Application of microfluidic chip analytical systems in ABO genotyping / 中国实验血液学杂志

Limitations of polyacrylamide gel or agarose gel electrophoretic methods in genotyping research affect the interpreting of detection results. In order to develop a simple and reliable method for appraising results of ABO genotyping detection, the microfluidic chip analysis system was established by using microfluidic chip to replace the gel electrophoresis and combining with multiplex-PCR-RFLP technique. 150 blood samples were tested by this microfluidic chip analysis system with multiplex-PCR-RFLP technique to evaluate its stability and accuracy. The results showed that all the testing results were consistent with serologic ABO genotyping results and 1 blood sample with decrease of B antigen caused by CML was identified. In conclusion, the established microfluidic chip analysis system is stable and reliable technique. Application of this technique enables the ABO genotyping results to be more objective and accurate.

ABO genotyping of Han population in Beijing / 中国实验血液学杂志

The aim of this study was to establish a diagnostic method for ABO genotyping and to investigate the distribution of ABO genotype in Beijing Han population so as to understand the distribution characteristics and regularity of ABO genotype. An ABO genotyping method was established by using multiplex-PCR-RFLP and PCR-SSP techniques, and the ABO allele frequency in Beijing Han population was investigated. The results showed that A102, O1 and B allele were more common genes in Beijing Han individuals. And A102 allele was predominant in the phenotype A group in this population. Three O2 alleles were found and no A201 allele was found while gene frequency investigation was performed. No A101A101, A101O2, A102O2, BO2 and O2O2 in this population were discovered. It is concluded that the primary regularity of ABO allele distribution in Beijing Han population is found through this study. It provides basic reference for further study of ABO types.

B to O erythrocyte conversion by the recombinant alpha-galactosidase / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Human group O red blood cells have great benefit in specialized transfusion areas such as armed conflict and natural calamity. The group B antigen differs structurally from group O antigen only by the addition of one terminal alpha-linked galactose residue. In this study we aimed to remove the terminal galactose from group B red blood cell to get group O red blood cell.</p><p><b>METHODS</b>alpha-galactosidase cDNA was cloned by RT-PCR from Catimor coffee beans grown on Hainan Island of China. The vector for alpha-galactosidase cDNA expression was constructed and transferred into Pichia pastoris cells by electroporation. The transgenic cells were cloned by fermentation and the recombinant alpha-galactosidase was purified by ion exchange chromatography. After studying the biochemical characters of alpha-galactosidase, we have used it in converting human erythrocytes from group B to group O.</p><p><b>RESULTS</b>The purity of recombinant alpha-galactosidase was higher than 96%, which was thought to be suitable for the use of blood conversion. Enzymatically converted human group O red blood cells (ECHORBC) exhibited membrane integrity, metabolic integrity, normal cell deformation and morphology. There were no coagulation between ECHORBC and any group of human blood. The ECHORBC will keep normal structure and function for a period of 21 days at 4 degrees C in monoammoniumphosphate nutrient solution. Experiments with Rhesus monkeys and gibbons showed that transfusion of enzymatically converted erythrocytes was safe.</p><p><b>CONCLUSION</b>ECHORBC can be easily obtained from group B red blood cell by alpha-galactosidase digestion. This study suggests that ECHORBC could be transfused to patients safely and efficiently.</p>

Animal trials for mPEG-modified red blood cells / 中国实验血液学杂志

This study was aimed to investigate the survival rate and difference of transfused modified and unmodified RBC at 24 hours. The modified and unmodified RBC from mice, monkey, pig and human were labeled by using FITC, then these blood RBCs were transfused to homogeneous and heterogeneous animals. The result showed that 24 hour survival rate of unmodified mice RBC transfused to mice was 74%, while survival rate of 2.0 mmol/L mPEG-SPA modified mice RBC transfused to mice was 45%, difference between them was significant. The 24 hour survived rate of unmodified human RBC transfused to mice was 8%, while 24 hours survival rate of 2.0 mmol/L mPEG-SPA modified human RBC transfused to mice was 5% without statistical difference. The 24 hour survived rate of homogeneous transfusion of modified monkey RBC was 90%, while survival rate of modified human and pig RBC was zero on 24 hours after transfusion to monkey. It is concluded that RBC labeling methods and mice species are unrelated to 24 hours survival rate, but mPEG variety and concentration are related to mouse RBC life-span. It is incredible to use mouse RBC homogeneous transfusion result instead of human RBC to evaluate longevity and safety of modified human RBC. But modified human RBC transfused to mice can be a model to evaluate longevity of modified human RBC. It is very difficult to get the result about modified RBC life span by RBC transfusion among great heterogeneous mammal animals. So evaluation in large mammal animal models needs to be further studied.

Stability of hepatitis C virus RNA in various processing and storage conditions / 中国实验血液学杂志

The study was purposed to investigate whether processing and storage conditions might influence the stability of the HCV RNA in whole blood or in plasma. The samples obtained from seven patients known to be positive for HCV RNA were kept in different storage conditions with different anticoagulants, and at the end of processing the plasma samples were frozen at -80 degrees C until fluorescent quantitative PCR testing. The results showed that there was no significant loss of HCV RNA titers in whole blood anticoagulated with CPDA or ACD or EDTA or none (P > 0.05), while differences in comparison of the EDTA-anticoagulant storage condition with three other anticoagulants storage conditions at 4 degrees C after 48 hours were significant (P < 0.05). The HCV RNA level decreased to 53.8%, 72.5% and 29.8% after 48 hours of storage of whole blood anticoagulated with ACD at 4 degrees C, 25 degrees C and 37 degrees C respectively. The HCV RNA level of plasma samples stored at 4 degrees C and at 25 degrees C (room temperature) after 7 days decreased to 70.9% and 25.1% respectively. After four freeze-thaw cycles the HCV RNA level decreased 38.9% in plasma samples. It is concluded that the HCV RNA is stable relatively. The HCV RNA is resistant to degradation under routine laboratory handling and storage conditions or blood collection, transport and processing conditions. The influence of different anticoagulants on the stability of HCV RNA is different. Blood samples would better be stored at 4 degrees C after collection and plasma separated within 48 hours. And it is important for the stability of HCV RNA undergoing asepsis blood collection process. HCV RNA remains stable at 4 degrees C for at least 7 days or at room temperature for 3 days, allowing greater flexibility in samples collection and transport in transfusion practice nowadays. HCV RNA in plasma samples subject to up to three short-term freeze-thaw cycles is still stable.

Correlation between signal/cutoff ratios of anti-HCV enzyme immunoassay (EIA) and their true positivity in blood donors / 中华肝脏病杂志

<p><b>OBJECTIVES</b>To evaluate the correlation between signal/cutoff (S/CO) ratios of anti-HCV EIA and their true positivity for determining the predictive value of S/CO ratios.</p><p><b>METHODS</b>One hundred and fifty-nine samples of blood from donors positive for anti-HCV at the initial screening were collected from Beijing, Guangzhou, Hangzhou, Kunming and Urumchi. All the samples were retested by Ortho and 6 Chinese domestic anti-HCV EIA kits in duplicate, and detected for HCV RNA (NAT) using Chiron Procleix HIV/HCV system (transcription mediated amplification, TMA). The HCV RNA negative samples were further tested for anti-HCV by Chiron RIBA 3.0. Either NAT or RIBA positive samples were interpreted as the true positive.</p><p><b>RESULTS</b>All 7 anti-HCV EIA kits had a significant correlation between S/CO ratios and true positivity. The S/CO ratio of Ortho > or = 3.8 predicted the true positivity in 96.1% of the samples tested. The S/CO ratios of BGI-GBI, GWK, SABC, KHB, InTec, and Wantai were > or = 7.0, > or = 10.0, > or = 6.0, > or = 10.0, > or = 8.6, > or = 14.0 and predicted 96.1%, 96.1%, 97.3%, 96.0%, 96.1%, 96.0% of the true positivity, respectively.</p><p><b>CONCLUSIONS</b>The S/CO ratios of anti-HCV EIA kits are associated with the true positivity. S/CO ratios of Ortho, BGI-GBI, GWK, SABC, KHB, InTec and Wantai predicting > or = 95% true positivity are > or = 3.8, > or = 7.0, > or = 10.0, > or = 6.0, > or = 1 0.0, > or = 8.6 and > or = 14.0, respectively.</p>

Preparation of transfusable human universal red blood cell with recombinant alpha-galactosidase / 中国实验血液学杂志

In order to meet the demand for safe transfusion in special conditions and to utilize the donated blood supply efficiently, technology has been developed to convert erythrocytes from type A, B, or AB to "universal donor" blood. Conversion of blood type B to O was performed by means of recombinant alpha-galactosidase digestion. The results showed that blood type B to O was converted successfully, 1 transfusion unit of red cells of group B (100 ml totally) could converted to universal blood cells in the optimal conditions including pH 5.6, 26 degrees C, 2 hours, obturation and sterilization. It is concluded that the universal red blood cells converted from group B to group O are conformed to demand of identification rules of biological products, no harmful effects of alpha-galactosidase on cell structure and function are observed. The converted red cells can stored in 4 degrees C for 21 days.