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Analysis of factors associated with successful allogeneic peripheral blood stem cell collection in healthy donors
Transfusion and Apheresis Science xxx (xxxx) xxxx
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Analysis of factors associated with successful allogeneic peripheral blood stem cell collection in healthy donors Jung Hee Konga,1, Youmi Hua,1, Hyoeun Shima, Eunyoung Leeb, Hyewon Leeb, Hyeon-Seok Eomb, Se-Na Leea,*, Sun-Young Konga,* a b
Department of Laboratory Medicine, National Cancer Center, Goyang, South Korea Center for Hematologic Malignancy, National Cancer Center, Goyang, South Korea
A R T I C LE I N FO
A B S T R A C T
Keywords: Allogeneic Peripheral blood stem cells Apheresis
Background: The collection of a sufficient number of stem cells is important for success of allogeneic hematopoietic stem cell transplantation (HSCT). This study aimed to investigate the factors associated with successful allogeneic peripheral stem cell (PBSC) collection in healthy donors. Methods: We retrospectively reviewed clinical data of allogeneic PBSC collection in 175 donors from 2007 to 2017 at the National Cancer Center, Korea. This study analyzed factors associated with the CD34+ cell yield such as the characteristics of donors, including age, laboratory results before apheresis, and data of procedures on the first day. The CD34+ cell dose of ≥ 4.0 × 106/kg have recently been the accepted minimum recommended dose in allogeneic HSCT settings, and this was the target dose in our study. Results: The factors associated with the CD34+ cell yield were age (p = 0.007), baseline platelet (PLT) (p = 0.014), and pre-collection hematopoietic progenitor cells (HPCs) (p = 0.001) by multivariate analysis. This study represented that age, baseline platelet count, and pre-collection HPC count are important predictive factors as shown in other previous studies. Conclusion: Our data suggest that young age, high baseline platelet counts and high HPC counts before collection might be useful for identifying successful mobilizers.
1. Introduction Allogeneic hematopoietic stem cell transplantation (HSCT) is an established procedure for many hematologic malignancies [1]. Peripheral blood stem cells (PBSCs) are increasingly used as the source of stem cells in HSCT because of improved neutrophil and platelet (PLT) engraftment, avoidance of general anesthesia for collection, and potentially better graft-versus-leukemia effects [2]. Factors associated with successful allogeneic peripheral blood stem cell transplantation (PBSCT) include age of the patient, disease stage, time interval from diagnosis to transplant, human leukocyte antigen (HLA) and ABO compatibility between donor and recipient, and number of harvested stem cells [3–5]. Especially, obtaining a sufficient number of stem cells using as few leukapheresis procedures as possible is important in regard to donor convenience, therapeutic results, and cost [6]. In allogeneic PBSCT, the optimal number of CD34+ cells for transplantation has not been definitively established, but a higher dose is often preferred at 5.0 × 106 CD34+ cells per kilogram (kg) of body
weight [7]. It is widely accepted that the minimum cell doses for autologous and allogeneic transplantation are 2.0 × 106 and 3.0 × 106 to 5.0 × 106 of CD34+ cells per kg, respectively [8]. However, ≥4.0 × 106 cells per kg have recently been the accepted minimum recommended dose [9,10]. In addition, a CD34+ cell dose of more than 4.0 × 106 per kg of body weight resulted in more rapid engraftment, lower transplant-related mortality, and better overall survival [11]. Thus, the target CD34+ cell dose was set at ≥4.0 × 106/kg in this study. Predicting the successful mobilization of CD34+ cells before the PBSC collection is preferable for the benefit of the donors and recipients. There are several factors reported to affect the collection of allogeneic PBSCs including sex, age, and method of mobilization, the number of peripheral white blood cells (WBCs), CD34+ cells, platelets count, and circulating immature cells [12–16]. Especially, a circulating CD34+ cell count before apheresis was widely used as an indicator for predicting the PBSC harvest yield, but is expensive and needs specific equipment and experienced staff. Hematopoietic progenitor cells
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Corresponding authors at: Department of Laboratory Medicine, National Cancer Center, 323 Ilsan-ro, Ilsandong-gu, Goyang-si Gyeonggi-do 10408, South Korea. E-mail addresses: [email protected] (S.-N. Lee), [email protected] (S.-Y. Kong). 1 Jung Hee Kong and Youmi Hu equally contributed to this work. https://doi.org/10.1016/j.transci.2019.102679 Received 18 September 2019; Received in revised form 7 November 2019; Accepted 11 November 2019 1473-0502/ © 2019 Elsevier Ltd. All rights reserved.
Please cite this article as: Jung Hee Kong, et al., Transfusion and Apheresis Science, https://doi.org/10.1016/j.transci.2019.102679
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(HPCs) quantification is rapid, inexpensive and requires minimal training [17]. Here, we retrospectively analyzed the yield of CD34+ cells from 175 healthy donors. The aim of this study was to investigate the factors influencing successful allogeneic PBSC collection of CD34+ cells.
Table 1 Characteristics of donors and variables of apheresis.
2. Materials and methods 2.1. Donor characteristics and variables of apheresis Two hundred thirty-four procedures were performed on 175 healthy donors and 59 donors required a second procedure between February 2007 and December 2017. This study was approved by the ethical review boards at the authors’ institution (NCC2016-0013). In our study, only the data of the first day were analyzed. The characteristics of the donors include sex, age, body weight (BW), and routes of vascular access (peripheral vein or central venous catheter). Blood parameters include counts of WBCs, mononuclear cells (MNCs) and HPCs measured before and after the procedure using XE-2100 automatic analyzer (Sysmex, Kobe, Japan). Enumeration of the HPC was performed in the immature myeloid information (IMI) channel, which identifies a population of hematopoietic progenitor cells based on cell size, complexity and differential cell lysis characteristics. The CD34+ cell count was measured by two-color flow cytometry using FACS Canto II (Becton, Dickinson and Company, San Jose, CA, USA) after immunofluorescence staining using anti-human CD34 monoclonal phycoerythrin-conjugated (PE) antibodies and CD45 monoclonal fluorescein isothiocyanate-conjugated antibodies (FITC) (Becton–Dickinson, New Jersey, USA). The number of CD34+ cells per BW in product was calculated by donor’s BW. We calculated the collection efficiency (CE) by using MNCs instead of CD34+ cells as the pre-collection CD34+ cell counts were not performed. The CE was estimated as described previously as [MNCs obtained from apheresis / MNCs (μL) × Apheresis volume processed (μL)] [6].
Variable
Median(range)
Number of donors Sex(male/female) Age (years) Donor-recipient relation Related Unrelated Donor BW (kg) Vascular access, N (%) Pheripheral vein Central venous catheter Baseline-PBSC collection WBC count (×109/L) Hct (%) PLT (×109/L) MNC (%) Pre- PBSC collection WBC count (×109/L) HPC (×106/L) Hct (%) PLT (×109/L) MNC (%) Post- PBSC collection WBC count (×109/L) Hct (%) PLT (×109/L) MNC (%)
175 107(61 %)/68(39 %) 35
(14–69)
103 72 69.2
(59 %) (41 %) (44.2–115.4)
134 41
(77 %) (23 %)
5.98 43.7 244 41
(2.98–13.15) (31.2–53.2) (164–460) (24–62)
44.78 60 40.9 196 19
(22.75–80.71) (2–296) (28.2–50.1) (99–339) (10–34)
34.68 41.3 85 11
(11.44–73.83) (29.4–48.9) (33–225) (3–36)
Abbreviations: SD, standard deviation; BW, body weight; WBC, white blood cell; MNC, mononuclear cells; HPC, hematopoietic progenitor cells; Hct, hematocrit; PLT, platelet. Table 2 Summary of procedures and collected products. Variable
Median(range)
Number of procedures 175 TBV (mL) 4557 TPV (mL) 17,000 Ratio of TPV to TBV 3.8 Volume of product (mL) 312 PBSC product blood parameters of collection Total WBCs (×108/kg) 9.73 Total MNCs (×108/kg) 8.00 Total CD34+ cells (/uL) 1113 Total CD34+ cells (×106/kg) 4.60 MNC efficiency (%)* 40.8 Platelet loss (%)† 56.3 Instrument, N (%) COBE Spectra 120 Spectra Optia 55 MNC collection efficiency according to instrument (%)* COBE Spectra 40.2 Spectra Optia 42.6
2.2. Mobilization and collection of allogeneic PBSC To mobilize the PBSCs, granulocyte-colony stimulating factor (GCSF) was administered subcutaneously once daily for 4 days to all donors at a dose of 10 mcg/kg. PBSC collection was initiated on the 5th day after mobilization. Stem cells were collected using either a central vein access via a perm catheter or peripheral vein access via 16 -18 gauge catheters. We used two different collecting machines. COBE Spectra (Terumo BCT, Inc.) was operated according to the manufacturer’s standard operating procedure based on the color scale 2–3% and Spectra Optia (Terumo BCT, Inc.), which has electronic features including optical sensors for continuous automatic monitoring. During the PBSC procedure, 3 to 4 times of the donor’s total blood volume was processed at a rate of 50 to 70 mL/min. The whole blood-to-anticoagulant (3000 IU heparin with 500 mL of acid-citrate-dextrose solution) ratio was 24:1 in both devices. The study investigated the number of procedures, total blood volume (TBV) of donors, total processed volume (TPV), TPV-toTBV ratio, and product volumes. We divided the donors into two groups according to the collected CD34+ cells in collected product on the first procedure: CD34+ in product < 4.0 × 106/kg and ≥ 4.0 × 106/kg.
(3010-6388) (9135-23,449) (2.1–6.5) (80–450) (2.10–22.30) (1.85–18.28) (156-4,404) (0.47–17.08) (13.9–96.9) (-9.2–77.7) (69 %) (31 %) (19.4–96.9) (13.9–73.2)
Abbreviations: TBV, total blood volume; TPV, total processed volume; PBSC, peripheral blood stem cell; WBC, white blood cell; MNC, mononuclear cells; CD34, Cluster of Differentiation antigen; *MNC efficiency % = MNCs obtained from apheresis / MNCs (μL) × apheresis volume processed(μL). † PLT loss % = (platelets/μL before apheresis – platelets/μL after apheresis) / platelets/ μL before apheresis.
univariate analysis. A multivariate backward stepwise selection regression analysis was used to derive a formula for predicting the cell yield. To predict successful mobilization, cut-off value of baseline platelet counts and pre-collection HPC counts were analyzed by receiver operating characteristic (ROC) analysis.
2.3. Statistics All statistical analyses were performed by using Stata Statistical Software Release 12 (StataCorp LP, College Station, TX, USA). The data was presented as the median with a range. A p value < 0.05 was considered statistically significant. Continuous variables were compared using the Kruskal-Wallis test, and categorical variables were analyzed using the Fisher’s exact test. The binary logistic regression model was used to analyze parameters found to be statistically significant at 2
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Table 3 Comparison between two groups according to the CD34+ cell yield.
Sex (male/female) Age (years) Donor BW (kg) Vascular access type, N (%) Peripheral vein Central venous catheter Instrument, N (%) COBE spectra Spectra Optia Apheresis variables TBV (mL) TPV (mL) Ratio of TPV to TBV Volume of product (mL) COBE Spectra Spectra Optia Baseline-PBSC collection WBC count (×109/L) Hct (%) PLT (×109/L) MNC (%) Pre-PBSC collection WBC count (×109/L) HPC (×106/L) Hct (%) PLT (×109/L) MNC (%) Post-PBSC collection WBC count (×109/L) Hct (%) PLT (×109/L) MNC (%) PBSC product Total WBCs (×108/kg) Total MNCs (×108/kg) Total CD34＋cell (/μL) Platelet loss (%)†
＜4.0 × 106 cells/kg
≥4.0 × 106 cells/kg
(N = 68, 39 %)
(N = 107, 61 %)
p value
38(56 %)/30(44 %) 40 67.9
(21–63) (47.7–115.4)
69(64 %)/38(36 %) 34 69.6
(14–69) (44.2–108.3)
0.255 0.005* 0.667 0.261
49 19
(72 %) (28 %)
85 22
(79 %) (21 %)
50 18
(74 %) (26 %)
70 37
(65 %) (35 %)
4344 18,000 3.8
(3,030-6388) (9135-23,449) (2.1–6.1)
4590 16,500 3.9
(3,010-6,330) (10,129-20,256) (2.1–6.5)
289 292
(170–364) (188–400)
311 342
(80–396) (198–450)
5.82 43.7 233 42
(2.98–11.82) (32.7–50.1) (164–359) (26–62)
6.11 43.7 249 40
(3.54–13.15) (31.2–53.2) (167–460) (24–55)
0.022* 0.895 < 0.001* 0.051
42.80 41 40.6 189 19
(23.01–67.14) (2–249) (28.2–48.4) (99–186) (10–33)
47.18 78 41.7 206 20
(22.75–80.71) (11–296) (31.3–50.1) (117–339) (10–34)
0.007* < 0.001* 0.741 0.006* 0.431
32.00 40.7 79 11
(11.44–54.30) (30.1–48.4) (33–186) (6–24)
37.27 41.7 89 12
(19.95–73.83) (29.4–48.9) (48–255) (3–36)
0.002* 0.382 0.003* 0.718
8.64 7.30 606 56.8
(2.10–16.44) (1.85–14.63) (156-1,524) (18.1–77.7)
10.66 8.55 1561 55.5
(5.26–22.30) (4.11–18.28) (582-4,404) (-9.2–76.7)
< 0.001* < 0.001* < 0.001* 0.690
0.260
0.465 0.384 0.292 0.030*
Abbreviations: CD34, Cluster of Differentiation antigen; BW, body weight; TBV, total blood volume; TPV, total processed volume; WBC, white blood cell; Hct, hematocrit; PLT, platelet; MNC, mononuclear cells; HPC, hematopoietic progenitor cells. * p < 0.05 statistically significant. † PLT loss % = (platelets/μL before apheresis – platelets/μL after apheresis) / platelets/μL before apheresis ×100.
median collected CD34+ cell count was 4.60 × 106/kg (0.47–17.08 × 106/kg). Furthermore, 120 (69%) and 55 (31%) procedures were performed on the COBE Spectra and Spectra Optia. The median MNC collection efficiency by the COBE Spectra and Spectra Optia were 40.2 % (19.4–96.9 %) and 42.6 % (13.9–73.2 %) (Table 2).
3. Results 3.1. Demographic details of the allogeneic donors and pre and post collection parameters of peripheral blood counts Among the 175 donors (107 male and 68 female), 103 (59 %) were related donors, and 72 (41 %) were unrelated donors. The median age and donor’s BW were 35 years (14–69 years) and 69.2 kg (44.2–115.4 kg), respectively. Venous access was peripheral vein in 134 (77 %) donors and central venous in 41 (23 %) donors. The median baseline WBC and PLT were 5.98 × 109/L (2.98–13.15 × 109/L) and 244 × 109/L (164–460 × 109/L), respectively. The median pre-collection WBC, PLT, and HPC counts were 44.78 × 109/L (22.75–80.71 × 109/L), 196 × 109/L (99–339 × 109/L), and 6 6 60 × 10 /L (2–296 × 10 /L). The median post-collection WBC and PLT counts were 34.68 × 109/L (11.44–73.83 × 109/L) and 85 × 109/l (33–225 × 109/L), respectively (Table1).
3.3. Comparison between the poor and good mobilizer groups We divided the donors into two groups according to the collected CD34+ cells in the product of the first procedure. We defined the poor mobilizer as < 4.0 × 106/kg and the good mobilizer as ≥ 4.0 × 106/kg CD34+ cells in product. 107 (61 %) donors reached a CD34+cell count ≥ 4.0 × 106/kg, and only 68 (39 %) did not. There were significant differences between the two groups for age (p = 0.005), volume of product (p = 0.030), baseline-collection WBC (p = 0.022), PLT (p < 0.001), pre-collection WBC (p = 0.007), HPC (p < 0.001), PLT (p = 0.006), post-collection WBC (p = 0.002), PLT (p = 0.003) and total WBCs (p < 0.001), MNCs (p < 0.001), CD34+cells (p < 0.001) in product. The median age of the group achieving a CD34+cell count of ≥ 4.0 × 106/kg was lower than that of the group achieving a CD34+cell count of < 4.0 × 106/kg. The count of baseline-collection PLT and pre-collection HPC in the group ≥ 4.0 × 106/kg was higher than that of the group < 4.0 × 106/kg (Table 3).
3.2. Results of PBSC collection and apheresis products A total of 175 PBSC collections were conducted during the study period. The median total blood volume (TBV) was 4557mL (30106388 mL). The median total processed volume (TPV) was 17,000 mL (9135–23,449 mL), and the TPV-to-TBV ratio was 3.8 (2.1–6.5). The 3
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Table 4 Univariate and multivariate analysis for predictive factors of optimal PBSC collection in donors. Odds ratio Sex Male Female Age (per 1 year increase) Donor BW (per 1 kg increase) Vascular access type Peripheral vein Central venous catheter Instrument COBE spectra Spectra Optia Apheresis variables TBV (mL) TPV (mL) Ratio of TPV to TBV Baseline-PBSC collection WBC count (×109/L) Hct (%) PLT (×109/L) MNC (%) Pre-PBSC collection WBC count (×109/L) HPC (×106/L) Hct (%) PLT (×109/L) MNC (%)
95 % CI
p value
1 0.698 0.961 1.003
0.375-1.298 0.936-0.987 0.982-1.024
0.256 0.003* 0.787
1 0.667
0.329-1.354
0.263
1 1.468
0.751-2.869
0.261
1.000 0.999 0.829
0.999-1.000 0.999-1.000 0.585-1.174
0.503 0.504 0.291
1.000 1.001 1.014 0.96
1.000-1.001 0.930-1.077 1.006-1.021 0.922-1.001
0.015* 0.982 < 0.001* 0.053
1.000 1.013 1.021 1.011 1.026
1.000-1.001 1.006-1.020 0.945-1.103 1.003-1.018 0.962-1.095
0.006* < 0.001* 0.595 0.005* 0.429
Odds ratio
95 % CI
p value
0.961
0.934-0.989
0.007*
1.010
1.002-1.018
0.014*
1.012
1.005-1.019
< 0.001*
Abbreviations: PBSC, peripheral blood stem cell; BW, body weight; TBV, total blood volume; TPV, total processed volume; WBC, white blood cell; Hct, hematocrit; PLT, platelets; MNC, mononuclear cells; HPC, hematopoietic progenitor cells. * p < 0.05 statistically significant.
Fig. 1. ROC curve of baseline PLT and pre-collection HPC count for predicting successful PBSC collection (4.0 × 106/kg CD34+ cells) in donor. (A) The cutoff value of the baseline PLT count was 210 × 109/L (sensitivity = 91.0 %, specificity = 76.6 %, AUC = 0.665). (B) The cutoff value of the pre-collection HPC count was 25 × 106/L (sensitivity = 91.0 %, specificity = 73.8 %, AUC = 0.667). Abbreviations: ROC receiver operating characteristic; PLT platelet; HPC hematopoietic progenitor cells; PBSC peripheral blood stem cell; AUC area under the curve
1.010, p = 0.014), and pre-collection HPC (OR 1.012, p < 0.001) (Table 4). The cutoff value of the baseline platelet count for predicting successful PBSC collection of 4.0 × 106/kg CD34+ cells was 210 × 109/L with 91.0 % sensitivity and 76.6 % specificity in ROC analysis. The cutoff value of the pre-collection HPC count for the same amount of CD34+ cells was 25 × 106/L with 91.0 % sensitivity and 73.8 % specificity. Area under the curve (AUC) of baseline PLT and precollection HPC count were 0.665 and 0.667, respectively (Fig. 1).
3.4. Predictive value of successful PBSC collection In univariate analysis, factors associated with a sufficient amount of stem cells were age [odds ratio (OR) 0.961, p = 0.003], baseline-collection WBC (OR 1.000, p = 0.015), PLT (OR 1.014, p < 0.001) and Pre-collection WBC (OR 1.000, p = 0.006), HPC (OR 1.013, p < 0.001), PLT (OR 1.011, p = 0.005). In multivariate backward stepwise selection regression analysis, significant predicting factors were donor’s age (OR 0.961, p = 0.007), baseline-collection PLT (OR 4
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4. Discussion
But, it was not a significant influential factor. It is known that the mobilization method is the most important factor influencing the allogeneic hematopoietic stem cell collection in healthy donors [18]. In particular, studies were conducted on the types of mobilization agent, doses, and duration of administration. In most published prospective and retrospective studies, G-CSF was administered to healthy donors for 4–5 days. The Italian Bone Marrow Donor Registry (IBMDR) recommends unrelated donors to limit the administration of G-CSF to a maximum of 5 days [10]. In several studies, PBSC mobilization using G-CSF alone showed in a higher CD34+ cell yield if the dose of G-CSF was applied 5mcg/kg twice daily rather than 10mcg/ kg once daily [28,29]. However, previous study reported that there were no significant differences in G-CSF induced mobilization of healthy donors between daily single and divided dose of G-CSF [30]. In our study, 10 mcg/kg/day of G-CSF was administered for 4 days, and the collection was initiated on the 5th day. More than 4.0 × 106/kg of the CD34+ cells were collected in 61 % (107/175) of the donors during the first leukapheresis. Recently, a study has reported that more than 4.0 × 106/kg CD34+ cells were collected from 62 % (31/50) of the donors in the first procedure after administration of G-CSF 10 mcg/kg/ day [22]. In conclusion, we found that the donor’s age and platelet count before mobilization were important factors for allogeneic PBSC collection in healthy donors. Furthermore, pre- collection HPCs were a significant factor that correlated with the CD34+ cell yield. Thus, young age and high baseline platelet counts and high HPC counts before collection might be useful for identifying successful mobilizers.
Several studies have also explored predictive factors for CD34+ cell yield. The number of WBCs, platelet count, CD34+ cells and circulating immature cells before and during apheresis have been reported to be factors [6,8,14,16,18]. Suzuya et al. reported that the factors associated with the CD34+ cell yield included the platelet count before and during mobilization [18]. Furthermore, Lysák et al. reported that a significant correlation was observed between CD34+ yield and donor platelet count measured before mobilization [14]. In this study, the platelet count before mobilization was observed to be a factor influencing the CD34+ cell yield. The CD34+ cell count on the day of apheresis is a powerful predictor [5]. However, the circulating CD34+ cell count before apheresis was widely used as an indicator for predicting the PBSC collection yield but was both time consuming and expensive. HPCs represent a specific population of immature myeloid cells, and can be measured quickly, inexpensively, and can be performed as part of a routine complete blood count study with minimal extra cost by automated hematology analyzer [8]. Therefore, a number of studies have been conducted to evaluate HPCs as a predictor of CD34+ cell yield. Many studies that investigated autologous PBSC collection demonstrated that there was an acceptable correlation between pre-collection HPC and CD34+ cell counts, as well as CD34+ cell yields after PBSC collection [19,20]. In our study, pre-collection HPC count was analyzed to be an important factor associated with the CD34+ cell yield. Also the cutoff value of pre-collection HPC count for predicting successful PBSC collection was 25 × 106/L. Yang et al. suggests that a preharvest HPC count of more than 21.3 × 106/L may predict adequate CD34+ (4.0 × 106/kg) cell yield [8]. In this study, the cutoff value of pre-collection HPC count was similar to the previous study. Namba et al. reported that significant factors for predicting successful collection included the sex of the donor [5]. Wang et al. noted that females had a lower mean collected outcome and a higher risk for poor yield than males [21]. However, the sex of donor was considered to have no effect on stem cell yield in many studies [11,14,22]. In our study, there was not a significant difference in donor sex among the two groups according to the number of CD34+ cells in the collected products. Age was reported to be an important factor in stem cell collection [11,14,18]. Pornprasertsud et al. reported that successful CD34+ cell counts of 5.0 × 106/kg or more were available from donors younger than 30 years of age [22]. Anderlini et al. reported that a donor age greater than 55 years was associated with a 3.8-fold greater risk for poor mobilization. There was a study that the donor’s age, with a threshold of 38 years or more, was a factor that significantly affected CD34+ cell mobilization and collection in healthy donors [11,16]. In this study, multiple regression analysis showed that age was significantly associated with the CD34+ cell yield. It has been reported that there is no significant difference between the peripheral vein and the central venous catheter in the donor’s venous access type [14,23]. Wang et al. (2008) reported that female donors had higher CD34+ cell yields if the circulation access was through an artery than if it was through a vein [21]. At our institution, central venous catheters are exclusively applied for collecting PBSCs from healthy donors with poor peripheral venous access. Furthermore, there have been no cases that used arterial access. There was not a significant difference between the two groups according to the type of venous access. Although there is a difference in total processing blood volume, blood flow rate and equipment, it has been previously shown that that there is no difference in CD34+ cell counts in the collected products [24–26]. Large-volume leukapheresis (LVL) is defined as processing 3 or up to 6 times the donor’s total blood volume. LVL has been suggested as a method of choice in donors and patients with a weak effect of mobilization [27]. In our study, the median TPV-to-TBV ratio was 3.6.
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Transfusion and Apheresis Science journal homepage: www.elsevier.com/locate/transci
Analysis of factors associated with successful allogeneic peripheral blood stem cell collection in healthy donors Jung Hee Konga,1, Youmi Hua,1, Hyoeun Shima, Eunyoung Leeb, Hyewon Leeb, Hyeon-Seok Eomb, Se-Na Leea,*, Sun-Young Konga,* a b
Department of Laboratory Medicine, National Cancer Center, Goyang, South Korea Center for Hematologic Malignancy, National Cancer Center, Goyang, South Korea
A R T I C LE I N FO
A B S T R A C T
Keywords: Allogeneic Peripheral blood stem cells Apheresis
Background: The collection of a sufficient number of stem cells is important for success of allogeneic hematopoietic stem cell transplantation (HSCT). This study aimed to investigate the factors associated with successful allogeneic peripheral stem cell (PBSC) collection in healthy donors. Methods: We retrospectively reviewed clinical data of allogeneic PBSC collection in 175 donors from 2007 to 2017 at the National Cancer Center, Korea. This study analyzed factors associated with the CD34+ cell yield such as the characteristics of donors, including age, laboratory results before apheresis, and data of procedures on the first day. The CD34+ cell dose of ≥ 4.0 × 106/kg have recently been the accepted minimum recommended dose in allogeneic HSCT settings, and this was the target dose in our study. Results: The factors associated with the CD34+ cell yield were age (p = 0.007), baseline platelet (PLT) (p = 0.014), and pre-collection hematopoietic progenitor cells (HPCs) (p = 0.001) by multivariate analysis. This study represented that age, baseline platelet count, and pre-collection HPC count are important predictive factors as shown in other previous studies. Conclusion: Our data suggest that young age, high baseline platelet counts and high HPC counts before collection might be useful for identifying successful mobilizers.
1. Introduction Allogeneic hematopoietic stem cell transplantation (HSCT) is an established procedure for many hematologic malignancies [1]. Peripheral blood stem cells (PBSCs) are increasingly used as the source of stem cells in HSCT because of improved neutrophil and platelet (PLT) engraftment, avoidance of general anesthesia for collection, and potentially better graft-versus-leukemia effects [2]. Factors associated with successful allogeneic peripheral blood stem cell transplantation (PBSCT) include age of the patient, disease stage, time interval from diagnosis to transplant, human leukocyte antigen (HLA) and ABO compatibility between donor and recipient, and number of harvested stem cells [3–5]. Especially, obtaining a sufficient number of stem cells using as few leukapheresis procedures as possible is important in regard to donor convenience, therapeutic results, and cost [6]. In allogeneic PBSCT, the optimal number of CD34+ cells for transplantation has not been definitively established, but a higher dose is often preferred at 5.0 × 106 CD34+ cells per kilogram (kg) of body
weight [7]. It is widely accepted that the minimum cell doses for autologous and allogeneic transplantation are 2.0 × 106 and 3.0 × 106 to 5.0 × 106 of CD34+ cells per kg, respectively [8]. However, ≥4.0 × 106 cells per kg have recently been the accepted minimum recommended dose [9,10]. In addition, a CD34+ cell dose of more than 4.0 × 106 per kg of body weight resulted in more rapid engraftment, lower transplant-related mortality, and better overall survival [11]. Thus, the target CD34+ cell dose was set at ≥4.0 × 106/kg in this study. Predicting the successful mobilization of CD34+ cells before the PBSC collection is preferable for the benefit of the donors and recipients. There are several factors reported to affect the collection of allogeneic PBSCs including sex, age, and method of mobilization, the number of peripheral white blood cells (WBCs), CD34+ cells, platelets count, and circulating immature cells [12–16]. Especially, a circulating CD34+ cell count before apheresis was widely used as an indicator for predicting the PBSC harvest yield, but is expensive and needs specific equipment and experienced staff. Hematopoietic progenitor cells
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Corresponding authors at: Department of Laboratory Medicine, National Cancer Center, 323 Ilsan-ro, Ilsandong-gu, Goyang-si Gyeonggi-do 10408, South Korea. E-mail addresses: [email protected] (S.-N. Lee), [email protected] (S.-Y. Kong). 1 Jung Hee Kong and Youmi Hu equally contributed to this work. https://doi.org/10.1016/j.transci.2019.102679 Received 18 September 2019; Received in revised form 7 November 2019; Accepted 11 November 2019 1473-0502/ © 2019 Elsevier Ltd. All rights reserved.
Please cite this article as: Jung Hee Kong, et al., Transfusion and Apheresis Science, https://doi.org/10.1016/j.transci.2019.102679
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(HPCs) quantification is rapid, inexpensive and requires minimal training [17]. Here, we retrospectively analyzed the yield of CD34+ cells from 175 healthy donors. The aim of this study was to investigate the factors influencing successful allogeneic PBSC collection of CD34+ cells.
Table 1 Characteristics of donors and variables of apheresis.
2. Materials and methods 2.1. Donor characteristics and variables of apheresis Two hundred thirty-four procedures were performed on 175 healthy donors and 59 donors required a second procedure between February 2007 and December 2017. This study was approved by the ethical review boards at the authors’ institution (NCC2016-0013). In our study, only the data of the first day were analyzed. The characteristics of the donors include sex, age, body weight (BW), and routes of vascular access (peripheral vein or central venous catheter). Blood parameters include counts of WBCs, mononuclear cells (MNCs) and HPCs measured before and after the procedure using XE-2100 automatic analyzer (Sysmex, Kobe, Japan). Enumeration of the HPC was performed in the immature myeloid information (IMI) channel, which identifies a population of hematopoietic progenitor cells based on cell size, complexity and differential cell lysis characteristics. The CD34+ cell count was measured by two-color flow cytometry using FACS Canto II (Becton, Dickinson and Company, San Jose, CA, USA) after immunofluorescence staining using anti-human CD34 monoclonal phycoerythrin-conjugated (PE) antibodies and CD45 monoclonal fluorescein isothiocyanate-conjugated antibodies (FITC) (Becton–Dickinson, New Jersey, USA). The number of CD34+ cells per BW in product was calculated by donor’s BW. We calculated the collection efficiency (CE) by using MNCs instead of CD34+ cells as the pre-collection CD34+ cell counts were not performed. The CE was estimated as described previously as [MNCs obtained from apheresis / MNCs (μL) × Apheresis volume processed (μL)] [6].
Variable
Median(range)
Number of donors Sex(male/female) Age (years) Donor-recipient relation Related Unrelated Donor BW (kg) Vascular access, N (%) Pheripheral vein Central venous catheter Baseline-PBSC collection WBC count (×109/L) Hct (%) PLT (×109/L) MNC (%) Pre- PBSC collection WBC count (×109/L) HPC (×106/L) Hct (%) PLT (×109/L) MNC (%) Post- PBSC collection WBC count (×109/L) Hct (%) PLT (×109/L) MNC (%)
175 107(61 %)/68(39 %) 35
(14–69)
103 72 69.2
(59 %) (41 %) (44.2–115.4)
134 41
(77 %) (23 %)
5.98 43.7 244 41
(2.98–13.15) (31.2–53.2) (164–460) (24–62)
44.78 60 40.9 196 19
(22.75–80.71) (2–296) (28.2–50.1) (99–339) (10–34)
34.68 41.3 85 11
(11.44–73.83) (29.4–48.9) (33–225) (3–36)
Abbreviations: SD, standard deviation; BW, body weight; WBC, white blood cell; MNC, mononuclear cells; HPC, hematopoietic progenitor cells; Hct, hematocrit; PLT, platelet. Table 2 Summary of procedures and collected products. Variable
Median(range)
Number of procedures 175 TBV (mL) 4557 TPV (mL) 17,000 Ratio of TPV to TBV 3.8 Volume of product (mL) 312 PBSC product blood parameters of collection Total WBCs (×108/kg) 9.73 Total MNCs (×108/kg) 8.00 Total CD34+ cells (/uL) 1113 Total CD34+ cells (×106/kg) 4.60 MNC efficiency (%)* 40.8 Platelet loss (%)† 56.3 Instrument, N (%) COBE Spectra 120 Spectra Optia 55 MNC collection efficiency according to instrument (%)* COBE Spectra 40.2 Spectra Optia 42.6
2.2. Mobilization and collection of allogeneic PBSC To mobilize the PBSCs, granulocyte-colony stimulating factor (GCSF) was administered subcutaneously once daily for 4 days to all donors at a dose of 10 mcg/kg. PBSC collection was initiated on the 5th day after mobilization. Stem cells were collected using either a central vein access via a perm catheter or peripheral vein access via 16 -18 gauge catheters. We used two different collecting machines. COBE Spectra (Terumo BCT, Inc.) was operated according to the manufacturer’s standard operating procedure based on the color scale 2–3% and Spectra Optia (Terumo BCT, Inc.), which has electronic features including optical sensors for continuous automatic monitoring. During the PBSC procedure, 3 to 4 times of the donor’s total blood volume was processed at a rate of 50 to 70 mL/min. The whole blood-to-anticoagulant (3000 IU heparin with 500 mL of acid-citrate-dextrose solution) ratio was 24:1 in both devices. The study investigated the number of procedures, total blood volume (TBV) of donors, total processed volume (TPV), TPV-toTBV ratio, and product volumes. We divided the donors into two groups according to the collected CD34+ cells in collected product on the first procedure: CD34+ in product < 4.0 × 106/kg and ≥ 4.0 × 106/kg.
(3010-6388) (9135-23,449) (2.1–6.5) (80–450) (2.10–22.30) (1.85–18.28) (156-4,404) (0.47–17.08) (13.9–96.9) (-9.2–77.7) (69 %) (31 %) (19.4–96.9) (13.9–73.2)
Abbreviations: TBV, total blood volume; TPV, total processed volume; PBSC, peripheral blood stem cell; WBC, white blood cell; MNC, mononuclear cells; CD34, Cluster of Differentiation antigen; *MNC efficiency % = MNCs obtained from apheresis / MNCs (μL) × apheresis volume processed(μL). † PLT loss % = (platelets/μL before apheresis – platelets/μL after apheresis) / platelets/ μL before apheresis.
univariate analysis. A multivariate backward stepwise selection regression analysis was used to derive a formula for predicting the cell yield. To predict successful mobilization, cut-off value of baseline platelet counts and pre-collection HPC counts were analyzed by receiver operating characteristic (ROC) analysis.
2.3. Statistics All statistical analyses were performed by using Stata Statistical Software Release 12 (StataCorp LP, College Station, TX, USA). The data was presented as the median with a range. A p value < 0.05 was considered statistically significant. Continuous variables were compared using the Kruskal-Wallis test, and categorical variables were analyzed using the Fisher’s exact test. The binary logistic regression model was used to analyze parameters found to be statistically significant at 2
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Table 3 Comparison between two groups according to the CD34+ cell yield.
Sex (male/female) Age (years) Donor BW (kg) Vascular access type, N (%) Peripheral vein Central venous catheter Instrument, N (%) COBE spectra Spectra Optia Apheresis variables TBV (mL) TPV (mL) Ratio of TPV to TBV Volume of product (mL) COBE Spectra Spectra Optia Baseline-PBSC collection WBC count (×109/L) Hct (%) PLT (×109/L) MNC (%) Pre-PBSC collection WBC count (×109/L) HPC (×106/L) Hct (%) PLT (×109/L) MNC (%) Post-PBSC collection WBC count (×109/L) Hct (%) PLT (×109/L) MNC (%) PBSC product Total WBCs (×108/kg) Total MNCs (×108/kg) Total CD34＋cell (/μL) Platelet loss (%)†
＜4.0 × 106 cells/kg
≥4.0 × 106 cells/kg
(N = 68, 39 %)
(N = 107, 61 %)
p value
38(56 %)/30(44 %) 40 67.9
(21–63) (47.7–115.4)
69(64 %)/38(36 %) 34 69.6
(14–69) (44.2–108.3)
0.255 0.005* 0.667 0.261
49 19
(72 %) (28 %)
85 22
(79 %) (21 %)
50 18
(74 %) (26 %)
70 37
(65 %) (35 %)
4344 18,000 3.8
(3,030-6388) (9135-23,449) (2.1–6.1)
4590 16,500 3.9
(3,010-6,330) (10,129-20,256) (2.1–6.5)
289 292
(170–364) (188–400)
311 342
(80–396) (198–450)
5.82 43.7 233 42
(2.98–11.82) (32.7–50.1) (164–359) (26–62)
6.11 43.7 249 40
(3.54–13.15) (31.2–53.2) (167–460) (24–55)
0.022* 0.895 < 0.001* 0.051
42.80 41 40.6 189 19
(23.01–67.14) (2–249) (28.2–48.4) (99–186) (10–33)
47.18 78 41.7 206 20
(22.75–80.71) (11–296) (31.3–50.1) (117–339) (10–34)
0.007* < 0.001* 0.741 0.006* 0.431
32.00 40.7 79 11
(11.44–54.30) (30.1–48.4) (33–186) (6–24)
37.27 41.7 89 12
(19.95–73.83) (29.4–48.9) (48–255) (3–36)
0.002* 0.382 0.003* 0.718
8.64 7.30 606 56.8
(2.10–16.44) (1.85–14.63) (156-1,524) (18.1–77.7)
10.66 8.55 1561 55.5
(5.26–22.30) (4.11–18.28) (582-4,404) (-9.2–76.7)
< 0.001* < 0.001* < 0.001* 0.690
0.260
0.465 0.384 0.292 0.030*
Abbreviations: CD34, Cluster of Differentiation antigen; BW, body weight; TBV, total blood volume; TPV, total processed volume; WBC, white blood cell; Hct, hematocrit; PLT, platelet; MNC, mononuclear cells; HPC, hematopoietic progenitor cells. * p < 0.05 statistically significant. † PLT loss % = (platelets/μL before apheresis – platelets/μL after apheresis) / platelets/μL before apheresis ×100.
median collected CD34+ cell count was 4.60 × 106/kg (0.47–17.08 × 106/kg). Furthermore, 120 (69%) and 55 (31%) procedures were performed on the COBE Spectra and Spectra Optia. The median MNC collection efficiency by the COBE Spectra and Spectra Optia were 40.2 % (19.4–96.9 %) and 42.6 % (13.9–73.2 %) (Table 2).
3. Results 3.1. Demographic details of the allogeneic donors and pre and post collection parameters of peripheral blood counts Among the 175 donors (107 male and 68 female), 103 (59 %) were related donors, and 72 (41 %) were unrelated donors. The median age and donor’s BW were 35 years (14–69 years) and 69.2 kg (44.2–115.4 kg), respectively. Venous access was peripheral vein in 134 (77 %) donors and central venous in 41 (23 %) donors. The median baseline WBC and PLT were 5.98 × 109/L (2.98–13.15 × 109/L) and 244 × 109/L (164–460 × 109/L), respectively. The median pre-collection WBC, PLT, and HPC counts were 44.78 × 109/L (22.75–80.71 × 109/L), 196 × 109/L (99–339 × 109/L), and 6 6 60 × 10 /L (2–296 × 10 /L). The median post-collection WBC and PLT counts were 34.68 × 109/L (11.44–73.83 × 109/L) and 85 × 109/l (33–225 × 109/L), respectively (Table1).
3.3. Comparison between the poor and good mobilizer groups We divided the donors into two groups according to the collected CD34+ cells in the product of the first procedure. We defined the poor mobilizer as < 4.0 × 106/kg and the good mobilizer as ≥ 4.0 × 106/kg CD34+ cells in product. 107 (61 %) donors reached a CD34+cell count ≥ 4.0 × 106/kg, and only 68 (39 %) did not. There were significant differences between the two groups for age (p = 0.005), volume of product (p = 0.030), baseline-collection WBC (p = 0.022), PLT (p < 0.001), pre-collection WBC (p = 0.007), HPC (p < 0.001), PLT (p = 0.006), post-collection WBC (p = 0.002), PLT (p = 0.003) and total WBCs (p < 0.001), MNCs (p < 0.001), CD34+cells (p < 0.001) in product. The median age of the group achieving a CD34+cell count of ≥ 4.0 × 106/kg was lower than that of the group achieving a CD34+cell count of < 4.0 × 106/kg. The count of baseline-collection PLT and pre-collection HPC in the group ≥ 4.0 × 106/kg was higher than that of the group < 4.0 × 106/kg (Table 3).
3.2. Results of PBSC collection and apheresis products A total of 175 PBSC collections were conducted during the study period. The median total blood volume (TBV) was 4557mL (30106388 mL). The median total processed volume (TPV) was 17,000 mL (9135–23,449 mL), and the TPV-to-TBV ratio was 3.8 (2.1–6.5). The 3
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Table 4 Univariate and multivariate analysis for predictive factors of optimal PBSC collection in donors. Odds ratio Sex Male Female Age (per 1 year increase) Donor BW (per 1 kg increase) Vascular access type Peripheral vein Central venous catheter Instrument COBE spectra Spectra Optia Apheresis variables TBV (mL) TPV (mL) Ratio of TPV to TBV Baseline-PBSC collection WBC count (×109/L) Hct (%) PLT (×109/L) MNC (%) Pre-PBSC collection WBC count (×109/L) HPC (×106/L) Hct (%) PLT (×109/L) MNC (%)
95 % CI
p value
1 0.698 0.961 1.003
0.375-1.298 0.936-0.987 0.982-1.024
0.256 0.003* 0.787
1 0.667
0.329-1.354
0.263
1 1.468
0.751-2.869
0.261
1.000 0.999 0.829
0.999-1.000 0.999-1.000 0.585-1.174
0.503 0.504 0.291
1.000 1.001 1.014 0.96
1.000-1.001 0.930-1.077 1.006-1.021 0.922-1.001
0.015* 0.982 < 0.001* 0.053
1.000 1.013 1.021 1.011 1.026
1.000-1.001 1.006-1.020 0.945-1.103 1.003-1.018 0.962-1.095
0.006* < 0.001* 0.595 0.005* 0.429
Odds ratio
95 % CI
p value
0.961
0.934-0.989
0.007*
1.010
1.002-1.018
0.014*
1.012
1.005-1.019
< 0.001*
Abbreviations: PBSC, peripheral blood stem cell; BW, body weight; TBV, total blood volume; TPV, total processed volume; WBC, white blood cell; Hct, hematocrit; PLT, platelets; MNC, mononuclear cells; HPC, hematopoietic progenitor cells. * p < 0.05 statistically significant.
Fig. 1. ROC curve of baseline PLT and pre-collection HPC count for predicting successful PBSC collection (4.0 × 106/kg CD34+ cells) in donor. (A) The cutoff value of the baseline PLT count was 210 × 109/L (sensitivity = 91.0 %, specificity = 76.6 %, AUC = 0.665). (B) The cutoff value of the pre-collection HPC count was 25 × 106/L (sensitivity = 91.0 %, specificity = 73.8 %, AUC = 0.667). Abbreviations: ROC receiver operating characteristic; PLT platelet; HPC hematopoietic progenitor cells; PBSC peripheral blood stem cell; AUC area under the curve
1.010, p = 0.014), and pre-collection HPC (OR 1.012, p < 0.001) (Table 4). The cutoff value of the baseline platelet count for predicting successful PBSC collection of 4.0 × 106/kg CD34+ cells was 210 × 109/L with 91.0 % sensitivity and 76.6 % specificity in ROC analysis. The cutoff value of the pre-collection HPC count for the same amount of CD34+ cells was 25 × 106/L with 91.0 % sensitivity and 73.8 % specificity. Area under the curve (AUC) of baseline PLT and precollection HPC count were 0.665 and 0.667, respectively (Fig. 1).
3.4. Predictive value of successful PBSC collection In univariate analysis, factors associated with a sufficient amount of stem cells were age [odds ratio (OR) 0.961, p = 0.003], baseline-collection WBC (OR 1.000, p = 0.015), PLT (OR 1.014, p < 0.001) and Pre-collection WBC (OR 1.000, p = 0.006), HPC (OR 1.013, p < 0.001), PLT (OR 1.011, p = 0.005). In multivariate backward stepwise selection regression analysis, significant predicting factors were donor’s age (OR 0.961, p = 0.007), baseline-collection PLT (OR 4
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4. Discussion
But, it was not a significant influential factor. It is known that the mobilization method is the most important factor influencing the allogeneic hematopoietic stem cell collection in healthy donors [18]. In particular, studies were conducted on the types of mobilization agent, doses, and duration of administration. In most published prospective and retrospective studies, G-CSF was administered to healthy donors for 4–5 days. The Italian Bone Marrow Donor Registry (IBMDR) recommends unrelated donors to limit the administration of G-CSF to a maximum of 5 days [10]. In several studies, PBSC mobilization using G-CSF alone showed in a higher CD34+ cell yield if the dose of G-CSF was applied 5mcg/kg twice daily rather than 10mcg/ kg once daily [28,29]. However, previous study reported that there were no significant differences in G-CSF induced mobilization of healthy donors between daily single and divided dose of G-CSF [30]. In our study, 10 mcg/kg/day of G-CSF was administered for 4 days, and the collection was initiated on the 5th day. More than 4.0 × 106/kg of the CD34+ cells were collected in 61 % (107/175) of the donors during the first leukapheresis. Recently, a study has reported that more than 4.0 × 106/kg CD34+ cells were collected from 62 % (31/50) of the donors in the first procedure after administration of G-CSF 10 mcg/kg/ day [22]. In conclusion, we found that the donor’s age and platelet count before mobilization were important factors for allogeneic PBSC collection in healthy donors. Furthermore, pre- collection HPCs were a significant factor that correlated with the CD34+ cell yield. Thus, young age and high baseline platelet counts and high HPC counts before collection might be useful for identifying successful mobilizers.
Several studies have also explored predictive factors for CD34+ cell yield. The number of WBCs, platelet count, CD34+ cells and circulating immature cells before and during apheresis have been reported to be factors [6,8,14,16,18]. Suzuya et al. reported that the factors associated with the CD34+ cell yield included the platelet count before and during mobilization [18]. Furthermore, Lysák et al. reported that a significant correlation was observed between CD34+ yield and donor platelet count measured before mobilization [14]. In this study, the platelet count before mobilization was observed to be a factor influencing the CD34+ cell yield. The CD34+ cell count on the day of apheresis is a powerful predictor [5]. However, the circulating CD34+ cell count before apheresis was widely used as an indicator for predicting the PBSC collection yield but was both time consuming and expensive. HPCs represent a specific population of immature myeloid cells, and can be measured quickly, inexpensively, and can be performed as part of a routine complete blood count study with minimal extra cost by automated hematology analyzer [8]. Therefore, a number of studies have been conducted to evaluate HPCs as a predictor of CD34+ cell yield. Many studies that investigated autologous PBSC collection demonstrated that there was an acceptable correlation between pre-collection HPC and CD34+ cell counts, as well as CD34+ cell yields after PBSC collection [19,20]. In our study, pre-collection HPC count was analyzed to be an important factor associated with the CD34+ cell yield. Also the cutoff value of pre-collection HPC count for predicting successful PBSC collection was 25 × 106/L. Yang et al. suggests that a preharvest HPC count of more than 21.3 × 106/L may predict adequate CD34+ (4.0 × 106/kg) cell yield [8]. In this study, the cutoff value of pre-collection HPC count was similar to the previous study. Namba et al. reported that significant factors for predicting successful collection included the sex of the donor [5]. Wang et al. noted that females had a lower mean collected outcome and a higher risk for poor yield than males [21]. However, the sex of donor was considered to have no effect on stem cell yield in many studies [11,14,22]. In our study, there was not a significant difference in donor sex among the two groups according to the number of CD34+ cells in the collected products. Age was reported to be an important factor in stem cell collection [11,14,18]. Pornprasertsud et al. reported that successful CD34+ cell counts of 5.0 × 106/kg or more were available from donors younger than 30 years of age [22]. Anderlini et al. reported that a donor age greater than 55 years was associated with a 3.8-fold greater risk for poor mobilization. There was a study that the donor’s age, with a threshold of 38 years or more, was a factor that significantly affected CD34+ cell mobilization and collection in healthy donors [11,16]. In this study, multiple regression analysis showed that age was significantly associated with the CD34+ cell yield. It has been reported that there is no significant difference between the peripheral vein and the central venous catheter in the donor’s venous access type [14,23]. Wang et al. (2008) reported that female donors had higher CD34+ cell yields if the circulation access was through an artery than if it was through a vein [21]. At our institution, central venous catheters are exclusively applied for collecting PBSCs from healthy donors with poor peripheral venous access. Furthermore, there have been no cases that used arterial access. There was not a significant difference between the two groups according to the type of venous access. Although there is a difference in total processing blood volume, blood flow rate and equipment, it has been previously shown that that there is no difference in CD34+ cell counts in the collected products [24–26]. Large-volume leukapheresis (LVL) is defined as processing 3 or up to 6 times the donor’s total blood volume. LVL has been suggested as a method of choice in donors and patients with a weak effect of mobilization [27]. In our study, the median TPV-to-TBV ratio was 3.6.
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