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Background: One of the complications of chronic transfusions in thalassemia is the development of red cell alloimmunization., Aims: The aim of the study was to determine the frequency, specificity of red cell alloantibodies, and factors influencing alloimmunization in multiply transfused thalassemia patients., Materials and Methods: The study was carried out prospectively on beta-thalassemia patients over 10 months. Plasma samples were used for antibody screening and identification using the column agglutination technique. Patients' clinical, laboratory, and transfusion details were obtained from hospital information system and patient files., Statistical Analysis: Continuous variables were reported as median and quartile, whereas categorical variables were provided as numbers and proportions. P < 0.05 was considered statistically significant., Results: Out of 255 patients, 17 (6.6%) patients developed alloantibodies. Alloimmunized patients had significantly higher median ages at their first transfusions (1 year vs. 0.5 years; P = 0.042) than nonalloimmunized patients. Alloimmunized patients had significantly higher conjugated bilirubin ( P = 0.016) and serum ferritin ( P = 0.007). The majority of alloantibodies had specificity toward K antigen, followed by E, C, D, JKa, and JKb antigens. Alloimmunized patients received more units per year than nonalloimmunized patients (median, 30 vs. 24 units; P < 0.001). The average transfusion interval time between two successive transfusions showed a significant difference ( P < 0.001)., Conclusions: The prevalence of alloimmunization in thalassemia patients in North India is relatively low. Since most of the alloantibodies belong to Rh and Kell blood group system, extended phenotype-matched blood for Rh and Kell will be helpful in further preventing or decreasing the development of alloantibodies in multiply transfused thalassemia patients., Competing Interests: There are no conflicts of interest., (Copyright: © 2023 Asian Journal of Transfusion Science.)

Background: The new cell separators make it simple to collect single donor platelets (SDP), although the platelet yield may vary depending on the cell separator used and donor-related clinical and laboratory variables., Aims: This study aims to study the factors affecting SDP yield and corrected count increment (CCI)., Materials and Methods: This retrospective study was carried out at a tertiary care facility in northern India, over 4 years (May 2017-April 2020), data were retrieved and analyzed., Statistical Analysis: Categorical variables were presented as proportions, while continuous variables were presented as mean with standard deviation, P < 0.05 was considered significant., Results: We found a positive correlation between predonation platelet count and yield ( r = 0.243, P = 0.000). No such significant correlation was found with Hb concentration ( r = 0.025, P = 0.720), age ( r = 0.016, P = 0.820), sex ( r = -0.038, P = 0.584), and weight ( r = -0.025, P = 0.714). Maximum platelet yield and minimum time were seen with Trima. Only 39.3% (33/84) meet the 24 h CCI. The majority of patients did not meet the desired CCI could be due to the patients' clinical condition. On logistic regression, we found a significant association of 24 h CCI with product yield (odds ratio [OR] = 0.168, P = 0.015) and posttransfusion platelet count (OR = 0.454, P < 0.05)., Conclusion: The only donor-related factor that influences yield is predonation platelet count, whereas 24 h CCI may depend on the clinical status of the patient and yield., Competing Interests: There are no conflicts of interest., (Copyright: © 2023 Asian Journal of Transfusion Science.)

Background: Hepcidin-25, a polypeptide hormone, plays a major role in iron metabolism and is found to be reduced during iron deficiency; therefore, testing for hepcidin can be utilized as an indicator of bioavailability of iron. Globally, reference range values for hepcidin have been established in different communities. The aim of the present study was to establish the normal reference range values of serum hepcidin in Indian blood donors to fathom the baseline and reference point of hepcidin., Materials and Methods: A total of 90 donors fulfilling the eligibility criteria were recruited in the study consisting of 28 males and 62 females. Blood samples collected were used to execute hemoglobin (Hb), serum ferritin, and hepcidin assays. Serum hepcidin-25 isoform was detected by a commercial competitive enzyme-linked immunosorbent assay kit according to the manufacturer's instructions. Hb and ferritin were evaluated by the standard methods., Results: The mean ± standard deviation (SD) of Hb level in males was 14.62 ± 1.34 g/dL and females was 13.33 ± 0.76 g/dL. The mean ± SD of ferritin level in males was 113 ± 56.12 ng/mL and females was 62.65 ± 40.8 ng/mL. Similarly, the mean ± SD of hepcidin level in male donors was 22.18 ± 12.17 ng/mL and female donors was 10.95 ± 6.06 ng/mL. The established reference range values of Hepcidin were 6.32-46.06 ng/mL for males and 3.44-24.78 for females., Conclusion: These findings suggest that further studies with larger groups of donors are mandatory to produce reference values of hepcidin that can be précised to the whole populace in India., Competing Interests: There are no conflicts of interest., (Copyright: © 2022 Asian Journal of Transfusion Science.)

The application of flow cytometry (FC) is diverse and this powerful tool in used in multiple disciplines such as molecular biology, immunology, cancer biology, virology, and infectious disease screening. FC analyzes a single cell or a particle very rapidly as they flow past single or multiple lasers while suspended in buffered solution. FC has a great impact in the field of transfusion medicine (TM) due to its ability to analyze individual cell population and cell epitopes by sensitive, reproducible, and objective methodologies. The main uses of FC in TM are detection of fetomaternal hemorrhage, diagnosis of paroxysmal nocturnal hemoglobinuria, quantification of D antigen, detection of platelet antibody, quality control of blood components, for example, residual leukocyte counts and evaluation of CD34-positive hematopoietic progenitor cells in stem cell grafts. In recent years, FC has been implemented as an alternative method for the detection and characterization of red cell autoantibodies in autoimmune hemolytic anemia. Many workers considered FC as a very good complement when aberrant expression of various erythrocyte antigens needs to be elucidated. It has been extensively used in the resolution of ABO discrepancies and chimerism study. FC has also been used successfully in various platelet immunological studies. In the recent past, FC has been used in several studies to assess the platelet storage lesions and elucidate granulocyte/monocyte integrity and immunology. FC analysis of CD34+ stem cells is now the method of choice to determine the dosage of the collected progenitor cells. The technique is vastly used to evaluate residual leukocytes in leukodepleted blood components. We conclude that flow cytometers are becoming smaller, cheaper, and more user-friendly and are available in many routine laboratories. FC represents a highly innovative technique for many common diagnostic and scientific fields in TM. Finally, it is the tool of choice to develop and optimize new cellular and immunotherapeutic trials., Competing Interests: There are no conflicts of interest., (Copyright: © 2022 Asian Journal of Transfusion Science.)

Background: Increasing demand of single donor platelet requires blood banks to expand the donor pool. A reassessment of donor deferral criteria for plateletpheresis is required to ensure that this increasing demand is met without compromising on product quality and donor safety., Aims: (1) To list the various causes of SDP donor deferral. (2) To discuss various approaches to minimize it., Materials and Methods: Data of plateletpheresis donor deferral were collected from records retrospectively over a period of 4 years from January 2017 to December 2020., Statistical Analysis: All statistical tests were performed using IBM SPSS software for Windows version 20. Categorical variables were presented as proportions, while continuous variables were presented as mean with standard deviation, mean calculated P < 0.05 was considered statistically significant., Results: Out of the 7478 donors screened for plateletpheresis procedure, 3232 (43.2%) were deferred among which 3089 (42.5%) were male and 142 (63.1%) were female donors. Majority (96.5%) of deferral were temporary. These included low platelet count (47.4%) followed by poor venous access (22.4%) and low hemoglobin (Hb) (7.2%). Among the donors deferred for low Hb, 24.7% (58 out of 234) had Hb between 12 and 12.4 g%. Similarly, among donor deferred for low platelet count, 12% (184 out of 1532) had platelet count between 140 and 149 × 10 3 /μl., Conclusion: There is potential for increasing the number of eligible plateletpheresis donors if the present donor selection criteria were relaxed to a minimum Hb of 12 g/dl and minimum platelet count of 140 × 10 3 /μl., Competing Interests: There are no conflicts of interest., (Copyright: © 2022 Asian Journal of Transfusion Science.)

Background & Objectives: RHD gene typing is highly complex due to homology with RHCE genes. Molecular polymorphism of the RHCE and RHD genes have been characterized among various populations, but no studies have been undertaken among Indians. This study was undertaken to assess the genetic basis of RHD-negative phenotype in Indian blood donor population., Methods: Sample from a total of 200 phenotypically RhD-negative blood donors were analyzed for presence of RHD gene using polymerase chain reaction (PCR). RHD genotyping was done using three primer sets designed for exons 4 and 10 and one set for identification of pseudo (RHDΨ) gene between introns (int) 3 and 4. Amplified PCR products were analyzed by gel-electrophoresis (XY Loper, Uvitech, Cambridge) and confirmed by nucleotide sequencing (ABI 3730 xl 96 capillary system)., Results: No PCR product was found in 195/200 (97.5%) of study samples indicating homozygous gene deletion. Of the 5/200 (2.5%) showing RHD gene polymorphisms, 4/200 (2%) were positive for presence of exon 10 only (RHD-CE-D hybrid). RHDΨ gene was not detected in any of the samples tested. One sample showed presence of all three tested regions and was negative for RHDΨ gene., Interpretation & Conclusions: RHD gene deletion was found to be the most common cause of an RHD-negative phenotype while RHDΨ gene was, reported to be present in up to 39 per cent of various ethnic populations, but was not detected. RHD-CE-D hybrid gene (found in 2.5% individuals) is important for predicting the requirement of Rh prophylaxis during the antenatal period.

Competing Interests: There are no conflicts of interest.

Background: Enzyme-linked immunosorbent assay (ELISA) used for screening blood donors for transfusion transmitted infections (TTIs) can sometimes fail to detect blood donors who are recently infected or possessing the low strength of pathogen. Estimation of a grey zone in ELISA testing and repeat testing of grey zone samples can further help in reducing the risks of TTI in countries where nucleic acid amplification testing for TTIs is not feasible., Materials and Methods: Grey zone samples with optical density (OD) lying between cut-off OD and 10% below the cut-off OD (cut-off OD × 0.9) were identified during routine ELISA testing. On performing repeat ELISA testing on grey zone samples in duplicate, the samples showing both OD value below grey zone were marked nonreactive, and samples showing one or both OD value in the grey zone were marked indeterminate. The samples on repeat testing showing one or both OD above cut-off value were marked positive., Results: About 119 samples (77 for hepatitis B virus [HBV], 23 for human immunodeficiency virus [HIV], and 19 for hepatitis C virus [HCV]) were found to be in grey zone. On repeat testing of these samples in duplicate, 70 (58.8%) samples (45 for HBV, 12 for HIV, and 13 for HCV) were found to be reactive. Six (5%) samples (four for HBV, one for HIV, and one for HCV) were found to be indeterminate., Conclusion: Seventy donors initially screened negative, were found out to be potentially infectious on repeat grey zone testing. Thus, estimation of grey zone samples with repeat testing can further enhance the safety of blood transfusion.

Background and Objectives: Platelets are a source of numerous growth factors which facilitate repair and healing. Thus platelet rich plasma has been increasingly used as a treatment modality in the field of reconstructive surgeries for wound healing. This preliminary study was carried out to explore whether platelet growth factors from platelet rich plasma could be used for enhancement of split thickness skin graft survival., Materials and Methods: Twenty patients (13 males and 7 females) requiring split thickness skin graft for various clinical reasons were enrolled in the study. Platelet rich plasma was collected by apheresis and frozen at -80° C. It was thawed at room temperature immediately before its intended application. PRP was applied only on one half of the wound, while another half served as control. Patient was followed for 6 weeks. The effect was assessed at first dressing in terms of graft uptake and subsequently as time taken for complete healing., Results: There was 100% uptake of the graft in the area where platelet rich plasma was applied. In the control area, there was complete graft loss in 4 cases, partial loss in 7 cases and complete uptake in 9 cases., Conclusion: This study demonstrated promising results on application of PRP to split thickness skin grafts. Further randomized studies with greater sample size may be undertaken to establish platelet rich plasma as a validated treatment modality.

Background: Thrombopoietin (TPO) is the key hematopoietic growth factor regulating the production of platelets from bone marrow megakaryocytes and maintaining platelet hemostasis. This study was done to find any relationship between the levels of thrombopoietin and the severity of disease in patients with aplastic anemia., Materials and Methods: Serum samples were collected from 52 patients with a confirmed diagnosis of aplastic anemia and 45 normal healthy blood donors of both sexes over a period of 2 years, and TPO was estimated by using commercially available TPO-specific-enzyme-linked immunosorbent assay., Results: The median TPO level of 1190 pg/ml (range 625-7651 pg/ml) in aplastic anemia patients was significantly higher than the median TPO level of 121.1 pg/ml (81.25-237.7 pg/ml) in normal healthy blood donors (P = 0.000). No significant difference was observed in TPO levels of male and female patients (P = 0.453). The median TPO concentrations observed in very severe aplastic anemia, severe aplastic anemia, and nonsevere aplastic anemia were 2765 pg/ml (range 625-6451 pg/ml), 1190 pg/ml (range 672.1-7651 pg/ml), and 1111.5 pg/ml (range 761.1-2289.2 pg/ml), respectively. TPO in patients of very severe aplastic anemia was significantly higher than patients of nonsevere aplastic anemia (P = 0.043), with no significant relation among rest of the groups., Discussion: TPO levels in aplastic anemia patients were significantly higher than in healthy blood donors; however, in aplastic anemia patients TPO levels were significantly higher only in patients with very severe disease.

Introduction: Young people, who tend to be healthy, idealistic, and motivated, are an excellent pool of potential voluntary unpaid blood donors. Recruiting and retaining young blood donors improves the long term safety and sufficiency of a country's blood supply. Knowledge, attitude, and beliefs about Human immunodeficiency virus (HIV) should play an important role in prevention of disease transmission., Materials and Methods: This study was a questionnaire based survey, conducted to explore the levels of knowledge, attitude, and beliefs about HIV in young college student blood donors., Results: The results showed that the proportion of participants with comprehensive knowledge of HIV prevention and transmission was lesser than expected. Increase in education level and male gender was found to be significantly associated with high HIV-related knowledge. The responses on the different aspects of HIV-related attitude were also varied and there is still stigma associated with Acquired Immunodeficiency Syndrome (AIDS) even in the educated groups., Discussion: There was a spectrum of myths and misperceptions emphasizing the need of education that recognizes the social context of attitude towards HIV. Results from this study may contribute to the development of appropriate educational and training material for this group of donors which in turn, may assist in achieving the elusive goal of safe blood supply in future.

Background: Direct antiglobulin test (DAT) is the most common test done in immunohematology lab, which detects immunoglobulin and fragments of complement attached to the red blood cells. These coated red blood cells are difficult to accurately phenotype, which may be required for selection of appropriate unit of red blood cells for transfusion., Aims: We have studied the efficacy of various elution methods in removing the antibodies coating the red cells and their impact on different blood group antigen activity., Materials and Methods: Patient samples sent for serological evaluation of autoimmune hemolysis were included in the study. DAT and Indirect antiglobulin test (IAT) were performed using gel cards (ID system, DiaMed Switzerland). Antibody coated red cells, either by in-vivo or in-vitro sensitization, were used to assess the outcome of three elution methods., Results: Out of 93 DAT positive samples already sensitized in vivo, 28 (30 %) samples became DAT negative post elution using either of three methods, while 36 (38.8%) showed reduction in strength of reaction, whereas in 29 (31.2%) there was no change in strength of reaction. Similarly, out of the 17 samples prepared by in vitro sensitization, 12 samples became completely negative after glycine-HCl/EDTA elution, 9 and 5 samples became negative after heat elution and chloroquine diphosphate elution methods, respectively., Conclusion: On comparative analysis glycine-HCl/EDTA elution method was better than the other two methods and can be used for eluting immunoglobulins from intact red cells.

Very few studies in humans have investigated the laboratory evidences suggestive of transfusion-associated immunologic changes. In this prospective study, we examined the effects of perioperative blood transfusion on immune response, by measuring various cytokines production, namely, interferon-gamma (IFN-γ), interleukin-10 (IL-10), and Fas Ligand (FasL). A total of 40 patients undergoing neurosurgery were randomly allocated into four groups: (a) no transfusion, (b) allogeneic non-leukofiltered transfusion, (c) prestorage leukofiltered transfusion, (d) autologous transfusion. Samples were collected before operation (day 0) and postoperative days (post-op) 1, 7, and 14. IFN-γ and IL-10 production capacity was measured in supernatant after whole blood culture and serum FasL levels in patients' sera using commercially available ELISA kits. Change in ratios (cytokine value after PHA stimulation/control value) of IFN-γ and IL-10 and percentage change from baseline for serum FasL levels across different transfusion groups during the sampling period were calculated. There was an increase in IL-10 production in patients receiving allogeneic non-leukofiltered transfusion on days 1 and 7 (mean ratio 2.22 (± 2.16), 4.12 (± 1.71), 4.46 (± 1.97) on days 0, 1, and 7, respectively). Similarly there was a significant (P<0.05) decrease in IFN-γ production in patients who received allogeneic non-leukofiltered red cell transfusion on post-op days 1, 7, and 14 (mean ratio 6.88 (± 4.56), 2.53 (± 0.95), 3.04 (± 1.38) and 2.58 (± 1.48) on day 0, 1, 7, and 14, respectively). Serum FasL production was increased across all patients till 7th day except for 'no transfusion' group and this increase was most significant in the non-leukofiltered group. We conclude that one time transfusion leads to quantitative changes in levels of these cytokines largely through interplay of Th2/Th1 pathways in allogeneic nonleukofiltered blood transfusion; however, soluble mediators like FasL which are also present in autologous and leukofiltered blood products may contribute toward minor immunologic effect in these settings.

Background: Despite the wide range of methods available for measurement of hemoglobin, no single technique has emerged as the most appropriate and ideal for a blood donation setup., Materials and Methods: A prospective study utilizing 1014 blood samples was carried out in a blood donation setting for quality evaluation of four methods of hemoglobin estimation along with cost analysis: Hematology cell analyzer (reference), HCS, CuSO4 method and HemoCue., Results: Mean value of HemoCue (mean +/- SD = 14.7 +/- 1.49 g/dl) was higher by 0.24 compared to reference (mean +/- SD = 13.8 +/- 1.52 g/dl) but not statistically significant ( P > 0.05). HemoCue proved to be the best technique (sensitivity 99.4% and specificity 84.4%) whereas HCS was most subjective with 25.2% incorrect estimations. CuSO4 proved to be good with 7.9% false results. Comparative cost analysis of each method was calculated to be 35 INR/test for HemoCue, 0.76 INR /test for HCS and 0.06-0.08 INR /test for CuSO4., Conclusion: CuSO4 method gives accurate results, if strict quality control is applied. HemoCue is too expensive to be used as a primary screening method in an economically restricted country like India.

Background & Objectives: Contaminating white blood cells (WBCs) in stored platelet concentrates (PC) are the main source of pro-inflammatory cytokines including interleukin-6 (IL-6), interleukin- 8 (IL-8) and tumour necrosis factor-alpha (TNF-alpha) that are implicated in transfusion reactions. We compared the levels of these cytokines in stored platelet preparations prepared by two methods. Effect of pre-storage leucofiltration on these cytokine levels was also studied., Methods: Twelve units of pooled PCs were prepared by platelet rich plasma (PRP) method and buffy-coat (BC) method each and stored for 5 days. IL-6, IL-8 and TNF-alpha levels were measured in platelet supernatants on day 0, 1, 3 and 5 of the storage using commercially available immunoassays. Pre-storage leucofiltration was done in 4-pooled units of PRP-PC and cytokine levels compared with unfiltered PCs., Results: Median IL-6 levels increased from day 0 to day 5 in both PRP-PC and BC-PC. In PRP-PC, IL-8 increased from <3 pg/ml on day 0 to 817 pg/ml on day 5, while in BC-PC the corresponding levels were 10 and 346.5 pg/ml, respectively. No significant increase in levels of TNF-alpha was observed in BC-PC during storage period, while levels increased significantly in PRP-PC on day 1 only. There was no significant change in the levels of all three cytokines in leucofiltered PCs over 5 days of storage., Interpretation & Conclusion: Findings of our study showed that method of preparation and WBC content are the critical factors in determining the cytokine levels in stored PCs.

Background and Objectives: IgA deficient patients are at risk of severe anaphylactic reaction on being transfused blood and blood products, and its prevalence varies in different parts of the world. No data are available from India. We did a blood donor survey to look for prevalence of IgA deficiency in north India., Methods: A sensitive enzyme linked immunosorbent assay developed in-house was used to detect IgA deficiency in a total of 3818 blood donors. Complete IgA deficiency was defined as value less than 5 mg/dl whereas partial IgA deficiency was defined as value between 5-30 mg/dl., Results: Of the 3818 blood donors screened, 3640 (95.3%) were males with a mean age of 31.2 yr. No donor was found to have complete IgA deficiency; however, 257 (6.7%) had partial IgA deficiency., Interpretation and Conclusion: Our study shows that IgA deficiency is rare in north India.

Background & Objectives: Platelet storage lesions (PSL) are detrimental to the post transfusion functional capacity of platelets. As EDTA enhances the storage-induced changes, changes in platelet indices with and without EDTA incubation are promising new tests to monitor the PSL. The present study was undertaken to monitor the PSL in 40 units of pooled platelet concentrates harvested by platelet rich plasma (PRP) method and stored for seven days in second generation platelet storage containers using conventional in vitro tests and platelet indices., Methods: Morphological changes in platelets were monitored by automated haematological cell counter for platelet count and mean platelet volume (MPV). Samples were incubated with K2EDTA for 1 h and platelet indices were repeated on the EDTA incubated samples. Difference between pre-and post-EDTA incubation of platelet count (dPLT) and MPV (dMPV) were calculated. Metabolic parameters such as pH, pO2, pCO2 and ATP were measured., Results: There was no significant change in the indices without EDTA during storage, however, after EDTA incubation, significant changes were noted in dPLT and dMPV. The mean dPLT on day 0 was 75.15 x 10(3)/microl decreasing to 44.4 x 10(3)/microl on day 7, while dMPV from 0.76 fl on day 0 increased to 1.34 fl on day 7 (P < 0.05). Metabolic parameters showed a significant decrease in pH and pCO2 concurrent with increasing pO2 during storage (P < 0.05). Average ATP level on day 0 was 21.09 micromol/dl falling to 10.59 micromol/dl on day 7., Interpretation & Conclusion: The results indicate that storage induced lesions take place even in second generation platelet storage containers under recommended conditions of storage. Platelet indices especially after EDTA incubation are useful in monitoring PSL. However, how much these changes contribute to poor post transfusion survival and haemostatic function of platelets need to be investigated.