakis
Angeline
A
Dean
Sue
S
Mosiman
Victoria L
VL
D018952:Antigens, CD34 / Q000378:metabolism*; D003114:Colony-Forming Units Assay; D005434:Flow Cytometry / Q000295:instrumentation* / Q000379:methods*; D006801:Humans; D011933:Reagent Kits, Diagnostic; D013234:Stem Cells / Q000166:cytology / Q000378:metabolism
D018952:Antigens, CD34; D011933:Reagent Kits, Diagnostic