|Year : 2017 | Volume
| Issue : 1 | Page : 38-43
Cumulative quality assessment for whole blood-derived platelets: A compliance review
Manish Raturi, Shamee Shastry, Pruthvi Raj
Department of Immunohematology and Blood Transfusion, Kasturba Medical College, Manipal University, Manipal, Karnataka, India
|Date of Web Publication||22-Mar-2017|
Department of Immunohematology and Blood Transfusion, Kasturba Medical College, Manipal University, Manipal, Karnataka
Source of Support: None, Conflict of Interest: None
Background and Objectives: Quality control (QC) results of platelet-rich plasma and buffy coat-reduced platelet concentrates (PCs) are presented with the goal to assess their compliance with published guidelines and corrective action taken for any process deviation during their manufacture. Subjects and Methods: Retrospective QC of in-house prepared whole blood-derived platelets (2009–2013) was conducted. Their cumulative results were compared to the published quality standards given by the American Association of Blood Banks, Council of Europe, and Indian guidelines. Data was analyzed using SPSS Statistics version 20. Results: A total of 36,053 PCs were prepared during the study period, and 1.43% (n = 516) was subjected to QC. The aggregate five years mean ± standard deviation (range) of product per bag were volume 58.4 ± 9.5 (37–90) mL, platelet yield 5.89 ± 1.28 (3.1–8.7) × 1010, residual leukocyte count 1.5 ± 1.2 (0.02–5.5) × 107, pH 6.67 ± 0.48 (6.0–7.3), and erythrocyte contamination 0.29 ± 0.2 (0.03-2.0) mL. Swirling was present in all the units. None of the bags showed any microbial growth. Against volume, yield, and erythrocyte contamination 90.0%, 94.3%, and 87.0% units showed compliance to the Indian standards, respectively. All the PCs had pH and leukocyte counts well within the recommended norms. Conclusions: Quality of our platelet product although suboptimal to International standards was well within the national requirements.
Keywords: Blood components, platelet concentrates, quality control
|How to cite this article:|
Raturi M, Shastry S, Raj P. Cumulative quality assessment for whole blood-derived platelets: A compliance review. Glob J Transfus Med 2017;2:38-43
|How to cite this URL:|
Raturi M, Shastry S, Raj P. Cumulative quality assessment for whole blood-derived platelets: A compliance review. Glob J Transfus Med [serial online] 2017 [cited 2020 Aug 6];2:38-43. Available from: http://www.gjtmonline.com/text.asp?2017/2/1/38/202719
| Introduction|| |
Quality is everybody's responsibility; it starts with me. It is defined as the consistent and reliable performance of services or products in conformance to the specified standards. Quality control (QC) of blood components is imperative to ensure adequate benefit of blood transfusion. Unlike other components, platelets concentrates (PCs) are “perishable” having a shorter shelf life thereby more concerns follow their demand and supply from the vision of transfusion services. Evidence-based prophylactic platelet transfusions are crucial to avoid catastrophic bleeding in patients. Dr. Scott Murphy and Frank Gardner (1969) proved that platelets could be stored at 22 ± 2°C temperature for up to three days and still maintain their hemostatic function. However, subsequent advancements like the availability of improved storage containers have enabled the provision of better quality and longer shelf life of platelets even up to seven days of storage with continuous agitation. Multiple factors influence the quality of platelets during storage. These include the preparation method, storage bag, and the ability of these bags to exchange gas across its surface. Other important factors affecting their quality are the donor-related biological variables (platelet count and/or total mass), storage temperature, type of anticoagulant used, actual PC in the bag, and their agitation. In addition, high-platelet quality would be expected to result in improved clinical efficacy determined by count increment, improved hemostasis, and lower risk for adverse reactions in recipients. Both national and international quality requirements have been implemented and are a part of the health quality management. Several approaches to perform QC of blood components have been promulgated by the American Association of Blood Banks (AABB), the Council of Europe (CE), and Director General of Health Services (DGHS) India. These are among the few that provide benchmarks for meeting the quality of blood components.
At our center, three different methods of platelet preparations are used, namely, single donor derived apheresis product, whole blood-derived platelets prepared by platelet-rich plasma (PRP-PC) method, and buffy coat (BC-PC)-reduced platelets. This study was undertaken with the goal to assess the compliance of whole blood-derived PC with the established standards and observe corrective action/s taken for any process deviation/s during their manufacture.
| Subjects and Methods|| |
Donors and procedures
Retrospective analysis of the routine QC of PC prepared from whole blood of eligible donors during the five years period (2009–2013) was undertaken at the Department of Immunohematology and Blood Transfusion supporting a 2032 bed tertiary care multispecialty center. Their trend and the cumulative QC results were analyzed and compared with the quality requirements of the AABB, CE, and DGHS [Table 1]. The protocol was approved by the Institutional Ethics Committee, prior, to the commencement of the study. The study was conducted in accordance with the principles of good laboratory practices.
|Table 1: Standards for platelet concentrates as required by the American Association of Blood Banks, Council of Europe, and Director General of Health Sciences|
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Preparation of whole blood-derived platelet concentrates
PCs were prepared from 450 ml whole blood bags within 8 h of collection using centrifuge (Thermo Fisher Scientific GmbH, Germany). PRP-PC method was routinely used since 2009 until mid-2010 from 450 mL triple bags containing CPDA-1 anticoagulant (Terumo Penpol Ltd., India). However, from the latter half of 2010, 450 mL quadruple bags containing CPD anticoagulant, with additive solution (SAGM; Terumo Penpol Ltd., India), were introduced. Thus, PC preparation was shifted to BC-PC method using automated component extractor (TACE -II; Terumo India Pvt Ltd. Chennai, India).
Department quality control protocol
The QC of PC is done once (weekly) as per our departmental standard operating procedure (SOP). The number of PC tested were 1% of the units prepared in alignment to the guidelines. According to AABB criteria, the platelet yield and pH parameters of 90% units and the sterility of 100% units should meet the prerequisites. CE requires 75% units should meet their quality standards against platelet count, residual leukocytes, and volume, whereas pH (6.4–7.4) should be met in 90% units. Interestingly, as per DGHS standards, 75% units should meet the prespecified quality requirements against platelet count, residual leukocytes, and red blood cell (RBC) contamination, whereas all 100% units should meet the criteria against pH, volume, swirling, and sterility [Table 1]. All the units subjected to QC were transfused to the patients on the same day of the testing.
Sampling and testing methods
Two to three milliliters of sample from PC was collected aseptically in a vial after thoroughly stripping the segment to ensure a representative product of the bag. The bag tubing was resealed using a sterile connecting device thereby ensuring the maintenance of a closed system. A total of 516 PCs (86 PRP-PC and 430 BC-PC, respectively) were selected and tested for the following parameters: PC volume, pH changes, platelet count, leukocyte count, erythrocyte contamination, swirling, and sterility.
Hematological values of the bag were obtained using a routinely calibrated automated cell counter (Sysmex K21, Japan). In case of aberrant results exceeding the upper limit of measurement, the samples were mixed with the equal volume of the diluent of the analyzer. The results of the diluted sample were multiplied by numerical value two. Platelet yield was calculated by multiplying the platelet count with product volume and expressed as n × 1010 per bag. Similarly, the leukocyte count was also calculated. For erythrocyte contamination, the red cell count was multiplied with the mean corpuscular volume of one red cell (80 fL) and the product volume was expressed in mL. Volume of the units was calculated by subtracting the weight of the empty bag from the original bag divided by the specific gravity of platelets (1.03). The pH estimation was done using pH strip during the period (2009–2012) and later in 2013 by pH bench meter (Eutech pH 700, Eutech Instruments Pte Ltd., Singapore). All tests were performed in accordance with our department protocol.
Swirling and sterility
Swirling in platelet units was assessed visually against the light source after a brief squeezing along the border of the bag on the day of QC and documented as “present” or “absent.” For sterility testing, samples (7–10 mL) from the blood bag were inoculated into sterile culture bottles containing biphasic “Brain heart infusion - Castaneda medium” under aseptic precautions (Level-II biosafety cabinet). These culture bottles were then sent to the department of microbiology for detection of any cultivable microbial growth.
Data was analyzed using SPSS software version 20 (IBM Inc., Armonk, NY, USA). Simple descriptive statistics were expressed as frequency distribution, mean ± standard deviation (SD), and qualitative data were expressed as percentage. We used the Student's t-test for comparative studies. P < 0.05 was considered to be statistically significant.
| Results|| |
During the study period, a total of 88,854 whole blood (450 mL) units was collected from January 2009 to December 2013. A total of 41% (n = 36,053) random donor platelets (PRP-PC - 6321 [17.5%] and BC-PC - 29,732 [82.5%]) were prepared. Of the total 36,053 PCs prepared, 1.43% (516) (PRP-PC - 86/6,321 [1.36%] and BC-PC - 430/29732 [1.44%]) were subjected for routine QA. The preparation of PCs according to the year and the percentage of units subjected for QA are given [Table 2]. The percentage conformability to the guidelines is given [Table 3].
|Table 2: Year-wise distribution of types of platelet concentrates prepared and tested for quality control|
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|Table 3: Aggregate five years mean values and percentage compliance to the standards|
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Volume of platelet concentrates
The volume of individual units was calculated and analyzed. The volume of all PCs during the study period was 58.4 ± 9.5 mL (mean ± SD) and ranged from 37 to 90 mL. The mean volume of the PRP-PC unit was 53.6 ± 7.0 mL and ranged from 37 to 70 mL. The mean volume of BC-PC was 59.3 ± 8.9 mL and ranged from 39 to 90 mL. A statistically significant difference (P = 0.0001) was observed on comparing the maximum number of BC-PC units with PRP-PC units which met the desired QC criteria against volume per unit (392/430 [91.1%] vs. 83/86 [96.5%]). Overall, 90.0% (n = 465/516) units were compliant to the Indian standards (>50 mL) and 99.0% (n = 511/516) units met the CE (>40 ml) standards.
The mean pH of all the PCs was 6.67 ± 0.48 (mean ± SD) and ranged from 6.0 to 7.3 units. The mean pH in PRP-PC (n = 86) was 6.32 ± 0.48 and ranged from 6.0 to 7.0 units, whereas the mean pH in BC-PC (n = 430) was 6.74 ± 0.42 and ranged from 6.0 to 7.3 units. On comparison, BC-PC units had higher pH than PRP-PC per bag. The difference was statistically significant (P < 0.0001). Overall, when considering pH, all the units were in conformance to the Indian standards (pH > 6.0 units/bag). With AABB and CE standards, the conformability was (70% [n = 360/516]) and (66% [n = 341/516]), respectively.
Platelet count per unit
The average platelet count during the study period was 5.89 ± 1.28 × 1010 (mean ± SD) per unit and ranged from 3.1 to 8.7 × 1010 per unit. In PRP-PC units, the mean platelet count was 5.70 ± 1.42 × 1010 per unit and ranged from 3.1 to 8.6 × 1010 per unit. In BC-PC, it was 5.93 ± 1.28 × 1010 per unit and ranged from 3.7 to 8.7 × 1010 per unit. A total of 91% (n = 78/86) PRP-PC units and 95% (n = 409/430) BC-PC units met the platelet counts >4.5 × 1010 according to the Indian standards (from 450 mL whole blood). On statistical analysis, of those meeting and not meeting the desired requirements no significant difference was observed between PRP-PC and BC-PCs (P = 0.305). Overall the minimum requirement for platelet yield of 4.5 × 1010 per bag ([DGHS] from 450 mL whole blood), 5.5 × 1010 per bag ([AABB] from 450 mL whole blood), and 6 × 1010 per single unit ([CE] from 450 mL whole blood) was achieved in 94.3% (n = 487/516), 79% (n = 409/516), and 64.0% (n = 331/516) PCs respectively.
Leukocyte count and erythrocyte contamination per unit
Leukocyte count was done on all the units of PCs. The mean residual leukocyte count of PCs was 1.5 ± 1.2 × 107 (mean ± SD) per unit and ranged from 0.02 to 5.5 × 107. In PRP-PC, it was 1.73 ± 1.7 × 107 and ranged from 0.02 to 4.4 × 107. In BC-PC, it was 1.46 ± 1.2 × 107 and ranged from 0.02 to 5.5 × 107. On comparison, BC-PC units had less mean leukocyte count than PRP-PC per product. The difference was statistically insignificant (P < 0.078). Almost all the units showed conformability for leukocyte counts to both DGHS (100%) and CE (99%). Similarly, the overall mean erythrocyte contamination calculated was 0.29 ± 0.2 mL (mean ± SD) per unit and ranged from 0.03 to 2.0 mL. The mean value in PRP-PC unit was 0.32 ± 0.2 mL and ranged from 0.03 to 2.0 mL. In BC-PC, mean erythrocyte count was 0.16 ± 0.1 mL per unit ranging from 0.03 to 0.89 mL. On comparison, BC-PC units had less mean erythrocyte contamination than PRP-PC per product. The difference was extremely statistically significant (P < 0.0001). Around 87% (n = 449/516) PCs had an acceptable erythrocyte contamination (<0.5 mL) which was conformable to both DGHS and AABB standards.
Swirling and sterility testing
Swirling was observed to be present in all the units. All the units were sterile and none showed any cultivable microbial growth.
| Discussion|| |
In bleeding patients, the utility of PCs is paramount and their hemostatic efficacy depends not just on the quantity but their quality as well. Their optimal safety greatly depends on the leukocyte content and pathogenic sterility. It has long been recognized that donor biological variables (platelet count and/or mass) as well as storage lesions all can contribute to poor platelet function which affect the posttransfusion survival. In our local population, 0.04% (n = 40/1000) donors had a predonation platelet count of <1.5 lakhs/mm 3 which could have contributed to the quality significantly. Changes in platelets fall into three broadly defined categories: Platelet activation, metabolic alterations, and platelet senescence. The parameters assessing the quality of these characteristics have to be checked at regular frequency and the results were compared with the established standards. The minimal need for testing frequency and quality checks are governed by legal provisions and professional standards. We separated 36,053 units of whole blood in five-year period subjecting more than 1% of PCs for QC and observed the compliance of our products as per the quality recommendations of AABB, CE, and DGHS, India.
Quality assessment of platelet concentrates
Once prepared, all PCs were stored at 20–24°C with continuous agitation until the time of issue. As a part of QC plan based on our departmental SOP, one percent of the units or 4 units/month was sampled (weekly once). This helped us to detect any process deviation and prepared us for a timely intervention to initiate any corrective action plan if needed. The minimum 1% criteria for the performance of QA (as per the requirements) were met. A total of 1.43% (516/36,053) units were subjected to QC during the study period [Table 2]. There is substantial heterogeneity in the utilization of either whole blood-derived or apheresis products globally, ranging from 10% to 90%, while in Europe the ratio is 50:50. At our center, PCs prepared from apheresis are very few (around 10% only) and are mostly demand based but 90% are those prepared from whole blood donations and are usually issued within 24–48 h of preparation to minimize chances of bacterial contamination and preserve their functional quality unlike five day storage and utilization of PCs practiced commonly in Europe.
Volume of platelet concentrate units
The platelets must be suspended in adequate plasma volume for their metabolism and to prevent platelet clumping. In this study, 90% of the PCs exhibited more than 50 mL plasma volume meeting DGHS requirements and >99% were in compliance to CE standards [Table 3]. However, AABB does not provide any criteria against volume of the PCs. Similar to our findings, Fijnheer et al. have shown that the mean volume values of PRP-PCs and BC-PCs were comparable. However, the range and SD in our units were wide, thus warranting standardization in their preparation methods.
pH determination is a straightforward method commonly used to report product viability and bacterial growth in PCs. Increased platelet glycolysis resulting in a fall in pH to the levels up to 6.0 in stored PCs is associated with substantial loss of viability. Acidosis adversely affects platelet behavior directly causing the cell to swell and adopt a spherical instead of discoid shape. A study by Snyder et al. reported that at the 5th day of storage, the pH of all PC was >7.0 and platelet count was above 5.5 × 1010 per bag (except for the PL-732) with the 6 rpm vertical rotator. In our study, during the first four years, pH was measured using litmus paper and from 2013 onward using pH meter. All the units tested were found within the acceptable limit (pH >6.0 units) as per DGHS standards which does not give any upper cut off limit.
Platelet count per unit
According to the DGHS  at least 75% of units tested for platelet yield (prepared from 450 mL whole blood) should be >4.5 × 1010 per unit, whereas 90% of units tested (prepared from 450 mL whole blood) should be >5.5 × 1010 per unit as per AABB recommendations and >6 × 1010 per unit (prepared from 450 mL whole blood) as per CE recommendations. Our results showed the average product yield (5.70 ± 1.28 × 1010 per unit) was consistent in 94.3% (n = 487/516) units meeting with Indian requirements. However, its compliance with AABB and CE standards was 79% (n = 409/516) and 64% (n = 331/516), respectively. In a study by Das et al. with the mean platelet yield of 5.47 × 1010 per unit (n = 267) tested, only 37.5%, 43.5%, and 71% of the units showed compliance with the CE, AABB and DGHS guidelines respectively. Fijnheer et al. reported 15% higher platelet yield in PRP-PC than BC-PC units. However, no such differences were observed in the present study and the platelet count per unit in both types of PCs was not found to be statistically significant [Table 4].
|Table 4: Comparison of various parameters for quality control of platelet-rich plasma-platelet concentrate and buffy coat-platelet concentrate|
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Leukocyte count and erythrocyte contamination per unit
Leukocytes in PCs have a detrimental effect on the storage medium, resulting in a significant drop in pH, increase in glucose consumption, lactic acid production, and LDH release during storage. They are also responsible for transfusion associated virus transmission (CMV etc.,) as well as most immune transfusion reactions affecting their quality. Fijnheer et al. reported residual leukocyte count per unit in PRP-PC was higher than the BC-PC. Similarly, our study also supported these findings thus highlighting the advantage of BC-PC over PRP-PC with regard to leukocyte contamination. Overall 100% and 99% of the units showed leukocyte count in the product to be within the acceptable limits in compliance with both DGHS and CE standards. Recommendations for erythrocyte contamination are proposed to be <0.5 mL in each bag. A total of 87% units showed conformability to both AABB and Indian standards. During instances of out of specifications where contamination of RBCs exceeded, the recommendations ([ >0.5 mL] in 13% [n = 67/516]) these units were issued as ABO group specific to the recipients.
A simpler technique to determine the quality of PCs in the laboratory is the “Swirling test.” The discoid morphology of platelets correlates with the in vivo survival and can be demonstrated by the swirling phenomenon. It only requires a trained eye (”swirl”) to generally discriminate poor quality PCs from good ones. However, the assay hardly has any dynamic range, is prone to subjectivity, and does not entirely or necessarily correlate with other platelet (quality) markers. During our study, all PCs exhibited swirling reflecting their intact functionality.
Unlike other components, PCs are at increased risk for bacterial contamination because of their storage at room temperature. The incidence of bacterial contamination of PCs has been estimated at one per 1000–3000 units. In a study at Croatia, the overall incidence of bacterial contamination of the leukocyte-reduced BC pooled platelets was 0.19% However, in our study, none of the units showed any microbial growth. Meticulous donor arm cleaning, use of potent disinfectant, preparation in an entirely closed system, and optimal processing of the product enabled us to provide PC without any cultivable microbial growth.
| Conclusions|| |
The quality of PCs prepared at our center was in harmony to the Indian standards. Nevertheless, improvement is desirable to meet the international benchmarks. Adherence to SOP, continual education of the personnel involved in the production and vigilant scrutiny of the quality indicators all contribute significantly to a robust quality management system enabling the blood components “fit for use” in the recipients.
The authors gratefully acknowledge the support and cooperation of Ms. Tensy George (Technologist) for her help in data collection during the study.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
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[Table 1], [Table 2], [Table 3], [Table 4]