A Study on the Improvement of Blood Donor Health Questionnaire / 대한수혈학회지

BACKGROUND: For the safety of blood resources, the Blood Donor Health Questionnaire (DHQ) should be modified and improved allowing donors to answer questions with further accuracy. To accomplish this, it is essential to identify any part of this questionnaire that is donors find inconvenient. METHODS: The problems of the current DHQ were examined through a poll of donors at the Korean Red Cross and other hospital blood service centers from November 2008. We also compared the structure and contents of the Korean DHQ to similar document in eight other countries. RESULTS: Donors thought that the current DHQ was too complicated, took too much time (27.3%) and probed too much into a donors private life (51.2%), making it difficult to answer honestly. The Korean DHQ focuses on a deferral period and uses special medical terminology in order for an interviewer to make easy decisions regarding donor eligibility. In contrast, other questionnaires tend to focus on a donor's recall of memory, use simple vocabulary, and emphasize donor's duties, and therefore, these documents are easy for donors to understand and complete CONCLUSION: Donor-oriented DHQs using simply terminology, help donors with memory recall and emphasize a donor's duty. Also, such a document allows donors to answer frankly. Therefore donor-oriented DHQs provide a great degree of blood resource safety than interviewer-oriented DHQs.

A Study to Establish Rare Blood Type Donor Registration System / 대한수혈학회지

BACKGROUND: To find compatible rare bloods, the Korean Red Cross (KRC) blood center is currently testing many units randomly selected from the units in storage. Since this procedure is usually very time-consuming, it is necessary to establish a new donor management system for the rare blood types. METHODS: We evaluated 261 units of red blood cells (RBCs), which were supplied as compatible bloods to the requests of hospitals in 2003. RESULTS: A total of 14 hospitals requested 248 units of compatible RBCs for their 39 patients through 64 occasions and 261 units were supplied. The blood types of 35 patients were identified using the medical records of the hospitals. A total of 19 kinds of specific antigens were negative and, among them, 10 kinds of single specific antigens were negative and 9 kinds of multiple antigens were negative. The frequencies of the negative antigens were C+e(17.9%), E+c(10.3%) and Jka(10.3%), respectively. The number of tested blood units was 1,894 and the rate of compatibility was 13.8%. The mean age of 257 blood donors was 22.5 and, among them, 220 donors (85.6%) were multiple donors with 7.5 collections on average. CONCLUSION: We conclude that the rare blood type donor registration system is necessary and suggest the following preconditions. First, the results of compatible blood test and irregular antibody screening test for the blood donor need to be registered automatically by the blood information management system(BIMS). Second, the test system should be improved to enhance the irregular antibody detection rate. Third, establishment of a frozen-thawing system for RBCs is needed. Fourth, the donors need to be informed of the results of their rare blood type, if any, including irregular antibody. Fifth, a new organization should be established to manage the rare blood type donor registration system. Finally, well cooperation with hospitals and KRC blood center is needed to ensure the stability of blood service for rare blood type RBCs.

Determining the Optimal Time for Peripheral Blood Stem Cell Harvest by Detecting Immature Cells using Hematology Analyzer, SE-9000TM IMI Channel / 대한임상병리학회지

BACKGROUND: A key to successful peripheral blood stem cell transplantation is to harvest a sufficient amount of hematopoietic stem cells. A method of quickly detecting hematopoietic stem cells in peripheral blood with simple procedures using the SE-9000TM IMI channel (TOA Medical Electronics Co., Ltd., Kobe, Japan) was developed. In this study, usefulness of determining the optimal time for peripheral blood stem cell harvest using IMI channel was investigated. METHODS: Seventy nine peripheral blood stem cell collections were performed from thirteen patients with hematologic malignancy and nineteen patients with solid organ malignancy. In 13 cases, G-CSF was administrered following chemotherapy. In 19 cases only G-CSF was used to mobilize the peripheral blood stem cells. The counts of leukocytes, mononuclear cells, CD34 positive cells, and IMI in peripheral blood and leukapheresis products were determined. RESULTS: The CD34 positive cell count in harvested PBSC showed positive correlation with leukocyte cell, mononuclear cell, CD34 positive cell, and IMI in peripheral blood, with correlation coefficients of 0.48, 0.27, 0.63, 0.66, respectively. Positive correlation was presented between IMI and CD34 positive cell in peripheral blood and harvested PBSC, with a correlation coefficient, 0.83 and 0.74, respectively. CONCLUSIONS: As the SE-9000TM enables determination of the number of PBSC easily and rapidly, within approximately 85 seconds, whereas CD34 assays is expensive and needs skilled operator, the measurement of IMI positive cells is clinically useful for monitoring the peripheral blood stem cell mobilization.

Evaluation of the Collection Efficiency of the Granulocyte in Leukapheresis Using 10% Pentastarch / 대한임상병리학회지

BACKGROUND: To collect high concentration of granulocytes for transfusion to neutropenic cancer patients with infections, we investigated the effect of G-CSF or dexamethasone as granulocyte mobilizers and 10% pentastarch (PS) as the sedimentation agent in granulocyte collection by leukapheresis. Subsequently, the therapeutic effect of the granulocyte transfusions was assessed. METHODS: Forty five leukapheresis were performed with CS-3000Plus (Baxter, Deerfield, IL, USA) using 10% pentastarch. The donors were classified into three groups according to their premedication drugs and the interface detector offset; group 1 used dexamethasone with offset 15 (n=16), group 2 used dexamethasone with offset 33 (n=16), and group 3 used G-CSF with offset 33 (n=10). We compared total collected granulocyte counts and granulocyte collection efficiency (GCE). RESULTS: The mean counts of total granulocytes collected and GCE were as follows; 0.9 0.5 x 1010 and 31.6 14.3% in group 1, 1.3 0.6 x 1010 and 39.0 14.2% in group 2, and 1.6 0.9 x 1010 and 63.9 32.2% in group 3, respectively. The counts of granulocytes collected in group 3 was significantly higher than that in group 1 (P<0.05). The GCE of group 3 was significantly higher than that of group 1 and group 2 (P<0.05). Sixteen granulocyte transfusions were performed to 11 patients. We observed successful therapeutic effects in 10 out of 16 transfusions (63%). CONCLUSIONS: G-CSF indicates greater potency than dexamethasone although its high cost is limitation of routine use as mobilizing agents and PS was an excellent red cell sedimenting agent in granulocyte collection. Large volume granulocyte transfusions allow high therapeutic effects in neutropenic patients with marrows of sufficient regenerating capacity.

Peripheral Blood Stem Cell Collections in Childhood Tumor Patients Weighing Less than 25 kg Using Large Volume Leukapheresis / 대한소아혈액종양학회지

BACKGROUND: Peripheral blood stem cell (PBSC) transplantation has been widely used to support the hematopoietic recovery after high dose chemotherapy in patients with advanced malignancies. The procedure of PBSC collection in pediatric patients is similar to that in adults, but needs the 'fine tuning' of the volume shift of each procedure and the technical factors to achieve specific target goals. This article provides our experience with fourteen collections of PBSC from five patients less than 25 kg in weight. METHODS: Patient's diagnoses were 2 stage IV neuroblastoma, 1 non-Hodgkin's lymphoma, 1 stage IV Ewing sarcoma, and 1 stage IV rhabdoid tumor of kidney. Their age ranged from 2 to 7 years old. Collections were performed using COBE Spectra or CS3000 plus that had been primed with leukoreduced, irradiated red blood cells. Patients underwent large volume leukapheresis. Radial artery was used as draw line and subclavian vein was used as return line. The blood to ACD ratio was 24:1 with 3000 units of heparin added to each 500 mL of ACD, in addition, heparin (1000 units) was added to collection bag when performed with COBE Spectra. Simultaneously, calcium chloride solution was dripped into an another venous line. During one course of large volume leukapheresis, about 5,000 mL of blood (>three total blood volume) were processed at a flow rate of 25~35 mL/min. RESULTS: The mean of total WBCs in collected components per procedure was 5.9+/-2.9x109 (3.0-10.5x109) with yield of 3.6+/-2.0x108 per kg of body weight. The mean of total CD34+ cells was 5.2+/-4.5x106 per kg (1.6-14.6x106/kg) for each collection. The patients tolerated well during the procedure without any apparent symptoms related to anemia or volume deficit or overload. CONCLUSION: In children weighing less than 25 kg, peripheral blood progenitor cell collection can be safely and efficiently performed with continuous flow blood cell separators, primed with red cells and additional heparin anticoagulation.

Multicenter Study on Flow Cytometric Enumeration of CD34+ Hematopoietic Stem Cells / 대한임상병리학회지

BACKGROUND: The increased use of peripheral blood stem cells (PBSC) to reconstitute hematopoiesis needs accurate methods to control the quality. Transplantation centers often rely on CD34+ cell quantitation by flow cytometry to ensure adequacy of hematopoietic progenitor cell collection. Because of variation in interpretation, a lack of interlaboratory proficiency studies, and no generally accepted methodology, comparison of CD34 positive cell data among centers is difficult. To assess the variability of CD34 assay, a multicenter survey involving 4 samples and 14 major laboratories was conducted to compare CD34 positive cell quantitation. METHODS: Fifty-three laboratories were asked to participate and participants were surveyed for their cell processing and staining methods as well as instrumentation, software, and analysis parameters. Four cord blood samples from delivered placenta were shipped by overnight carrier to fourteen participating laboratories for analysis. Total leukocyte count, total mononuclear cell count, CD34 positive cell percentage of leukocytes, absolute CD34 positive cell count were assayed items. RESULTS: Fourteen laboratories were recruited. Six laboratories used only CD34 monoclonal antibody and five laboratories included one more additional antibody (CD45 or CD14) in their procedure. Two laboratories used a commercial kit (ProCOUNTTM). One laboratory analyzed with indirect immunofluorescent assay. Coefficient of variants of CD34 positive cell percentage of leukocytes of each sample were 74.1%, 100.0%, 73.1%, 70.0%, that of absolute CD34 positive cell count were 64.2%, 84.7%, 79.5%, 75.8%, respectively. CONCLUSIONS: We observed an alarming variation among the CD34 positive cell counts reported from different laboratories. An effort to standardize the procedure, reporting policy, quality control as well as to make close communications with physicians and laboratorians should be made for proper management of the patients.

A Severe Thrombocytopenia Associated with Cyclosporine-Dependent Antiplatelet Antibody Against Platelet Glycoproteins Ib/IX and IIb/IIIa Complexes / 대한임상병리학회지

A 51 year-old woman underwent living related renal transplantation under cyclosporine A immunosuppression. After surgery, she did well initially, but the serum creatinine level subsequently rose to 3.6 mg/dL on postoperative day 95, she was admitted at Severance Hospital for further evaluation. On admission day 4, a renal biopsy was performed, and the microscopic findings revealed an interstitial mononuclear cell infiltrate, suggestive of severe allograft rejection. Because of persistently impaired renal function, the patient was began on twice weekly hemodialysis, and the progression of renal deterioration paralleled the onset of a thrombocytopenia. The platelet count dropped to 13x109/L despite daily platelet transfusion. On admission day 19, antiplatelet antibody against the glycoprotein Ib/IX (GP Ib/IX) and glycoprotein IIb/IIIa (GP IIb/IIIa) complex was detected in the presence of cyclosporine A (CsA) with modified antigen capture ELISA (MACE) assay, thereby implicating the drug. CsA was stopped immediately and immunosuppression drug was changed to FK506. After CsA was discontinued 7 day later, her platelet count returned to normal, up to 170x109/L without requirement of any platelet concentrates. This paper presents the first case of CsA induced thrombocytopenia in Korea which was confirmed by in vitro CsA dependent antiplatelet antibody detection test.

Platelet transfusion support for splenectomy in patients with chronic immune thrombocytopenic purpura / 대한수혈학회지

BACKGROUND: Splenectomy is often performed for the patients with refractory chronic immune thrombocytopenic purpura (ITP). Still, there are no generally accepted guidelines for the minimum level of platelet count and the average requirement of platelet transfusion so that the patients can safely undergo splenectomy. We evaluated the changes of platelet count and transfusion requirements around the splenectomy in patients with chronic ITP. METHODS: We reviewed the medical records of 25 patients with chronic ITP. We compared the platelet counts at admission, immediately pre-op and several post-op days. We also investigated the number of platelet concentrates transfused around splenectomy. We determined the effect of splenectomy according to Difino's classification. RESULTS: The median platelet counts of the patients was 18x109/L (7-238x109/L) on admission and recovered to 108x109/L (22-460x109/L) on preoperation day by platelet transfusion and immunosuppressive treatment. The platelet counts were rapidly recovered after splenectomy from the day of operation. Only 3 patients needed platelet transfusion after splenectomy. Thirteen among twenty five patients (52%) underwent operation without platelet transfusion support. Most transfusions were done before the surgery and 80% (12/15) of the patients required transfusion of more than 10 units of random donor platelet concentrate. Twenty one patients (84%) showed the complete remission after splenectomy. CONCLUSION: Splenectomy can lead to rapid remission even in most cases of refractory chronic ITP. Many cases can undergo the operation only with treatment modalities other than transfusion such as immunosuppressive agents and/or immunoglobulin. The minimum level of platelet counts for splenectomy may be safe over 50x109/L and about 10 units of platelet concentrate may be enough for preparation of splenectomy.

Application of ABO genotyping in determination of ABO subgroups / 대한수혈학회지

BACKGROUND: The knowledge about the nucleotides sequence of 9th chromosome that regulates the phenotype of ABO blood group has made the ABO genotyping possible. Since the genotyping can be done with only a small amount of DNA sample, it was primarily applied to the field of forensic medicine. When applied to the blood bank, it is useful in the resolution for ABO discrepancies between the cell and serum typing and determination of A and B subgroups. Rapid ABO genotyping using the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method and its value in determination of ABO subgroups is presented. METHODS: ABO genotyping was performed in seven patients and three families, seven were the cases of ABO discrepancies in routine ABO grouping and three families were for the confirmation of the ABO group. To identify the 261th nucleotide, a 252 bp PCR amplifed fragment was amplified by PCR and digested with Kpn I. For 703th nucleotide, a 128 bp PCR amplified fragment was designed and digested with Alu I. To determine the ABO genotype, the patterns of digestion in DNA fragment were examined. RESULTS: Among the seven cases of ABO discrepancies, B3 and Ael were two cases each. Weakened B due to leukemia was the one, and the other two cases were cis-AB and Am. The three families for confirmation of the ABO group were acquired B due to infection one family, cis-AB two families. CONCLUSIONS: ABO genotyping is a rapid and reliable method that can be used in the case of ABO discrepancies and determination of ABO subgroups.

Evaluation of a domestic second generation platelet storage container / 대한수혈학회지

BACKGROUND: Polyvinyl (PVC) plastic container plasticized with di-(2-ethylhexyl) phthalate (DEHP) has been used for the storage of platelet concentrates for five days in Korea. Authors evaluated a second generation platelet storage container plasticized with tri (2-ethylhexyl) phthalate (TOTM) which was recently produced by Green Cross Medical Corp.(Korea). METHODS: 30 units of platelet concentrates were stored in TOTM-PVC container at 22'C in a flatbed agitator. Samples were taken at day 1,3,5, and 7 from the containers and tested for platelet count, MPV, PDW, pH, HCO3-,LDH, lactate, hypotonic shock response and beta-thromboglobulin (beta-TG). Electron microscopic examination was also performed. RESULTS: The number and functions of platelets were well preserved during storage. pH was maintained above 6.8 and any evidence for platelet activation was minimal. CONCLUSION: The TOTM-PVC second generation platelet storage container recently produced by the Green Cross Medical Corp.(Korea) was able to preserve platelets for at least five days without significant storage lesions.

Preapheresis WBC, mononuclear cell and CD34+ cell counts as predictors of optimal timing of peripheral blood stem cell collection / 대한수혈학회지

BACKGROUND: Peripheral blood stem cells (PBSC) transplantation has been widely used as a substitute of bone marrow transplantation in patients with hematologic malignancies and solid tumors. Because, PBSC harvest by serial daily apheresis procedure is expensive and time consuming, it is important to determine the best time to start the collection for reducing the number of apheresis procedure. We analyzed our experiences of PBSC collections and evaluated the preapheresis hematologic parameters that may predict the PBSC yields. METHODS: One hundred seventy six PBSC harvests from seventy cancer patients (median age : 32 yrs; fourty five males and twenty five females) were performed using our large volume leukapheresis protocol (total blood volume processed : over three total blood volume) after chemotherapy and infusion of G-CSF. Peripheral blood obtained immediately before the start of apheresis was analyzed for total WBC, mononuclear cell (MNC), and CD34+ cell counts. Total WBC, MNC, and CD34+ cell count were performed on selected samples of PBSC from each patient before freezing for determining the PBSC yields. Linear regression analysis was performed on logarithmized data whether preapheresis WBC, MNC, and CD34+ cell counts on the day of harvest in the peripheral blood might correlate well with the PBSC yield, respectively. RESLUTS: With the use of linear regression analysis, preapheresis WBC counts and MNC counts were not correlated significantly with the CD34+ cell yield in PBSC harvests (WBC/microliter in PB vs. CD34+ cell/kg in harvests, r=0.35, p=0.10; MNC/microliter in PB vs. CD34+ cells/kg in harvests, r=0.42, p=0.07). But the CD34+ cell count (CD34+ cells/microliter in peripheral blood) correlated most closely with the progenitor cell yield in the corresponding leukapheresis product (CD34+ cells/kg body weight, r=0.75, p<0.001). A number of 20 circulating CD34+ cells/microliter blood ensured 2.0 x 106 CD34+ cells/kg, that is known to be a threshold dose for rapid hematologic recovery, and the best time for the collection on the same day by a single leukapheresis in more than 85% cases. CONCLUSIONS: The number of CD34+ cells/microliter blood allows a reliable prediction of the CD34+ progenitor cell yield in subsequent leukapheresis procedure, while WBC and MNC counts did not predict the progenitor cell yield. A level of more than 20 CD34+ cells/microliter indicates that the threshold quantity of 2.0 x 106 CD34+ cells/kg is likely to be obtained by a single leukapheresis processing 15~20 liters of peripheral blood.

Comparison of Molecular Biologic Methods for Detecting HBV-DNA in the Sera which Showed Both Hepatitis B Surface Antigen and Antibody Positivity / 대한임상병리학회지

BACKGROUND: Serologic markers are used to screen and diagnose the hepatitis B virus infection. In endemic area of hepatitis B, the coexistence of HBsAg and anti-HBs was frequently observed. This finding is unusual and difficult to interpret. In this study, we performed three molecular biologic assays-polymerase chain reaction (PCR), chemiluminescent molecular hybridization assay (CMHA), branched DNA (bDNA) nucleic acid hybridization assay- to detect HBV-DNA in the sera which showed both HBsAg and anti-HBs positivity. To define the patients` exact clinical conditions, we analysed the characteristics of the patients according to their diagnoses, other serologic markers and clinical findings. METHODS: HBsAg and anti-HBs were detected by EIA (Enzygnost, Behringwerke, Germany) from clinical specimens of Yonsei University College of Medicine Severance Hospital collected In the period between January 1996 and December 1996. Eighty three specimens from Severance Hospital and twenty two specimens from Health Care Center were randomly selected and were subjected to HBV PCR, HBV CMHA and HBV bDNA assay for the presence of HBV-DNA. RESULTS: The patients were arbitrarily divided into 4 groups on the basis of the optical density values of enzyme immunoassay results. Group I (high HBsAg and high antral-HBs) consisted of 6 cases; group II (high HBsAg and low anti-HBs) consisted of 70 cases, group III (low HBsAg and high anti-HBs) consisted of 1 case; group IV (low HBsAg and low antral-HBs) consisted of 6 cases. Among 83 cases, the positive rate was 51.8% (43 cases) using PCR method, 53.0% (44 cases) using CMHA, 60.2% (50 cases) using bDNA assay. HBeAg and anti-HBc IgM were helpful to predict the presence of HBV-DNA in the sera. CONCLUSIONS: More than half of the patients who showed both HBsAg and anti-HBs positivity were positive for HBV-DNA by molecular biologic methods. In contrast, no one whose serologic markers with only anti-HBc positivity with out HBsAg and anti-HBs positivity showed HBV-DNA positive in the sera from Health Care Center. Taken together, the management and follow-up of the patients of both HBsAg and anti-HBs positivity could be greatly aided by combined adoption of any one molecular biologic assay of HBY-DNA with other serologic markers such as HBeAg and anti-HBc IgM.

Platelet Activation and Storage lesions in Apheresis Platelet Concentrates / 대한수혈학회지

BACKGROUND: Three cell separators are being used and they collect platelets with different centrifuge speed and duration. Because centrifugation may cause platelet activation, the differences of centrifuge speed and duration are important in controlling the quality of apheresis platelet products. We compared many parameters of activation of platelets collected by Spectra (Cobe BCT, Lakewood, CO, USA), CS3000plus (Baxter Healthcare, Fenwal Division, Round Lake, IL, USA) and Mobile Collection SystemTM (MCS, Haemonetics co., Braintree, MA, USA). METHODS: Platelets were collected from ninety-five normal donors with Spectra (n=39), CS3000plus (n=19) and MCS (n=37). We underwent the procedure according to the automatic program set. We measured platelet yield and assayed pH, hypotonic shock respose (HSR), CD62 (p-selectin, GMP140) expression and beta-thromboglobulin in each stored unit on day 0 and day 3 for evaluation of the storage lesions. RESULTS: Platelet yield per product was 3.7 +/- 1.2 x 1011, mean final product volume was 316 +/- 69 mL and mean procession time was 100 +/- 19 minutes. Mean collection efficiency was 42.5 +/- 8.3%. The cell separator volume of product collected by CS3000plus was the smallest while platelet concentration and total yield were the highest in the product collected by Spectra. The pH of the products were 7.1 +/- 0.1 on day 0 and 6.7 +/- 0.4 on day 3. Hypotonic shock response was 69 +/- 13 % on day 0 and 28 +/- 17 % on day 3. P-selectin expression was 19 +/- 9 % (4.2 +/- 1.9 relative fluorescence intensity, RFI) on day 0 and 60 +/- 22 % (17.9 +/- 14.2) on day 3. beta-thromboglobulin was 28.5 +/- 7.0 IU/107 platelets on day 0 and 31.3 +/- 7.2 IU/107 platelts on day 3. The comparison of the three cell separators showed that on day 0 platelet product of MCS has lower pH and higher beta-thromboglobulin release than others (p<0.05). And on day 3 platelet product of MDS has better hypotonic shock response than others (p<0.05). Other parameters revealed no differences among three cell separators. The expression of p-selectin was shown to correlate highly with pH reduction (r=0.72), but not with the release of beta-TG (r=0.24). CONCLUSIONS: Most parameters showed no differences among three cell separators, but apheresis platelet concentrates processed by MCS showed lower pH on day 0 and higher beta-thromboglobulin concentration on day 0 and day 3 than apheresis platelet concentrates processed by Spectra or CS3000plus and hypotonic shock response on day 3 was the lowest in CS3000plus. So platelet activation produced during apheresis processing was lowest in apheresis platelet concentrates with Spectra.

Production and Assessment of Goat Antihuman Globulin Reagent / 대한수혈학회지

BACKGROUND: Since the 1960 s rabbit antihuman globulin reagent has been used widely. Recently most conjugate of enzyme immunoassay is produced from goat, and precise purification method is developed. Therefore we evaluated the commercial value of the goat antihuman globulin as a blood bank test reagent. METHODS: The human IgG was purified by protein-A gel, and injected into goat. The goat antihuman globulin was coupled by CNBr activated sepharose 4B-gel and purified by 0.2M glycine elution buffer. For verification of this reagent, commercial reagents(Ortho rabbit reagent & DiaMed Gel test) were used. RESULTS: The minimal concentration for detecting antibody of goat reagent was 9 ng/mL. The results of direct antiglobulin tests, with 400 samples collected from donated blood in CPDA-1, were all negative(false positive rate: 0%). With 613 samples collected from inpatients of Severance Hospital, the results were positive in 35 patients(positive rate:5.7%), and those results were in complete agreement with commercial reagent(concordance rate: Goat vs. Ortho :99.8%, Goat vs. DiaMed :98.4%). And with 30 samples of artificially prepared complement-coated RBC, the results were all negative. Indirect antiglobulin test showed higher agglutination score than commercial reagents. CONCLUSIONS: Goat reagent showed high sensitivity and specificity in comparison with rabbit reagent. Because false positive reaction was not observed in negative control samples, the heterophil agglutinin reaction, which was the major problem when the reagent was initially developed, might be excluded. In conclusion, goat reagent seems to be more economical than rabbit reagent because the former can be obtained in a large quantity and of high potency.