Plateletpheresis: the Process, Devices, and Indicators of Product Quality
Plateletpheresis: the Process, Devices, and Indicators of Product Quality
Journal of Life Science. 2014. Sep, 24(9): 1030-1038
Copyright © 2014, Korean Society of Life Science
This is an Open-Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License ( which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
  • Received : June 20, 2014
  • Accepted : July 25, 2014
  • Published : September 30, 2014
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About the Authors
철수, 장
성인, 김
현경, 김
창오, 권
병원, 김
동찬, 김
윤석, 김
기종, 이
재기, 류

Platelet products are used to treat hemorrhagic or platelet dysfunction diseases. Plateletpheresis involves collecting the platelet components of blood using an apheresis blood-collection system. Various indicators are available for evaluating the qualities of the apheresis platelets. The productivity of platelet collection is evaluated through both the collection efficiency and collection rates. Platelet storage quality can be evaluated in vitro using several indicators, including visual appearance, metabolic activities, volume, platelet count, white blood cell count, microparticles, and various platelet activation markers. Platelet activation markers have been used as indicators of storage quality in various studies. Post-transfusion platelet quality can be evaluated based on the corrected count increment and the percentage of platelet recovery. Although various studies have investigated the aspects of plateletpheresis, no article has systemically presented assessments of the platelet products obtained from different plateletpheresis devices. The present study provides a review of plateletpheresis, including the specifics of the process, the types of devices employed, the platelet quality, the overall efficacy, and the evaluation indicator qualities. Furthermore, the differences in functionality among the different apheresis devices are discussed. Although adverse reactions to the citrate anti-coagulant have been reported, apheresis processing may provide a safer option for donors who are at a high risk for presyncopal or syncopal reactions related to whole blood collection.
Apheresis is the process of collecting blood components such as plasma, platelets, red blood cells, and granulocytes from donor blood. The term “apheresis” is derived from the Latin word “aphaeresis”, which means “withdrawal”. Apheresis is accomplished using an apheresis instrument termed a cell separator. Whole blood from the donor is separated by the device through centrifugation, based on the specific gravity and/or filtration parameters. The selected component of the blood is retained, while remaining blood components are returned to the donor through automated circulation. The processing time is approximately 1-2 hr [7 , 24] . Apheresis instruments are used with various settings, depending on the specific needs of different cases [8] . Cell separators employ either continuous flow centrifugation, which uses 2 venipunctures to collect blood, or intermittent flow centrifugation, which returns the unwanted blood components to the donor after it is temporarily collected in the cell separator. In practice, apheresis presents several obstacles. For example, apheresis requires a trained cell separator operator; is difficult to apply in high-capacity, emergency settings; requires large initial expenditures; and can result in adverse reactions among donors, such as paresthesia, tingling, seizure, and muscle cramps. These adverse reactions result from the use of citrate anticoagulants during apheresis. Nonetheless, the collection and use of blood components through apheresis has increased over the past few decades. The transfusion of blood components collected through apheresis has many benefits. Indeed, it provides larger quantities of components than does whole blood separation, produces consistent product volumes, makes efficient use of the same donor, reduces the donor’s physical stress and blood cell count recovery times, and decreases the risk of bacterial contamination. By eliminating the recipient’s exposure to multiple donors, apheresis additionally reduces the risks of human leukocyte antigen (HLA) alloimmunization and transfusion-transmitted diseases [3 , 8 , 15 , 24 , 41 , 63] .
Platelets are anucleate blood cells (150-450×10 3/μ l) that form a platelet plug by adhesion and aggregation, thereby contributing to hemostasis [14] . Platelets are transfused for (i) treating hemorrhage due to severe thrombocytopenia, (ii) increasing platelet counts that are less than 20,000 per μl, or (iii) treating platelet dysfunction disease [40 , 55] . Plateletpheresis is the process of collecting the platelet component through apheresis. Various studies have investigated the aspects of apheresis platelet products, including quality evaluation, efficacy, the comparative performance of different devices, and methods designed for different patient symptoms. However, to our knowledge, no article has systematically presented assessments of the quality and efficacy of platelets obtained from different plateletpheresis instruments. In the present review, we address the quality and efficacy of apheresis platelets.
Current Status of Methods and Results
- Donor screening for plateletpheresis
With the exception of the donation interval, the requirements for apheresis and whole blood donation are the same. Apheresis donors must meet the requirements for whole blood donation and additionally satisfy criteria that are particular to the selected apheresis. Donor screening is required to ensure a safe transfusion for the recipient. Prospective donors must complete several steps before actual blood donations, including physical examination, a donor history questionnaire, and testing for transmissible diseases. The American Association of Blood Banks (AABB) recommends that prospective donors receive physical examinations including an assessment of weight: hemoglobin, hematocrit, ABO and Rh typing: and inspection for marks from intravenous drug use. Tests are performed for the presence of syphilis, human immunodeficiency virus (anti- HIV-1/2 and HIV-1 RNA), hepatitis C virus (anti-HCV and HCV RNA), hepatitis B virus (HBsAg and anti-HBc), human T-cell lymphotropic viruses I and II (anti-HTLV-I/II), Trypanosoma cruzi (anti- Trypanosoma cruzi ), and West Nile virus (West Nile virus RNA). The Korean Society of Blood Transfusion recommends the tests for ABO and Rh typing, ABO subtyping, ALT, Total Protein, Antibody Screening Test, HBV (HBs Ag, HBV-NAT), HCV (anti-HCV, HCVNAT), HIV (anti-HIV, HIV-NAT), anti-HTLV, anti-Syphilis, and anti-Malaria for the donated blood.
Further, the platelet counts of plateletpheresis donors must be >150×10 3 per μl [32] . Individuals can donate 3 days after ingesting aspirin-related medications.
- Plateletpheresis instruments
Plateletpheresis instruments include the Fenwal CS3000 and Fenwal CS3000+ (Baxter Healthcare Corp., Round Lake, IL, USA); Fenwal Amicus (Fenwal, Inc., Lake Zurich, IL, USA); Cobe Spectra (Terumo BCT, Lakewood, CO, USA); Fresenius AS104 and Fresenius COM.TEC (Fresenius AG, Bad Homburg, Germany); Trima (Version 4) and Trima Accel (Terumo BCT, Lakewood, CO, USA); and Haemonetics MCS LN9000 and MCS+LN3000 (Haemonetics Corp., Stoughton, MA, USA). These different devices have been compared according to a variety of measures, including platelet yield, collection efficiency, and collection rates [3 , 8 , 9 , 10 , 58] .
- Platelet yield
The platelet volume collected by plateletpheresis ranges from 150 to 300 ml and 75% of the collection must meet the standard of 3×10 11 platelets/unit. Platelet yields in excess of 6×10 11 present several advantages. For example, when large quantities of platelets can be collected from donors who have been matched with patients according to HLA type, the collected blood can be divided into two parts and used [5] . Burgstaler et al. [9] reported that plateletpheresis with a double-needle Amicus provided more double units than did Cobe Spectra double-needle version 5 leukoreduction systems, Cobe Spectra double-needle version 7 leukoreduction systems, or Fresenius AS 104. The frequency of double units is 65% when Amicus or Trima Accel is used [9 , 10] . If pre-donation platelet counts are high, platelet yields are generally high as well.
- Reduction of white blood cell count
If a recipient’s white blood cells (WBCs) are exposed to donor WBCs, allogenic immune reactions may result in complications. In such cases, the leukoreduction process is employed to reduce the complications that were induced by the transfusion. The choice of leukoreduction is motivated by several major goals: reducing febrile non-hemolytic transfusion reactions, diminishing HLA sensitization and platelet refractoriness, reducing the rate of cytomegalovirus infection, and reducing the rate of transfusion-associated graft versus host disease [33 , 48] . Newly developed apheresis instruments have built-in leukoreduction systems that eliminate the need for further, post-apheresis reduction of leukocytes counts. Leukoreduced blood products are required to meet AABB, which indicate that WBC counts per unit must be <5×10 6 .
- Collection efficiency, collection rates, and WBC count
Evaluations of platelet collection performance are based on collection efficiency (CE) and collection rate (CR). CE compares the number of platelets in the collection bag with the number of platelets that pass through the device. Is calculated according to the following formulas:
  • CE = platelet yields in collection bag (1011) ×100/average platelet count × blood volume processed.
  • Average platelet count = (pre-apheresis platelet count + post-apheresis platelet count)/2
  • Blood volume processed = total blood volume processed − volume of anticoagulant
CR is defined as the platelet yield per minute of processing time. However, it does not reflect the time that is required to collect platelets. To estimate the time to collect required quantities of platelets, it is necessary to have the CRs for the instruments that are being used. CR is determined by the following formula:
  • CR = platelet yields in a collection bag (1011)/min of processing time
Various studies have compared the function and characteristics of different apheresis instruments. Table 1 summarizes previous reports of CE, CR, and WBC for different apheresis equipment. According to these reports, the single- needle Amicus and single-needle Trima Accel have the highest CE and CR [3 , 8 , 9 , 10 , 58] . Flesch et al. [21] reported that the double-needle Amicus and the single-needle Trima Accel showed similar platelet yields and donation times.
Collection efficiency (CE), collection rates (CR), and white blood cell (WBC) content for samples obtained from different plateletpheresis instruments
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AS104: Fresenius AS104 double-needle, AMDN: Amicus double-needle, V7-LRS: Cobe Spectra double-needle version 7 leukoreduction system, V5-LRS: Cobe Spectra double-needle version 5 leukoreduction system, AMSN: Amicus single-needle, TASN: Trima Accel single-needle, CT4DN: COM.TEC version4, CS3000+: Fenwal CS3000+, N: number of donation, PLT yield: median platelet count of plateletpheresis product, CE: median collection efficiency, CR: median collection rates. [ ]: reference number
- Quality evaluation indicators of apheresis platelet
Platelet products that are stored at room temperature (20-24℃) with agitation may remain viable for 5 days [32] . The storage time allowed by the US Food and Drug Administration (FDA) was increased to 7 days in 1984, but returned to 5 days in 1986 because of events of bacterial transmission by transfused platelets. Slichter et al. [55] reported in vivo platelet recovery and survival in a platelet product obtained from MCS+ LN 9,000 and stored for 8 days. The quality of platelets that are stored after donation is influenced by many factors, including differences in blood component processing, gas permeability of the storage bag, anticoagulants, agitation, and storage temperature [4 , 38 , 56] . Platelet quality can be evaluated in vitro using several parameters such as visual appearance, metabolic activity, volume, platelet count, and WBC count per unit. After transfusion, platelet quality can be evaluated using corrected count increment (CCI) and percentage platelet recovery (PPR) [27 , 54] .
- Visual appearance
The visual appearance of platelets, such as evidence of swirling, is used to evaluate platelet functionality [34] . Swirling is caused by the reflection of light when a discoid platelet is exposed to a light source. Generally, platelet concentrate with a pH of 6.7-7.5 will show swirling when the platelets retain their discoid shape in vitro at the time of transfusion, and are expected to be functional in vivo [6 , 34] . Singh et al. [54] found that apheresis-platelet concentrate units from the CS3,000+ instrument showed better swirling and platelet counts than platelet rich plasma-platelet concentrates; however, Mintz et al. [37] confirmed that spherical platelets may revert to the discoid form, and that non-discoid platelets may retain suitable functionality post-transfusion.
- Metabolic activity
Metabolic activities of apheresis platelets are mainly measured using pH, pCO 2 , pO 2 , bicarbonate, glucose, lactate, adenosine triphosphate (ATP), and hypotonic shock response (HSR).
The AABB requires that the pH of the platelet products remains ≥6.2 during storage with agitation at 20-24℃. During storage, pH decreases as a result of increased lactic acid production due to glycolysis. At pH 6.0, the platelets irreversibly transition from the discoid to the spherical form [39] . Accordingly, pH has been identified as an indicator of stored platelet functionality. However, Tudisco et al. [59] found that pH had little utility for the quality control of platelets produced by MCS + LN 3,000, Amicus, and CS-3,000+ apheresis instruments. Most platelet products obtained from apheresis collections have a pH ≥6.2 on day 5 of storage, and the pH may even exceed 7.0 at 7 days of storage [18 , 20 , 26 , 45 , 55] .
Assessments of pCO 2 , pO 2 , and bicarbonate changes in apheresis platelets showed that pCO 2 and bicarbonate were both significantly reduced in all platelet concentrates collected using MCS+ LN9,000 or Trima instruments. However, differences in pO 2 were observed to increase or decrease, depending on the sample. Results were not significant in platelets from Trima that had been stored 7-9 days. Furthermore, in apheresis platelets, glucose levels decrease and lactate significantly increases during storage [18 , 20 , 26 , 45 , 55] .
ATP is the major energy source for platelet survival. Naturally, ATP also decreases significantly during storage [18] . ATP was reduced in platelet concentrates collected using Trima [18 , 20 , 45] .
HSR measures the capacity of platelets that have been restored to normal morphology after hypotonic shock due to the addition of hypotonic solution. HSR is regarded as a predictor of platelet functionality [20 , 46] . The HSR of apheresis platelets decreases as storage times increase [20 , 26 , 45 , 55] .
- Microparticles (MPs)
MPs are membrane vesicles that originate primarily from platelets (PMP) or endothelial cells (EMP). The level of PMP generally increases in plateletpheresis concentrates during storage because PMPs are released by activation [43 , 47 , 52 , 61] . The aim of plateletpheresis is to reduce HLA sensitization through leukodepletion: however, platelets that are collected primarily from plateletpheresis contain MPs, including EMP, which have high densities of HLA antigen [47 , 50] . Therefore, improvements in apheresis technology may be necessary to reduce MP loads.
- Platelet activation
Platelets play 2 key roles in hemostasis: platelet adhesion to exposed subendothelial membrane and platelet aggregation among activated platelets result in platelet plug formation. Negatively charged platelet surfaces promote the activation of coagulation factors II and X [8 , 39] . Various studies have investigated platelet activation, which has been used to evaluate platelet quality during processing and storage of plateletpheresies products. Activated platelet markers that have been used in studies of plateletpheresis include P-selectin (CD62P) [2 , 27 , 30] , CD63 [20 , 22 , 47] , glycoprotein (GP)Ibα [25 , 30 , 36] , CD40 ligand (CD40L, CD154) [12 , 13 , 19 , 28 , 62] , and coated platelets [1 , 11 , 57] .
P-selectin (CD62P) is stored in granules of endothelial cells and platelets. It is expressed on the surface of activated cells, where it functions as a cell adhesion molecule [2 , 30] . During storage, increased P-selectin expression on activated platelet surfaces triggers rapid clearance of transfused platelets from circulation. Comparisons of platelet concentrates obtained from different apheresis instruments have demonstrated that the double-needle Amicus has higher rates of P-selectin-positive platelets than does the single-needle Trima Accel: this suggests that increased P-selectin expression may result from delayed processing times within the apheresis device [27 , 30] .
GPIbα is a platelet adhesion molecule that interacts with von Willebrand factor bound at damaged sites of blood vessels [25] . GPIbα may function as a platelet activation marker, as does P-selectin [30 , 36] . GPIbα of activated platelets is redistributed from the platelet surface to the internal membrane during conventional storage: as storage time increases, GPIb levels of platelet surface decrease. During storage of the platelet concentrates, platelet products from the Haemonetics PCS-Plus change less than those from platelet- rich plasma [36] .
CD63 is a 53-kDa lysosomal membrane glycoprotein that is translocated to the plasma membrane after platelet activation. It is considered to be a platelet activation marker [23] . Gutensohn et al. [22] reported that the CD63 levels of platelets collected using Amicus, Cobe Spectra, and Trima devices increased after storage for 5-8 days [20 , 47] .
CD40L, a member of the tumor necrosis factor family, is a trimeric transmembrane protein that is mainly contained within platelets [12 , 28] . CD40L is generally sequestered inside resting platelets. However, it translocates to the cell surface after activation, and is subsequently cleaved from the platelet surface to generate soluble CD40L (sCD40L). Therefore, increased sCD40L levels can provide a marker of platelet activation [12 , 13 , 19 , 28 , 62] . Platelet products that are collected with MCS+, Trima Accel, and Amicus instruments show increased sCD40L levels during storage [19 , 28 , 62] . Further, the sCD40L levels of platelet products that are collected using the Amicus or Gambro instruments do not differ substantially the sCD40L levels of platelet products that are prepared from whole blood. Coated platelets provide another marker of platelet activation. Coated platelets are activated by collagen and thrombin, representing approximately 30% of the total platelet population [1 , 11] . The surfaces of coated platelets contain procoagulants [1 , 11] . The percentage of coated platelets that is obtained from whole blood and apheresis products decreases as post-production storage time increase. The plateletpheresis method has also been observed to influence platelet activation; in addition to the storage phase itself, the plateletpheresis process conducted with the Trima Blood Collection System decreases the number of coated platelets [11 , 57] . Table 2 presents the changes in metabolic activity during apheresis platelet storage [11 , 55 , 59] .
Changes in the function and metabolic activity of apheresis platelets after storage
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Changes in the function and metabolic activity of apheresis platelets after storage
- CCI and PPR
When evaluating the in vivo recovery and survival of post-transfusion platelets, CCI and PPR are routinely used to determine the response to platelet transfusions in vivo , as well as to assess platelet transfusion refractoriness. CCI and PPR are calculated according to the following formulas [27 , 44] .
  • CCI = (platelet increment/μl) × (body surface area in m2)/number of platelets transfused (×1011)
  • PPR = (platelet count increment/μl) × blood volume (ml) × 103/number of platelets transfused
Successful platelet transfusion is defined as a CCI greater than 7,500 platelets × m 2 /μl and a PPR greater than 30% within 1 hour of transfusion [44] . In a quality evaluation of apheresis platelet products, Pandey et al. [44] found that use of the COM.TEC instrument provided an optimal response to platelet transfusion, in terms of both CCI and PPR values. However, Julmy et al. [27] reported that the transfusion efficacy of the double-needle Amicus was significantly lower than that of the single-needle Trima Accel. Julmy et al. [27] considered this finding to be a consequence of high platelet concentrations or increased platelet activation, showing that Amicus is not suitable for high-yield plateletpheresis (≥6.0×10 11 ). However, CCI and PPR results are sensitive to the recipient’s medical state and the apheresis process [51] . It has been observed that alloimmunized patients have difficulty managing their thrombocytopenia after platelet transfusion, and are refractory to fresh allogeneic platelets [42 , 60] . Additionally, some studies have found decreased CCI values using platelet additive solution (PAS), and further, it has been observed that all patients who were refractory to fresh allogeneic platelet transfusion had successful CCI results to autologous cryopreserved platelets [16 , 29 , 60] . Table 3 presents the transfusion efficacy of apheresis platelets according to different approaches to apheresis, which showed that the successful platelet transfusion in all apheresis instruments.
Transfusion efficacy after transfusion of apheresis-platelet concentrates
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AMDN: Amicus double-needle, TASN: Trima Accel single-needle Cobe Spectra: Cobe Spectra version 4.0autologous: cryopreserved in ThromboSol and 2% dimethyl sulfoxide [ ]: reference number
- Effect of PAS
When processing the platelet component, the use of a PAS to suspend the platelet concentrates (instead of plasma) may reduce the incidence of adverse reactions due to plasma [17 , 53] . In a quality comparison of extended platelet storage (up to 9 days) using the different Trima systems, it was found that platelets collected in PAS were more likely to retain acceptable metabolic and cellular characteristics than platelets collected in plasma-stored apheresis units [26] . However, some studies have reported that platelets suspended in PAS show significantly lower CCI than platelets suspended in plasma [16 , 29] . Table 4 presents details of the changes in metabolic activity of apheresis platelet according to the use of different additive solutions when the same device is used [18 , 26 , 45] . PAS can affect the functionality of apheresis platelets during storage.
Changes in metabolic activity of apheresis platelet according to the use of different additive solutions
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Trima: Trima AccelM: pathogen reduction technology (PRT) treatment with the Mirasol PRT system C: Mirasol PRT system untreated control [ ]: reference number
- Donor adverse reactions
In the apheresis collection process, the most common donor adverse reactions are related to citrate toxicity [31 , 35] . Symptoms including tingling, paresthesia, and muscle cramps can result from citrate reactions [63] . Benjamin et al. [5] reported that plateletpheresis using an Amicus instrument required larger quantities of acid citrate dextrose than did plateletpheresis using a Cobe Spectra instrument. Benjamin et al. [5] further reported than use of the Amicus instrument resulted in greater rates of citrate toxicity among apheresis donors. However, citrate-induced adverse events that result from apheresis processing can be treated with calcium supplementation or reductions in the quantity of anticoagulant that is employed. Compared with whole blood collection, blood collection using apheresis may be a safer option for donors who have risk factors for presyncopal or syncopal reactions related to whole blood collection [3 , 63] . Relevant risk factors include younger age, female gender, and small total blood volume.
Plateletpheresis has become more popular because single- donor platelet products reduce the incidence of adverse reactions and increase the efficiency of blood usage. Various studies have evaluated the quality of platelet products during apheresis platelet storage. Quality can be assessed using platelet functionality markers such as visual appearance, metabolic activity, MPs, platelet activation, CCI, and PPR. In particular, various markers of platelet activation have been developed and used for studies related to cell activation. The quality of platelets can also be influenced by the processing method and additive solutions. Additionally, apheresis may provide substantial benefits for donors who have risk factors for presyncopal or syncopal reactions related to whole blood collection.
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