Comparison of the methods evaluating post thawing viability of peripheral blood stem cell graft

Varan, Hacer Doğan; Bay, Meltem; Ozturk, Aydin; Dalva, Klara; İlhan, Osman

doi:10.1016/j.transci.2019.03.016

Cited by 7 publications

(17 citation statements)

References 19 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Flow cytometry. CD45+, CD34+, and CD34+/7-AAD positive cells were counted by flow cytometry as a measure of viable cells [22]. Following a total nucleated-cell count, outlined above, samples were diluted to 1-2x10 7 cells ml -1 and incubated with CD45 FITC/CD34 PE antibody (BD Bioscience, Wokingham, Berkshire, UK, #341071) and 7-AAD viability dye (BD Bioscience, #559925) in BD Trucount tubes (BD Bioscience, #555899) for 15 minutes at room temperature in the dark.…”

Section: Post-thaw Analysis Of Cryopreserved Leukapheresis Samplesmentioning

confidence: 99%

Automated dry thawing of cryopreserved haematopoietic cells is not adversely influenced by cryostorage time, patient age or gender

et al. 2020

View full text Add to dashboard Cite

Cell therapies are becoming increasingly widely used, and their production and cryopreservation should take place under tightly controlled GMP conditions, with minimal batch-to-batch variation. One potential source of variation is in the thawing of cryopreserved samples, typically carried out in water baths. This study looks at an alternative, dry thawing, to minimise variability in the thawing of a cryopreserved cell therapy, and compares the cellular outcome on thaw. Factors such as storage time, patient age, and gender are considered in terms of cryopreservation and thawing outcomes. Cryopreserved leukapheresis samples from 41 donors, frozen by the same protocol and stored for up to 17 years, have been thawed using automated, water-free equipment and by conventional wet thawing using a water bath. Post-thaw viability, assessed by both trypan blue and flow cytometry, showed no significant differences between the techniques. Similarly, there was no negative effect of the duration of frozen storage, donor age at sample collection or donor gender on post-thaw viability using either thawing method. The implication of these results is that the cryopreservation protocol chosen initially remains robust and appropriate for use with a wide range of donors. The positive response of the samples to water-free thawing offers potential benefits for clinical situations by removing the subjective element inherent in water bath thawing and eliminating possible contamination issues.

show abstract

Section: Post-thaw Analysis Of Cryopreserved Leukapheresis Samplesmentioning

confidence: 99%

Automated dry thawing of cryopreserved haematopoietic cells is not adversely influenced by cryostorage time, patient age or gender

et al. 2020

View full text Add to dashboard Cite

show abstract

“…Ten microliters taken from this mixture was pipetted and placed on the counting chamber. If the cell number exceeded 1×10 7 cells/ml, the samples were diluted with saline solution at the ratio of 1:9, and the count was repeated thereafter [6].…”

Section: Cell Count Automated Cell Count Via Trypan Blue Exclusionmentioning

confidence: 99%

“…The viability of the cells is displayed on the screen in terms of percentage of viability. The 7-aminoactinomycin dye (7-AAD) method Flow cytometric analysis was performed at Ankara University Hematology Laboratory as explained in the studies of Varan et al [6] and Kilic et al [12]. Briefly, the yielded cellular products were evaluated for 7-AAD uptake during the flow cytometric count of CD34+ cells.…”

Section: Viability Automated Cell Counter Via Trypan Blue Exclusionmentioning

confidence: 99%

“…Also some manual methods-i.e. acridine orange, eosin staining can be used for viability detection [6]. Alternatively, the viability dye 7aminoactinomycin D (7-AAD) can also be used to determine the number of viable CD34+ cells [7,8].…”

Section: Introductionmentioning

confidence: 99%

“…In this manner, automated techniques present themselves to be utile in performing both cell count and viability estimation, and when necessary being able to give both results realtime. Previously, researchers made various comparisons of manual and automated cell count and viability techniques in different cell types [2,5,6,[9][10][11]. However, none of these studies show comparison between automatic and/or manual cell count and viability techniques by means of hematopoietic progenitor cells.…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Utility investigation of automated techniques in hematopoietic progenitor cell count and viability assessment in the Good Manufacturing Practice (GMP) setting

Kilic

Bay

Baydin

et al. 2020

j-ebr

Self Cite

View full text Add to dashboard Cite

To compare our parameters as regards: i) cell count via two different automated cell count techniques, and ii) viability via automated trypan blue exclusion and 7-aminoactinomycin D (7-AAD) staining. Method: We used the trypan blue exclusion technique and an automated cell counter and for viability testing, and the trypan blue exclusion technique and the 7-AAD evaluation by flow cytometry. The trypan blue exclusion and the radio frequency techniques were used for automated cell counting. Flow cytometric analysis was performed by evaluating the yielded cellular products for 7-AAD uptake during the cell count of CD34+ cells. Results: The mean values for cell count were estimated as 3.44±1.22x10 6 /ml (range, 2.48-5.71x10 6 /ml) and 4.14±1.94x10 6 /ml (range, 1.77-7.43x106/ml) for the trypan blue exclusion and radio frequency techniques, respectively. Additionally, the mean values for viability analyses via the automated trypan blue exclusion and 7-AAD were 93.38±6.09% (range, 79.00-98.00%) and 99.49±0.60% (range, 98.40-100.00%), respectively. Conclusions: Our study has responded to two fundamental questions: whether the results of both of the automated techniques for cell count correspond with each other, and whether the results of the automated viability assessment conform those of the 7-AAD technique during the manufacturing processes of cellular therapy products intended for clinical use. Even though we have the opportunity to use the hemocytometer in our laboratory setting, the automated trypan blue exclusion technique gives cell count results in concordance within the range of the expectations of our Quality Management System (QMS).

show abstract

Variable recovery of cryopreserved hematopoietic stem cells and leukocyte subpopulations in leukapheresis products

Rimac

Bojanić

Dabelić

et al. 2023

Transfusion and Apheresis Science

View full text Add to dashboard Cite

Comparison of the methods evaluating post thawing viability of peripheral blood stem cell graft

Cited by 7 publications

References 19 publications

Automated dry thawing of cryopreserved haematopoietic cells is not adversely influenced by cryostorage time, patient age or gender

Automated dry thawing of cryopreserved haematopoietic cells is not adversely influenced by cryostorage time, patient age or gender

Utility investigation of automated techniques in hematopoietic progenitor cell count and viability assessment in the Good Manufacturing Practice (GMP) setting

Variable recovery of cryopreserved hematopoietic stem cells and leukocyte subpopulations in leukapheresis products

Contact Info

Product

Resources

About