Flow cytometry using annexin V can detect early apoptosis in peripheral blood stem cell harvests from patients with leukaemia and lymphoma

Anthony, Robert S.; McKelvie, Nicola D.; Cunningham, Alan; Craig, J. I. O.; Rogers, Steven; Parker, A. C.

doi:10.1038/sj.bmt.1701134

Cited by 35 publications

(26 citation statements)

References 12 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Because annexin V has been reported and generally accepted to be able to detect early apoptosis, 6,[8][9][10] we have compared Syto R 16 staining with annexin V staining. Our experiments with the KB3-1 cell line confirm the finding of others [12][13][14] that annexin V may fail to detect early apoptosis although at the same time it is able to stain those cells that have permeable membranes.…”

Section: Discussionmentioning

confidence: 99%

“…One such case is the application of human CD34-positive progenitor cells for stem cell transplantation after high-dose chemotherapy. 9 We have found that the quality of CD34 + preparations strongly depends on the pre-treatment history of the transplant such as freeze-thawing. 22 Therefore, the recognition and quantification of early irreversible apoptosis not detected with conventional techniques might well contribute to clarifying the uncertainties found in literature concerning the numbers of CD34 + cells necessary for successful transplantation (Ref.…”

mentioning

confidence: 99%

“…When using annexin V, the physiological substrate for phosphatidylserine, 6,7 these changes have been claimed to represent an early phenomenon in the execution phase of apoptosis. 6,[8][9][10][11] Recently, however, it has been claimed that the annexin V technique may not detect apoptosis in all apoptosis models. [12][13][14] Other plasma membrane changes, eg in antigenic determinants, have been described, suggesting that the plasma membrane may be an early target in the cascade of apoptotic events.…”

mentioning

confidence: 99%

See 2 more Smart Citations

Large populations of non-clonogenic early apoptotic CD34-positive cells are present in frozen-thawed peripheral blood stem cell transplants

Schuurhuis

Muijen

Oberink

et al. 2001

Bone Marrow Transplant

110

View full text Add to dashboard Cite

Summary:Apoptosis is the common cell death pathway which is initiated by a variety of different stimuli. The recognition of early apoptotic events would markedly improve reliability and convenience of apoptosis assays. In the present study the vital stain Syto R 16 in combination with the permeability marker 7-amino actinomycin D, (7-AAD) has been used to identify an early stage of apoptosis, not detected with trypan blue or 7-AAD alone or with conventional apoptosis tests and not consistently and only partly detected by the early apoptosis marker annexin V. The method was established using solid tumour cell lines treated with TNF. Subsequently we applied it to determine apoptotic populations in CD34 + peripheral blood progenitor cells obtained from growth factor and/or chemotherapy mobilised patients and frozen/thawed according to standard stem cell transplantation protocols.

show abstract

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

See 1 more Smart Citation

Large populations of non-clonogenic early apoptotic CD34-positive cells are present in frozen-thawed peripheral blood stem cell transplants

Schuurhuis

Muijen

Oberink

et al. 2001

Bone Marrow Transplant

110

View full text Add to dashboard Cite

show abstract

“…Other viability dyes such as Syto16 and AnnexinV are markers of early apoptosis and may underestimate the number of viable CD34 þ cells within samples. 13,15,16 The yield of CD34 þ cells was directly compared before and after cryopreservation. The overall median recovery of viable CD34 þ cells post thaw was 93%.…”

Section: Discussionmentioning

confidence: 99%

“…Various groups have reported the recovery of viable CD34 þ cells post thawing. 1,[13][14][15][16][17] In all cases the thawed product was manipulated, either by washing 1 or diluting in PBS, 17 potentially resulting in nonspecific loss of CD34 þ cells and thus not reflecting the actual number of viable CD34 þ cells available for transplant. Ultimately, the enumeration of viable CD34 þ cells for reinfusion is likely to be a clinically important variable to predict graft success or failure.…”

mentioning

confidence: 99%

Recovery of viable CD34+ cells from cryopreserved hemopoietic progenitor cell products

Sartor

Antonenas

Garvin

et al. 2005

Bone Marrow Transplant

View full text Add to dashboard Cite

Summary:The number of CD34 þ cells infused into patients at the time of autologous or allogeneic transplantation is a clinically important variable, but the viability of these cells has not been extensively documented. In this study, we analyzed the recovery of viable CD34 þ cells before and after cryopreservation on 79 autologous stem cell products, using a novel flow cytometry assay without red cell lysis. For 70 PBSC harvest samples, the mean viable CD34 þ cell count was 5.98 Â 10 6 /kg (range 0.3-23 Â 10 6 / kg) before freezing and 5.4 Â 10 6 /kg (range 0.2-23 Â 10 6 / kg) after thawing. The median recovery was 93% (range 48-107%), with 90% recovery for NHL (range 48-100%, n ¼ 34), 83% for multiple myeloma (range 56-106%, n ¼ 11), 92.3% for acute leukemia (range 71-100% n ¼ 7) and 94.5% for nonhematological malignancies (range 50-107% n ¼ 18). Similarly, for autologous bone marrows (n ¼ 9) the median recovery of viable CD34 þ cells was 90% (range 68-100%). The recovery of viable CD34 þ cells for adult (n ¼ 51) and pediatric (n ¼ 28) stem cell collections was 91 and 94%, respectively. Further examination of the correlation between the kinetics of hematological recovery and the number of viable progenitor cells infused, particularly at the lower end of the accepted dose range, may be warranted.

show abstract

The dynamic process of apoptosis analyzed by flow cytometry using Annexin‐V/propidium iodide and a modified in situ end labeling technique

Span¹,

Pennings

Vierwinden

et al. 2001

Cytometry

View full text Add to dashboard Cite

BackgroundTo study the apoptotic process in time, we used the following flow cytometric (FCM) techniques: phosphatidylserine (PS) translocation by Annexin‐V (AnV), DNA fragmentation by in situ end labeling (ISEL), and propidium iodide (PI) staining. Because PS translocation is assumed to be an early feature of programmed cell death (PCD), we questioned if AnV positivity implies inevitable cell death.MethodsApoptosis was induced in Jurkat cells by γ‐irradiation, incubation with camptothecin (CPT), or cytosine β‐D‐arabinofuranoside (Ara‐C). At different time intervals, PCD was quantified by AnV/PI and ISEL. To analyze the influence of cell handling procedures on PCD, we applied these three FCM techniques on CD34+ bone marrow (BM) stem cells after selection and after a freeze‐thaw procedure. Various AnV/PI− CD34+ fractions were cultured in a single‐cell single‐well (SCSW) assay.ResultsJurkat cells under three different detrimental conditions showed essentially the same pattern of apoptosis in time. Initially developed AnV+/PI− cells subsequently (within 1 h) showed ISEL positivity, after which they turned into AnV+/PI++ cells with even higher levels of ISEL positivity (80–90%). Eventually, they lost some of their PI and ISEL positivity and formed the AnV+/PI+ fraction. Cell handling of CD34+ cells caused high and variable AnV+/PI− fractions (overall range 23–62%). Within total AnV+ and AnV+/PI− populations, only a minority of CD34+ cells showed ISEL positivity (range 4–8% and 0.8–6%, respectively). Different fractions of AnV+/PI− CD34+ cells did have clonogenic capacity.ConclusionsPCD of cell suspensions in vitro can be followed accurately in time by these three FCM techniques. PS translocation is followed rapidly (within 1 h) by oligo‐nucleosomal DNA fragmentation, after which cell (and nuclear) membrane leakage occurs. Detection of PS asymmetry by AnV‐fluorescein isothiocyanate (FITC) is not always associated with (inevitable) apoptosis, as can be concluded from the proliferative capacity of AnV+ /PI− CD34+ cells in the SCSW assay. Cytometry 47:24–31, 2002. © 2001 Wiley‐Liss, Inc.

show abstract

Flow cytometry using annexin V can detect early apoptosis in peripheral blood stem cell harvests from patients with leukaemia and lymphoma

Cited by 35 publications

References 12 publications

Large populations of non-clonogenic early apoptotic CD34-positive cells are present in frozen-thawed peripheral blood stem cell transplants

Large populations of non-clonogenic early apoptotic CD34-positive cells are present in frozen-thawed peripheral blood stem cell transplants

Recovery of viable CD34+ cells from cryopreserved hemopoietic progenitor cell products

The dynamic process of apoptosis analyzed by flow cytometry using Annexin‐V/propidium iodide and a modified in situ end labeling technique

Contact Info

Product

Resources

About