Scientific Journal of

  Abstract

 Background and Objectives

  Blood group antigens include more than 260 antigens on the erythrocyte cell membrane, forming 24 different blood group systems. These can be categorized in 11 major systems which account for 172 of the antigens. Incompatibility of some of these antigens, especially in multiple transfused patients, results in hemolytic reactions. Although intermediate thalassemics need fewer blood transfusions than major ones, alloimmunization is more frequent in the former. This can be due to the different frequenct rates of blood group phenotypes in intermediate thalassemics as compared with healthy people.

  Materials and Methods 

  Red blood cell antigen typing was done in 39 intermediate thalassemics and 150 healthy blood donors for Rh, Kidd, MNSs, Lutheran and Duffy blood systems. K and M antigens had different frequency rates in both groups (p<0.01, p<0.0005) showing higher frequency in patients.

 Results

 Complementary studies are recommended to be performed in order to evaluate high-risk alloimmunization in intermediate thalassemias. Assessment of the frequency of other red blood cell antigens, antibody screening, and identification of alloantibodies in immunized intermediate thalassemics can be helpful in this regard.

 Conclusions

 Kantigen is very immunogenic however its prevalence rafe is high among intermediate thalassaemics, but this contradiction can not be posed as the increasing cause of alloimmune reactions in such patients. Supplemental studies including screening and detection of post-transfusion created antibody types not only help in selecting the more appropriate blood type for thalassaemics but also help in justifying increasing alloimmune reactions in thalassaemics.

  Abstract

 Background and Objectives

  In this study our aim was to determine HLA-Class I and II antigens freguencies of Hamedani ethnic group. In addition to demographic studies and disease association, it has wide application in bone marrow donor registeries.

  Materials and Methods 

  In order to establish DNA-based HLA typing in central Laboratory of Iranian Blood Transfusion Organization, a comparison of serological and molecular (sequence specific primers “SSP”) methods for HLA-DRB has been performed. The study was descriptive and the population under study were selected out of the native people of Hamedan 100 healthy volunteer blood donors were chosen by questionnaire. 10ml heparinized and 3ml EDTA blood were collected from each selected donor. EDTA (PCR) samples were then frozen.

 Results

  N.I.H standard microlymphocytotoxicity and Nylon wool T and B cells separation was used for serological I and II typing. HLA-Class I plates were prepared from Iranian Blood Fractionation and Research Company and for Class II we used Biotest DR/DQ Typing Trays. PCR was done using “Roche high pure DNA extraction” Kit and HLA-DRBSSP (Biotest). The most and least frequent HLA-B antigens were B5 group (B51/B52) and B16 (38,39) respectively. Because of low resolution of HLA-DRB Kit, no significant difference was observed between serological and PCR methods. Although some blanks have been determined by PCR.

 Conclusions

  The HLA-DRB determination by PCR is mandatory for donor/recipient pairs (even sibling) for bone marrow transplantation for donors it should be done by high resolution kits.

Abstract

Background and Objectives

Hemophilia B is a genetic disorder due to deficiency or complete absence of factor IX coagulation factor. Treatment of choice for these patients is use of factor IX concentrates. Therefore, purification of plasma proteins and seperation of factor IX have been major objectives for scientists involved in this field. In this respect, purification procedure using ion exchange chromatography is widely used, but in the past decade affinity chromatography was also introduced. The objective of the present study has been to apply both techniques for the purification of factor IX and compare the quality and yield of the product.

Materials and Methods

For the purification procedure, chromatography columns (XK-16), containing DEAE sepharose and Heparin sepharose were used. Factor IX coagulation activity was measured using a one-stage coagulation assay and factor IX antigen was quantified using ELISA technique.

Results

The specific activity and relative increase in purity of factor IX was calculated and it was demonstrated that specific activity improved from 3.1 IU/mg using DEAE ion exchange to 29 IU/mg when affinity chromatography was added and purity was increased from 155 to 1450 respectively.

Conclusions

The present study demonstrates that addition of an affinity chromatography step using heparin sepharose is a major improvement in the purification of factor IX, where both specific activity and purity are increased considerably.

Key words:  Heparin affinity chromatography, Factor IX, Hemophilia B, Ion exchange chromatography

  Abstract

 Background and Objectives

  Conditions for preparation and storage of platelets for transfusion purposes may lead to platelet activation which in turn contributes to decreased ability of stored platelets to function and to survive in vivo after transfusion as compared with freshly prepared platelets. We investigated platelet membrane expression of CD62P, CD63 in platelet stored for up to 3 days under standard blood banking conditions.

  Materials and Methods 

  Twenty-four platelet units prepared by platelet-rich-plasma and platelet concentrates were evaluated during storage for markers CD62P, CD63 and pH.

 Results

 During storage for up to 3 days platelet units displayed no significant pH (p>0.05). During storage for up to 3 days (days 1 and 3) platelet units were significant in the CD62P and CD63 expressions as compared with day 0 (p<0.05).

 Conclusions

 Storage of platelet concentrates causes activated platelets. Moreover, these markers (CD62P and CD63) can act as useful in vitro means in the quality control of platelet components.

 Key words: Platelet-rich-plasma, CD62P, CD63

  Abstract

 Background and Objectives

 Diversity of heavy chain immunoglobulin (IgH) and T-cell receptor (TCR) molecules occures during B- and T-lymphocyte differentiation through the rearrangement of variable (V), diversity (D), junction (J) and constant (C) gene segments. Lymphoid leukemia cells are similar to normal precursors and have rearranged IgH, Igκ and TCR (cross-lineage rearrangement) genes which can be used as a marker of clonality and evaluation of minimal residual disease (MRD). The purpose of this study is to evaluate the pattern of TCR- δ/γ gene rearrangements using polymerase chain reaction (PCR) in B-precursor acute lymphoblastic leukemia (ALL) in Iranian children.

  Materials and Methods

 In our prospective study, bone marrow aspirates of 183 children with early diagnosis of acute leukemia were collected at admission before any chemotherapy. After reviewing cytomorphology and immunophenotyping, only 140 subjects with diagnosis of B-precursor ALLs were selected for study. Sixteen were excluded from our study due to various reasons including cellular degeneration. Mononuclear cells including leukemic blasts were isolated by density gradient. After DNA extraction, hyper-variable regions TCR- δ (Vδ2-Dδ3 and Dδ2-Dδ3) and TCR- γ(V γ V γ I and V γ II) were amplified by consensus primers using PCR. PCR products were analyzed after heteroduplex analysis and polyacrylamide gel electrophoresis (silver stain). The DNA sequences were compared and aligned to the homologous sequences of Gene Bank for confirmation. T-test, Mann whitney, Fisher exact test and Chi-square were used for data analysis.

 Results

 Clonal rearrangement of TCR-γ (Vγ) and VγI/II were present in 79.3% and 64.9% of patients respectively and only 5% of cases showed biclonal pattern. The VγII rearrangement was the most common (46.8%) type in TCR-γ. 47 (45.2%) and 11 (16.6%) of patients had Vδ2- Dδ3 and Dδ2–Dδ3 partial gene rearrangements, respectively. Biclonal/oligoclonal patterns were present respectively in 27.7% and 4.3% of cases with Vδ2-Dδ3 rearrangement. Only one patient had biclonal Dδ2 Dδ3 rearrangement.

 Conclusions

 Clonal rearrangement of TCR-δ (Vδ2-Dδ3 and Dδ2-Dδ3) genes had similar pattern to other populations. Frequency of TCR- γ (V γ I and V γ II) rearrangements were slightly higher than previous reports. and in contrary to others except Brazilian report the V γ II rearrangement was the most common type. We found no significant correlation between presence of different types of rearrangements and quantitative variables and the only significat point was the reduction of Vδ2Dδ3 with increase in age . According to preliminary results, these clonal markers can be used in diagnosis and follow up of MRD.

 Key words: Gene rearrangement, Clonality, Minimal Residual Disease (MRD)

  Transfusion medicine

  past, present, future

  Pourfathollah A.A.^1,2(PhD)

  ¹ Tarbiat Modarres University

  ² Iranian Blood Transfusion Organization-Research Center

 Abstract

 Background and Objectives

  Blood transmitted infections have always made problems in the use of blood and blood products. In this study, the prevalence of hepatitis B, C and HIV and relevant factors were evaluated among regular and non-regular donors in Arak Blood Center in the first six months of the year 1383 (2004).

  Materials and Methods

  11615 donors of Arak Blood Transfusion Center were selected. The required data were gathered by reviewing donor forms. Finaly, the subjects divided into regular and non-reguler donors according to demographic properties were compared. Results were analyzed based on Fisher Exact Test and Logistic Regression in spss software.

 Results

  40% of donors were regular and 60% non-regular. According to confirmed tests, prevalence of HBV, HCV and HIV were 0.68% , 0.2% and 0% in blood donors. These fiqures were 0.1%, 0.02% and 0% in regular donors and 0.05%, 0.45% and 0% in non-regular donors respectively. 1.4% of all blood donations were discarded for being positive in Elisa tests this rate is 111 times higher among non-regular donors. In this study viral infections in non-regular donors had more prevalence (p<0.0001) prevalence of these infections in regular donors was higher among men and lower in employees (p<0.05) as tested by Elisa. The results for both groups of subjects were higher in mobile units (p<0.05). Prevalence of infections was lower among non-regular donors as appeared in confirmed tests in subjects with bachelor degree or higher (p<0.05).

 Conclusions

  Prevalence of viral infections among regular donors was much lower than non-regular donors. Proper awareness-raising of donors about viral diseases, criteria for blood donation, appropriate behavior for blood donation in order to promote regular donation are ways to decrease viral infections.

 Key words: Prevalence, HIV, HCV, HBV

 Abstract

  Background and Objectives

 The most important component of fibrinolytic system is the proenzyme plasminogen that through various activators is converted to its active form plasmin and performs its vital functions that is fibrin clot lysis. The first anti human plasminogen antibody was prepared by Ploplis in 1982 and its effect was studied. Since then many researchers have attempted to prepare and study anti plasminogen antibodies to elucidate important aspects of structure and activation mechanism of plasminogen , physiologic condition of fibrinolysis, etc. In the present study, we studied the probable effects of three antihuman plasminogen monoclonal antibodies A₄D₁₀, A₅E₁₀ and A₂C₈ on the activation of the fibrinolytic system.

  Materials and Methods

  After separate steps like the culture of antibody-producing hybridoma cells, their injection to mice, extraction of the ascites fluid and purification of antibodies were taken, various methods such as optical evaluation of plasma clot lysis in the presence of antibodies, quantitative measurement of DD/E in D-dimer assay, evaluation with ELIZA assay using S-2251 synthetic substrate lysis and the like were used to study the effects of thses antibodies.

  Results

  Primary observations with human pooled plasma showed that in the presence of plasminogen activators (t-PA, u-PA and SK), A₅E₁₀ and A₄D₁₀ can enhance activation of fibrinolytic system, but A₃B₂ has no effect on this system. According to D- dimer assay, it was shown that the lytic effects of A₅E₁₀ and A₄D₁₀ antibodies were dose dependent the higher the amount of antibodies, the lower their effects. The other test performed with S-2251 synthetic substrate showed plasminogen activation in the presence of Urokinase therefore, lysis of this substrate enhanced in the presence of A₅E₁₀ and A₄D₁₀ antibodies. Moreover, in this test ineffectiveness of A3B2 on plasminogen activation was confirmed .

  Conclusions

  In conclusion, we suggested A₄D₁₀ and A₅E₁₀ antibodies by modification of structural from of plasminogen facilitate its activation in presence of plasminogen activators.

  Key words : Fibrinolytic, Plasminogen, Monoclonal antibody, Plasminogen activators 

 Abstract

  Background and Objectives

  Human plasma is a valuable and unique material containing vital proteins such as albumin, immunoglobulin, and coagulation factors which have pharmaceutical applications. The most current process for purification of immunoglobulin is the Cohn fractionation procedure using ethanol. The basic material for production of immunoglobulin in this procedure is fraction II. In the present study, Cohn fractionation was modified appropriately in order to achieve fraction II with a quality suitable for production of intravenous immunoglobulin.

  Materials and Methods

  Various plasma proteins were precipitated using varying concentrations of ethanol under controlled physiochemical conditions such as temperature , pH and ionic strength. The fractionation products as well as the resulting fraction II were tested and analyzed.

  Results

  The results demonstrate that the modified procedure used can result in a pure fraction II with high yield . The procedure was also suitable for all other plasma proteins. The purified fraction II contained acceptable PKA levels according to specifications of pharmacopoeia. The protein structure was also within normal limits. The repeatability of the modified procedure reported was acceptable.

  Conclusions

  The fraction II obtained in the modified Cohn procedure was a suitable intermediate product to be used in production of intravenous immunoglobulin . The PKA levels as well as the protein aggregation were minimal , and the quality of fraction II was standard . The results showed that the present modified Cohn procedure can be easily scaled up to industrial and semi-industrial levels. The resulting fraction II can be used as an intermediate in production of IVIg.

  Key words: Intravenous immunoglobulin, Cohn fraction, Molecular conformation

  Abstract

 Background and Objectives

 Blood transfusion has a critical role in clinical practice, but there has always been a possibility for transmission of infections. A lot of research have been conducted recently to find the cause of these infections. In 1997, TTV was first found by Nishizawa. Our aim was to estimate the prevalence rate of TTV in healthy blood donors and recipients of blood components in Tehran and prepare PCR kits to detect this virus.

 Materials and Methods

  In this research, we studied the prevalence rate of TTV in 250 thalassemic, hemophilic and dialysis patients and 250 blood donors. After extraction of DNA and amplification by semi-nested polymerase chain reaction, the bonds of DNA were observed in electrophoresis with 2% gel agarose.

 Results

 250 patients were studied 66.9% of whom were positive and 33.1% negative for TTV. TTV-PCR results were also studied in 250 blood donors 41% of whom were positive and 59% negative. There was a significant difference (p=0.0001) between patients and the control group.

 Conclusions

 TTV has a high prevalence in recipients and blood donors. Blood transfusion probably is not the only way for transmission of TTV, and other ways such as oral-fecal route can also play a role for transmission of TTV.

 Key words : TTV, Blood donor, Blood recipient

  Abstract

 Background and Objectives

  Umbilical cord blood (UCB) contains a high number of primitive progenitor cells for transplantation. However, the rate of UCB CD34⁺ stem cell engraftment is low. In this study we examined the effect of human MSCs (mesenchymal stem cell) on engraftment of human UCB-derived CD34⁺ cells in irradiated Balb/c mice.

 Materials and Methods

  Human UCB CD34⁺ cells were obtained from full-term normal deliveries by immunomagnetic techniques and MSCs were isolated by a standard methodology from bone marrow aspirates. Isolated MSCs were evaluated for cell-surface antigens of CD166 and CD105 by flowcytometry. The absolute count of CD34⁺ UCB stem cells was also determined by flowcytometry. Irradiated (7 Gy) Balb/c mice were transplanted intravenously with 0.2 to 1.0 × 10⁶ human UCB CD34⁺ cells in the presence or absence of 0.25 to 1 × 10⁶ human bone marrow-derived MSCs. The mice in every group on day 11 after transplantation were killed and their spleen dissected. In every group, colony assay and H and E staining were performed. To make sure of the presence of human stem cells in spleen colonies, UCB CD34⁺ cells were labeled with super paramagnetic iron oxide (SPIO) before transplantation. The colonies in spleen then underwent Prussian blue staining. In the statistical analyis, Kruskal-Wallis test using SPSS software was employed to determine the number of colonies.

 Results

  Flowcytometry assay showed that after purification, 90% of the cells were CD34⁺ and 96% marker positive for MSCs. Viablity of cells was 100%. Cotransplantation of low doses of UCB CD34⁺ cells (0.2 and 0.3 × 10⁶) and MSC (0.5 and 1 × 10⁶) resulted in a significant increase of the number of colony forming units spleen, in comparison with the engraftment of UCB CD34⁺ stem cells without MSC after 11 days (p<0.01). Prussian blue staining showed the Fe⁺² granules in UCB stem cells. This indicated that cells in the colonies were engrafted UCB CD34⁺ stem cells.

 Conclusions

  The results showed that cotransplantation of MSC with UCB CD34⁺ cells promotes engraftment of UCB CD34⁺ cells.

 �

  Key words : Stem cell, Cotransplantation, Umbilical cord blood, Mesenchymal stem cell

  Abstract

 Background and Objectives

 Occurrence of new infectious agents threatens access to zero risk in blood transfusion and enhancement of blood safety. Although sensitive methods are available for diagnosis of hepatitis, yet some hepatitis cases do not have a known etiology. In 1997, the novel DNA virus was isolated from post-transfusion serum samples of patients affected by non-A-G hepatitis. Nowadays this novel virus is known as transfusion-transmitted virus. This circular single stranded unenveloped and virucidally resistant virus is the first human circovirus and has universal distribution. It is believed that TTV may cause hepatitis and aplastic anemia. This study was conducted to determine the prevalence of TTV in healthy blood donors in Ahwaz and set up N22 PCR for subsequent first-time viral studies in south region in Iran.

 Materials and Methods

 In 2003, We studied the presence of TTV DNA by using Okamoto primers with PCR in plasma of blood donors in whom serologic tests for hepatitis A-C and HIV-Ab were negative.

 Results

 Our study showed that the virus prevalence in blood donors was 23.7% (60/253) and there was not any significant differences between prevalence of TTV and background variables.

 Conclusions

 Our findings showed the same prevalence rate as in neighboring countries however, in comparison with thalassemic patients that were studied in parallel with the present research, the difference was significant (143/250 57.3%). It shows the importance of blood transfuison in transmission of the virus.

  Abstract

 Background and Objectives

 Acute myeloid leukemia (AML) comprises a heterogenic group of malignant disorders involving cell maturation arrest at an undifferentiated stage in bone marrow. Activation of N-RAS proto-oncogene due to point mutations plays a major role in AML malignancy. Since there was no report on the frequency of N-RAS gene mutations in Iranian AML patients, therefore, we decided to determine its frequency and compare the results with age, sex and FAB subtypes.

 Materials and Methods

 In this descriptive study, 60 de novo AML patients from Tehran Shariati hospital, hematology-oncology and bone marrow transplantation center were screened for the mutations of N-RAS gene at codons 12, 13 and 61. DNA was extracted from peripheral blood samples before the start of chemotherapy. The above mentioned codons were amplified by PCR and analyzed by restriction endnuclease enzymes.

 Results

 We were able to detect mutations in 12 out of 60 (20%) patients. Most of the mutations were detected in men with an age over 40 years old. The frequency of mutations for codons 12, 13 and 61 were 15% ,11.6% and 5% respectively. Most of the mutations (33.3%) were found to happen in AML-M4 FAB subtype. We could not detect any mutation in AML-MO, M6 and M7.

 Conclusions

 We detected mutations in 20% of our AML patients. In general, the frequency of the mutations we found was in agreement with the results of other studies. However, a study with more patients and a wider range of age using a combination of PCR-RFLP and direct gene sequencing is highly recommended.

 Key words : Acute myelocytic leukemia, Mutation. PCR

  Abstract

 Background and Objectives

 The process of platelet concentrate production by plasma rich (PRP) method could activate the platelet and granules secretion of beta thromboglobulin, LDH and CD62P. Platelets activated during the preparation process do not have sufficient efficiency for hemostasis in vivo. It seems that platelet preparation by buffy coat method has an ability less than PRP to activate the platelet. Measuring platelet activation indices, such as CD62P expression and beta thromboglobulin, is a useful means to evaluate the percentage of activated platelet concentrates and compare the two methods of buffy coat and PRP.

 Materials and Methods

 In this experimental study, 15 concentrates were prepared via PRP method and 15 via BC method 15 intact blood units were also considered as control group. The percentages of CD62P expression, soluble CD62P concentrates, IL-8 level, and CD14 positive cells were evaluated. Special monocolonal antibodies that conjugated with flourecence dye in flocytometric method were used for CD62P and CD14. ELISA method was used for evaluation of soluble CD62P and IL-8.

 Results

 The average platelet count in both methods showed no significant difference, but WBC contamination rate in PRP-PCs was more than BC-PCs. In PRP-PCs, we found a little decrease in CD62P expression and increase in soluble form and IL-8 level during reservation time. The level of CD14 showed no significant difference in these components. In BC method during the three day reservation, expression of CD62P, its soluble form, and IL-8 concentrates increased and the level of monocyte surface CD14 showed slight decrease ranging from 0.4 to 0.1.

 Conclusions

 It is concluded that there is a close relationship between IL-8 and WBC count in platelet concentrates. In PRP method in contrary to BC method, high speed centrifuge causes adhesion, aggregation and platelet activation.

 �

 Key words : Platelet, Platelet rich plasma, CD62P, IL-8, CD14.�

  Abstract

  Background and Objectives

  Diversity of IgH and Ig κ molecules is generated during B and T Lymphocyte differentiation through the rearrangement of variable, diversity, junction and constant gene segments. Additionally, random insertion and deletions of nucleotides between gene segments make unique sequences which are cell or clone specific. Similar IgH and Igκ genes rearranged in normal cells of lymphoid leukemia cases can be used as a marker of clonality and for evaluation of minimal residual disease (MRD). The purpose of this study is to evaluate the pattern of IgH chain and Igκ gene rearrangements using polymerase chain reaction (PCR) in B-precursor acute lymphoblastic leukemias (ALL) to follow the MRD at day 14, day 28 (end of remission induction), week 10 , 3-6 months and 6-12 months after the initiation of treatment.

  Materials and Methods

  In our prospective study bone marrow aspirates of 183 children at the mean age of 63.6 months with diagnosis of acute leukemia were collected at admission before any chemotherapy. After reviewing cytomorphology and immunophenotyping, only 140 cases with diagnosis of B-precursor ALLs were selected for study. Mononuclear cells including leukemic blasts were isolated by density gradient. After DNA extraction, IgH and Igκ ( Vκ I-IV / Kde) were amplified by consensus primers using PCR. PCR products were analyzed after heteroduplex analysis and polyacrylamide gel electrophoresis (silver stain). The DNA sequences were compared and aligned with the sequences homologous for IgH and IgK published by Gene Bank. The follow up specimens were collected at day 14, day 28 (end of remission induction), day 45-month 3 , and 3-6 months and 6-12 months after initiation of treatment. After routine cytomorphologic analysis, similar PCR was done on follow up extracted DNAs in parallel with diagnosis DNA. MRD was considered to be approved positive if bands similar to those at the time of diagnosis were present. Statistical analysis using SPSS software (version 11.5) was performed.

  Results

  90.5% of patients had clonal IgH gene rearrangements. Monoclonal, biclonal and oligoclonal patterns were observed in 57.8%, 34.9% and 5.5% of patients with IgH (CDR III) rearrangement, respectively. Clonal patterns of Igκ-Kde were detected in 59 (67% n: 88) of BP-ALLs. According to cytomorphology about 92% of patients were in complete remission. MRD positivity decreased from more than 90% to 20% using different gene rearrangements in defined time points. Four patients who relapsed during follow up were MRD positive using 1-3 rearrangements and all except one were in clinical remission.

  Conclusions

  Clonal rearrangement of IgH had a pattern similar to other populations. IgK was slightly more frequent than previously reported and the VKI (25%) was the most common type. These differences can be explained by different techniques, DNAs and clonality markers. According to the results, these clonal markers can be used in diagnosis and follow up of MRD.

  Key words: Gene rearrangement, Minimal residual disease (MRD), Acute lymphoblastic leukemia

  Abstract

 Background and Objectives

 The incidence and severity of Graft Versus Host Disease following the use of umbilical cord blood as a source of stem cells for bone marrow reconstitution challenge the scientific findings of the immunocompetence of newborn immune cells. The reports show that self renewal characteristics and the proliferative capacity of primitive hematopoietic progenitors in the preterm cord blood are higher in comparison with term cord blood and bone marrow. In this study, the characteristics of preterm cord blood immune cells were analyzed from a naive point of view especially in comparison with its counterparts in term cord blood.

 Materials and Methods

 Term and pretem MNCs were isolated and cultivated in complete media containing PMA (50 ng/ml) and Ionomycine (1µg/ml) at the presence of monensin they were then permeabilized with %0.1 saponin. After cell stimulation with PMA and Ionomycine, staining was performed with Moab anti-CD69 antibody to estimate the level of activation and was also conjugated with anti- IL-10, IFN-γ, IL-4, IL-2 antibody to evaluate the production of cytokine. Mean percentage frequency of cytokine producing cells and the level of cytokine expression in CD4+/CD8+, CD45RA+/RO+ cells were analyzed by Epics-XL and IMMUNO-4 software. Statistical analysis was carried out using Kolmogrov-Smirnov and Student's t-test .

 Results

 Cellular phenotypic analysis showed no significant differences in CD4 ⁺ CD45RA⁺, CD8⁺CD45RO⁺ and CD4⁺CD45RO⁺ cells in term and preterm cord blood (p< 0.05). Mean percentage of CD3⁺ , CD4⁺ , CD8⁺, HLA-DR+CD3⁺, DR - HLA⁺CD4⁺ and CD25⁺ cells had significant increment in term cord blood. No statistically significant differences in the level of expression and the frequency of cytokine producing cells were observed in distinct gestational ages.

 Conclusions

 Considering the lack of any significant functional differences between term and preterm cord blood immune cells, hematopoietic stem cells of preterm cord blood not only have the same immunological behavior, especially with respect to GVHD, but also have the higher frequency and proliferative capacity. The presence of immature progenitor cells may have priority to term cord blood and be applied in transplantation settings.

 �

 Key words:�Cord blood, CD4 positive T lymphocyte , CD8 positive T lymphocyte, Cytokine

  Abstract

 Background and Objectives

 White blood cells (WBCs) are present in all cellular blood components which are prepared by standard techniques. Studies have shown that leucocytes can cause a variety of side effects after transfusion. Clinical reactions may be attributed to specific leucocyte subsets. Leukodepletion filters reduce leucocytes and some of their adverse effects. The aim of this study was to evaluate leucocyte subpopulations in whole blood prior and after Prestorage filter leukodepletion it was performed in Iranian Blood Transfusion Organization.

 Materials and Methods

 In this experimental study, different leukocyte subpopulations in whole blood were identified and quantified before and after filtration in a blood collection system having integral whole blood filtration (Baxter Fenwal, USA) equipped with RZ 2000 (Asahi, Japan) filters. Leucocytes were analysed by flowcytometry with monoclonal antibodies specific for CD45, CD19, CD3, CD14 and CD13. Statistical analysis was performed through SPSS software.

 Results

 Leukodepletion by this type of filter reduced the leukocyte load by 2.8 log 10 on average. The proportion of PMNs (CD13⁺) and monocytes (CD14⁺) to total number of leucocytes (CD45⁺) significantly reduced, but that of CD3⁺ and CD19⁺ cells showed no considerable differences before and after filtration.

 Conclusions

 This type of filter caused difference in leucocyte subset distribution after filtration. It appears that these distributions were cell size dependent. Filtration also reduced the absolute number of WBCs significantly.

 �

 Key words : Filtration, Flowcytometry, White blood cell