Clotting during autologous hematopoietic progenitor cells collection

Autologous hematopoietic progenitor cell (HPC) transplant through peripheral blood mobilization and leukapheresis is a standard treatment for many patients with hematopoietic malignancies. Although leukapheresis is usually completed with no complications, we present a case in which the hematopoietic progenitor cells clotted during collection. The patient had no history of hypercoagulopathy. It was identified that the anticoagulant infusion line was partially constricted by a blood warmer clamp. The machine did not alarm. Most of the multiple Food and Drug Administration reports of clotting occurring during apheresis procedures were due to the patients' preexisting hypercoagulopathy or insufficient anticoagulant solution being used. The machine alarmed in most of these cases. Our case demonstrates that inadequate anticoagulation can occur during an HPC collection procedure without activation of an alarm.     

Crude collection efficiency of CD34+ hematopoietic stem cell apheresis

Understanding the apheresis principles for harvesting hematopoietic stem cells (HSCs) is critical for performing efficient procedures. However, despite significant advances in estimating the collection efficiency (CE) of aphereses, many confounding factors still need to be addressed in the classical calculations. The CE values are unrestricted, and many procedures exhibit CEs of a given cell population greater than 100%. This report introduces a simple equation that estimates the “crude” CE, which ranges from 0% to 100% and intrinsically considers the contribution of donor-related variables such as the pre-procedure mobilization and intra-apheresis recruitment of CD34⁺ cells (as a convenient marker for HSCs), as well as the performance of the apheresis system itself.     

Evaluation of the spectra optia apheresis system for mononuclear cell (MNC) collection in G‐CSF mobilized and nonmobilized healthy donors: Results of a multicenter study

The Spectra Optia apheresis system is a newer centrifugation‐based device that in comparison with the COBE Spectra includes features that enhance procedure automation and usability. In this FDA‐approved three‐center two‐arm observational study we characterized the performance of the Spectra Optia for collection of MNCs and CD34+ cells from nonmobilized and granulocyte‐colony stimulating factor (G‐CSF) mobilized healthy donors, respectively. There were a total of 15 evaluable subjects in each arm. Key performance indicators included collection efficiency of MNCs/CD34+ cells, product purity and cellular viability. For nonmobilized donors, median MNC collection efficiency, platelet collection efficiency, product hematocrit and granulocyte contamination were 57%, 12%, 4%, and 1.7%, respectively. For mobilized donors, median MNC collection efficiency, CD34+ cell collection efficiency, platelet collection efficiency, product hematocrit and granulocyte contamination were 61%, 77%, 19%, 4%, and 15%, respectively. Average WBC viability in the mobilized products was 99%. There was one severe (grade 3) adverse event related to citrate toxicity. This study demonstrates that the Spectra Optia can be used for safe and efficacious collection of MNCs, and results obtained are in line with expectations on collection efficiency and product characteristics. Adverse events were limited to those that are well documented in the stem‐cell mobilization and leukapheresis process. As of the time of this writing, FDA 510(k) approval for use of the Spectra Optia device for MNC collection was achieved in the US based partly on the results of this study. J. Clin. Apheresis 29:273–280, 2014. © 2014 Wiley Periodicals, Inc.     

Three-hour collection of committed and multipotent hematopoietic progenitor cells by apheresis

Although multipotent progenitor cells have been collected from blood in humans by apheresis in 1- to 2-hour collections, longer-term collections of multipotent progenitor cells have not been reported. To determine whether such collections could be sustained, the authors performed six 3-hour collections on five normal volunteers, processing a mean of 8024 ml of blood. Blood samples were taken at 0, 1.5, and 3 hours, and component samples were taken at 1.5 and 3 hours. Cell counts, differentials, and committed myeloid (CFU-GM), erythroid (BFU-E), and multipotent (CFU-MIX or CFU-GEMM) progenitor cell assays were performed. Collection efficiency for progenitor cells, the number of cells collected divided by the number in the blood processed, did not diminish significantly. The number collected in the second 1.5-hour period resembled the number collected during the first. Circulating numbers did not fall significantly during apheresis. Prolongation of apheresis collections may reliably increase the number of progenitor cells available for eventual hematopoietic reconstitution.     

幼儿外周血造血干/祖细胞采集得率影响因素分析

目的考察利用COBE Spectra血细胞分离机采集幼儿外周血造血干/祖细胞的影响因素。方法根据患儿体重(≤15 kg,>15 kg)、采集时外周血白细胞计数(≤5×109/L,>5×109/L)、淋巴细胞+单核细胞比例(≤18%,>18%)、Hct(≤25%,>25%)、采血速度(≤20 mL/min,>20 mL/min)血细胞分离机报警次数(≤3次,>3次)分别分组,比较2组患儿的每公斤体重单个核细胞计数、CD34+细胞计数及所占比例。结果淋巴细胞+单核细胞比例>18%组与≤18%组比较,Hct>25%组与≤25%组比较,血细胞分离机报警次数≤3次组与>3次组比较,在每公斤体重单个核细胞计数、CD34+细胞计数及所占比例均有显著差异(P<0.01)。其余组别之间差异不显著(P>0.05)。结论采用COBE Spectra血细胞分离机采集幼儿外周血造血干/祖细胞时,淋巴细胞+单核细胞比例、患儿红细胞比积及血细胞分离机报警次数是影响采集得率的因素。     

Safety of large-volume leukapheresis for collection of peripheral blood progenitor cells

Large volume leukapheresis (LVL) reduces the number of procedures required to obtain adequate peripheral blood progenitor cells (PBPCs) for autologous hematopoietic reconstitution. LVL involves the processing of >15 L or 5 patient blood volumes using high flow rates. We report our experience with LVL evaluating its efficiency and adverse effects in 71 adult patients with hematologic or solid organ malignancies. All were mobilized with chemotherapy and granulocyte colony-stimulating factor (G-CSF). All collections used a double lumen apheresis catheter. Means values per LVL were as follows: blood processed, 24.6 L; patient blood volumes processed, 5.9; ACD-A used. 1.048 ml; heparin used, 6,148 units; collect time, 290 min; blood flow rate, 89 ml/min. Eighty percent of the collections were completed in one or two procedures to obtain ≥6.0 × 10⁸ MNCs/kg body weight. The most frequent side effect (39%) was parasthesia due to citrate-related hypocalcemia. This was managed with oral calcium supplements and or slower flow rates. Post-LVL electrolyte changes were generally asymptomatic. Prophylactic oral potassium supplements were administered in 57% of cases. Other reactions included hypotension (4%), prolonged parasthesia (1.4%), and headache (1.4%). Catheter problems in 9 (13%) of the procedures were attributed to clot formation (37%) or positional effects (63%). No bleeding occurred. Post-LVL decreases in hematocrit and platelet count averaged 3.5% and 46%, respectively. Six (4%) of the procedures required red blood cell transfusions. Platelet transfusions were given in 19 (13%) of the procedures. We conclude that adverse reactions with LVL are similar to those reported for conventional PBPC collections, making it safe and efficacious as an outpatient procedure. J. Clin Apheresis 12:10–13, 1997. © 1997 Wiley-Liss, Inc.     

Evaluation of a new protocol for peripheral blood stem cell collection with the Fresenius AS 104 cell separator

In this report we analyzed sixty leukapheresis procedures on 35 patients with a new protocol for the Fresenius AS 104. Yields and efficiencies for MNC, CD 34+ cells, and CFU-GM indicate that the new protocol is able to collect large quantities of hemopoietic progenitors. Procedures were performed processing 8.69 ± 2.8 liters of whole blood per apheresis and modifying 3 parameters: spillover-volume 7 ml, buffy-coat volume 11.5 ml, centrifuge speed 1,500 rpm; blood flow rate was 50 ml/min and the anticoagulant ratio was 1:12. No side effects were observed during apheresis procedures except for transient paresthesia episodes promptly resolved with the administration of calcium gluconate. Yields show a high capacity of the new program to collect on average MNC 17.28 ± 10.85 × 10⁹, CD 34+ 471 ± 553.5 × 10⁶ and CFU-GM 1278.7 ± 1346.3 × 10⁴ per procedure. Separator collection efficiency on average was 49.91 ± 23.28% for MNC, 55.1 ± 35.66% for CFU-GM, and 62.97 ± 23.09% for CD 34+ cells. Particularly interesting are results for MNC yields and CD 34+ efficiency; these results make the new program advantageous or similar to the most progressive blood cell separators and capable to collect a sufficient number of progenitor cells for a graft with a mean of 1.80 ± 0.98 procedures per patient. J. Clin. Apheresis 12:82–86, 1997. © 1997 Wiley-Liss, Inc.     

Safety and efficacy of peripheral blood progenitor cell mobilization and collection in patients with advanced coronary heart disease

Information on the safety of mobilization and collection of peripheral blood progenitor cells (PBPC) in patients with advanced coronary heart disease (CHD) is limited. We report herein our early experience with patients participating in a Phase I trial of injection of autologous CD 34(+) cells into threatened, ischemic myocardium for neovascularization and symptom relief in patients with chronic refractory myocardial ischemia. All patients had advanced inoperable CHD despite the best medical therapy. Granulocyte colony stimulating factor (G-CSF, 5 microg/kg/day) was administered subcutaneously for 5 days for mobilization of CD34(+) cells into the peripheral blood. PBPCs were collected in the outpatient apheresis suite on day 5. Nine patients from our institution were evaluable. Adverse effects of mobilization included: increase in frequency and/or intensity of angina in 8 patients (88.8%); bone pain in 7 patients (77.7%); headaches in 4 patients (44.4%); 2 patients (22%) were hospitalized. Collection phase toxicities included: tingling in 5 patients (55.5%) and angina in 3 patients (33%). All procedures were completed without new myocardial infarction, congestive heart failure, or death. The median peripheral blood CD34(+) cell count on day 5 of G-CSF was 21 cells/microl (range 10-40 cells/microl). A median of 1.65 x 10(6) CD34(+) cells/kg (range: 0.13-3.0 x 10(6)/kg) were harvested. We conclude that mobilization and collection of PBPC in patients with advanced CHD can be safely performed as an outpatient procedure. Apheresis professionals should be aware of the intensity and frequency of angina in this patient population.     

Reflections on methodical approaches to hematopoietic stem cell collection in children

Pediatric peripheral blood stem cell collection (PBSC) is challenging because it has potentially more side effects than in adults due to the small body mass and unique physiology of children. The extracorporeal volume of the cell separator device, poor venous access and metabolic complications due to citrate toxicity are the main problems to face during PBSC collection. These aspects are more relevant in very low body weight (BW) children of 20?kg or lower. An efficient, experienced and well-prepared team of pediatricians, apheresis physicians and nurses, and physicians involved in CVC positioning is crucial to performing a safe PBSC collection. Despite the growing demand for PBSC collection in the pediatric setting, there is not an actual unique standardized detailed practice approach to be employed, therefore, on reflection, we believe that it is timely to draw up useful evidence-based recommendations on which guidelines can be developed for use by those groups with limited or no experience.     

Counterflow centrifugation apheresis for the collection of autologous peripheral blood stem cells from patients with malignancies: a comparison with a standard centrifugation apheresis procedure

Two apheresis methods used to collect hematopoietic stem cells from peripheral blood were compared in eight patients with a variety of malignancies. The standard lymphocyte collection method was alternated with the counterflow centrifugation or lymphocyte surge protocol. The number of clonogenic cells (CFU-GM and BFU-E), the red cell volume, and the number of mononuclear cells in each collection were assessed as well as the changes in circulating leukocytes, platelets, and blood hemoglobin produced by each apheresis procedure. There was no statistically significant difference found in the number of clonogenic cells collected with either method, but the number of mononuclear cells collected with the standard procedure was significantly higher (P = 0.001). The red cell volume collected with the standard procedure was significantly higher, (P = 0.0001), but corrected for the number of mononuclear cells the difference was not significant. The counterflow centrifugation apheresis produced significantly less thrombocytopenia (P = 0.005). The counterflow centrifugation apheresis procedure used collected fewer mononuclear cells than the standard procedure, however, with less red cell contamination but a comparable number of CFU-GM and BFU-E in four hour apheresis procedures. Each collection method resulted in a comparable amount of anaemia and leukopenia but the lymphocyte surge method produced less thrombocytopenia following the collection.     

Thrombosis and ELISA optical density values in hospitalized patients with heparin-induced thrombocytopenia

The natural history of heparin-induced thrombocytopenia (HIT) in the absence of thrombosis was previously established using functional assays for confirmation of diagnosis (e.g. 14C serotonin release assay). An enzyme-linked immunosorbent assay (ELISA) that detects the presence of antibodies directed against the heparin-platelet factor-4 (PF4) complex has largely replaced functional assays in many medical centers. Although the ELISA is highly sensitive for detecting HIT antibodies, its usefulness for predicting thrombotic outcomes has not been clearly established. We performed a retrospective chart review of all hospitalized patients at a university hospital who tested seropositive for HIT by a commercial ELISA during 2001 and 2002. A total of 63 inpatients were identified as HIT positive by ELISA. Forty-eight patients had no apparent HIT-associated thrombosis at the time of HIT seropositivity (i.e. isolated HIT) and only one was treated prophylactically with a direct thrombin inhibitor. The 30-day thrombosis rate for patients with isolated HIT was 17% (eight of 48). Higher ELISA optical density (OD) measurements correlated significantly with thrombosis (1.41 +/- 0.87 vs. 0.79 +/- 0.46, P <0.001). Patients with isolated HIT and an OD measurement of > or = 1.0 demonstrated nearly a 6-fold increased risk of thrombosis compared with those with OD values between 0.4 and 0.99 (odds ratio 5.74, 95% confidence interval 1.73, 19.0; absolute rate of thrombosis, 36% vs. 9%, respectively, P=0.07). We conclude that in hospitalized patients with isolated HIT, the presence of heparin-PF4 antibodies detected by ELISA was associated with a significant risk of subsequent thrombosis and higher ELISA values were observed among patients suffering thrombotic events.     

Manual color monitoring to optimize hematopoietic progenitor cell collection on the Fenwal Amicus

A technique was developed to improve consistency of MNC transfers from the centrifuge to the collection bag in the Fenwal Amicus. The operator assures that RBCs completely fill the cassette by the end of the transfer by adjusting the RBC offset in succeeding cycles. We compared yields and crosscellular content before and after implementation of the monitoring technique. Retrospective data from 400 consecutive HPC collection procedures (200 for each technique) were compared. In 40 monitored collections, the RBC offset was adjusted to 6-9 mL to ensure that RBCs completely filled the cassette. Collections requiring these adjustments were not associated with a specific diagnosis. Median values were compared between the 40 collections requiring offset adjustment and those performed before implementation of monitoring. Baseline peripheral CD34+ cell (17 vs. 14 cells μL(-1)), lymphocytes (2 vs. 1.3 × 10(9) /L), WBCs, HCT, and PLTs were significantly higher in the group requiring offset changes. The group requiring offset changes had significantly more CD34+ cells per collection (190.8 × 10(6) or 2.04 × 10(6) /kg vs. 84.3 × 10(6) or 0.89 × 10(6) /kg) and more lymphocytes per collection (16.9 × 10(9) vs. 11.6 × 10(9)). Crosscellular content of the group requiring offset changes was significantly higher for WBCs (41.8 vs. 33.1 × 10(9)), granulocytes (9.6 vs. 7.2 × 10(9)), RBCs (23 vs. 17 mL), and PLTs (2.1 vs. 1.2 × 10(11)). Manual monitoring is a simple, inexpensive method to optimize each HPC collection to maximize CD34+ cell and lymphocyte yields.     

Blood component collection by apheresis

Apheresis component collection is a rapidly growing area in the blood collection field. Several instruments with varying capabilities are available. This is a brief review of the equipment available for granulocyte and apheresis component collection and indications for their use. In the United States, granulocytes are collected with the Fenwal CS3000, Fenwal CS3000 Plus, COBE (Gambro) Spectra, Haemonetics LN9000, and Fresenius AS 104. The use of hetastarch for sedimenting agent and stimulation with G-CSF and G-CSF plus dexamethasone have substantially increased granulocyte yields. Plateletapheresis is performed in the United States on the Fenwal CS3000, Fenwal CS3000 Plus, Fenwal Amicus, COBE (Gambro) Spectra, Gambro Trima Version 4, Gambro Trima Accel (Version 5), and Haemonetics LN9000. Automated red blood cell (RBC) collections are performed with the Haemonetics MCS+LN8150, Gambro Trima Version 4, Gambro Trima Accel (Version 5), Amicus, and Baxter Alyx. The RBC can be collected concurrently (with other components) in some instruments or separately in others. Plasma is collected concurrently on several instruments. Plasmapheresis for plasma only is performed on the Fenwal Autopheresis C and Haemonetics PCS2. Granulocyte yields range from 0.46 x 10(10) to 1.0 x 10(10) for unstimulated donors and 2.1 x 10(10) to 2.6 x 10(10) for donors stimulated with dexamethasone or prednisone. The use of G-CSF and G-CSF with dexamethasone has substantially increased granulocyte yields with yields of 4.1 x 10(10) to 10.8 x 10(10) reported. Platelet collection rates of 0.045-0.115 x 10(11) plt/min have been reported. Collection efficiencies of 46-85.7% have been reported. Automated (apheresis) component collection has the advantages of controlled volumes or doses of component, efficient use of the donor, multiple components from the same donor, better inventory control, and better quality control due to less manipulation of the individual components. Disadvantages of automated component collection include the use of expensive equipment and disposables, the need for specially trained machine operators, and lower capacity to collect large volumes of blood compared to whole blood donation. The use of apheresis component collection is rapidly growing to provide the best blood components in the most efficient manner.