Objective To prepare B-lymphoblastoid cell lines of HLA novel allele B*5610 in a family for further study and identification . Method Isolate mononuclear cells under aseptic conditions from the peripheral blood. After infection with Epstein-Barr virus, the cells were cultured in 20% FBS, 2?g/ml CsA RPMI 1640. Results Immortalized B-lymphoblastoid cell lines of five B *5610 carriers in a family were achieved, and the new genes were inherited stably. Conclusion Our work is important for storing and breeding the precious material of biomedicine because the B *5610 genes in the immortalized B-lymphoblastoid cell lines were inherited stably.

Objective To gain purified platelets for further research of the immune function of platelet.Methods Platelets were purified by positive(MS) and negative(LD) magnetic cell sorting(MACS) separation.The percentage of activated platelets was detected by flow cytometry and nucleated cell clearance was evaluated by white cell count.Results The percentage of activated platelets before separation was(2.39?1.10)%,and increased to(2.56?1.08)% and(16.76?4.04)%,respectively,after MS and LD MACS.The clearances of nucleated cells after MACS MS and LD were(98.44?0.24)% and(98.47?0.18)%,respectively.The recovery rate of purified platelet after MS and LD was(76.50?1.49)% and(79.20?2.61)%.Conclusion MACS LD method was more suitable for the platelet purification.

ObjectiveTo analyze the polymorphism of HLA-A,B,and DRB1 alleles and their haplotypes in Chinese Man population.Methods Frequencies of HLA-A,B and DRB1 alleles and haplotypes were estimated by maximum-likelihood estimation method based on the genotypes of 2183 Chinese Man bone marrow donors.ResultsA total of 18 HLA-A alleles,44 HLA-B alleles and 15 HLA-DRB1 alleles were detected in Man population,and the most frequent alleles were A*02,A*11,A*24,A*30,A*33,B*13,B*35,B*46,B*51,B*40(B60),B*40(B61),B*15(B62),DRB1*04,DRB1*07,DRB1*08,DRB1*09,DRB1*11,DRB1*12,DRB1*13,DRB1*14 and DRB1*15.A*30-B*13,A*02-DRB1*15,B*13-DRB1*07 and A*30-B*13-DRB1*07 were the most frequent haplotypes in Man population for A-B,A-DRB1,B-DRB1 and A-B-DRB1 haplotype,respectively.The number of haplotypes with frequency ≥ 0.01 for A-B,A-DRB1 and B-DRB1 haplotype was 31,24 and 27,respectively,and ≥ 0.005 for A-B-DRB1 haplotype was 32.There were 14 in A-B,3 in A-DRB1,14 in B-DRB1 and 38 in A-B-DRB1 haploypes that showed strong linkage disequilibrium with ALD≥0.40.ConclusionsThe distribution of HLA-A,B and DRB1 alleles and haplotypes in Man population was similar to that in Northern Chinese Han population.

Objective To study the inter-allelic recombination event occurring in the HLA-C locus in a family of Chinese Han nationality, and to evaluate the molecular genetic background of the new HLA allele.Methods Peripheral blood samples were collected from a Chinese leukemia woman patient, as well as her healthy parents and two brothers.HLA-A, C, B, DRB1 and DQB1 alleles were typed by high-resolution PCR-sequence-based typing (SBT) method using Atria Genetic AlleleSEQR HLA SBT kits.The Protrans S4 HLA-C single allele-specific sequencing strategy was used to separate the two HLA-C alleles and to determine novelty of the allele.The full length sequences of HLA-C alleles of the patient and her parents were further analyzed using cloning and haplotype sequencing method. The HLA five loci linked haplotypes and the recombination site were analyzed by family study, meanwhile the full length sequences of the five HLA-C alleles were compared with the IMGT/HLA database by the program BLAST.Results The two haplotypes of the father and mother were a:A*0207-C*010201-B*550201-DRB1*090102-DRQ1*030302 and b:A*240201-C*120202-B*5201-DRB1*1502-DRQ1*0601, c:A*300101-C*060201-B*130201-DRB1*0405-DRQ1*0401 and d:A*110101-C*070201-B*4001-DRB1*080302-DRQ1*0601,respectively.The two brothers inherited their parent′s haplotypes a, d and b, c respectively.The two haplotypes of the patient were the maternal c and paternal recombinant a/b haplotype.The recombinant a/b haplotype A*240201-C*new-B*550201-DRB1*090102-DRQ1*030302, A*240201 came from the paternal haplotype b,while B*550201-DRB1*090102-DRQ1*030302 came from the other paternal haplotype a.When comparing the full length sequences of the HLA-C new allele with the father′s allele C*010201 and C*120202, it could deduce that the recombinant a/b haplotype derived from a recombination event occurring between the paternal chromosome 6 during meiosis.The crossover site was between genomic nt273 and nt330 of HLA-C alleles, which created a HLA-C new allele and the fifth haplotype of the family, and inherited it to the patient.The full length sequences of the new allele had been submitted to Genbank, and officially named C*0121 by WHO nomenclature committee.Conclusion This study demonstrates a rare inter-allelic recombination event occurring in the HLA-C locus within a Chinese Han family and illustrates the process of novel allele and haplotype, and provides direct theory for further studying the mechanisms of gene recombination and HLA polymorphism.

Objective Study on fetomaternal immuno state and RHD type of a pregnant woman of weak D phenotype.MethodsThrough polymerase chain reaction (PCR)、direct genomic DNA sequencing and flow cytometry.ResultsIn both sequence specific promer (SSP) PCR and the sequencing PCR tests, the sample was detected negative in exons 3 6 of the RHD gene, whereas all other exons (exons 1 2,7 10) were tested positive. And the sequence of detected exons (exons 1 2,7 10) are the same with normal RHD in GenBank (accession no. AJ299020 1 and AJ299026 9). Serologically and genetically, the sample can be designated as D category VI type Ⅲ. Through a duce tube PCR method, the RhD zygosity of this individual was typed CD VI e/cde。In flow cytometry, a few fetal erythrocytes were detected in peripheral blood of the mother. However there were no anti D detected in sera and hemolytic disease of the newborn(HDN) observed at all.ConclusionSevere cases of HDN have occurred in D positive babies born to partial D mother with anti D, although HDN don't take place in this case. We may still consider D VI phenotype individuals as D positive donors and D negative receiptions in our transfusion practice and in clinical anti D allo immune prophylaxis and monitoring.

Objective To compare efficacy of female sterilization by modified Uchida technique and silver clips and to evaluate the influence on operation procedure and clinical effect with or without surgery training of service providers. Methods A comparative, multicenter clinical trial was performed in 18 county and township-level service centers. Totally 2198 women underwent sterilization from these 18 study center were divided into 1116 women sterilized by modified Uchida technique and 1082 women by silver clips.Those 18 centers were classified into 9 training groups which provide surgical skills of sterilization and other contents and 9 non training groups. Clinical documents of sterilization were recorded. All women were followup at 3, 6 and 12 months after surgery. Results There were no complications during surgery by both sterilization. The failure rate was 2.03% (22/1082) in silver clip method and the mean operative time were ( 12. 4 ± 6. 4 ) minutes in training group and ( 14. 4 ± 8. 1 ) minutes in non training group. In modified Uchida method, the failure rate was 0. 18% (2/1116) and the mean operative time were (16. 2 ± 4. 9)minutes in training group and (19.0 ±8.6) minutes in non training group. The mean operative time between two groups reached statistical difference ( all P ＜ 0. 05 ). Total ended rate in modified Uchida technique were 2. 2/hundred women year in training group and 2. 5/hundred women year in non training group, and the rate of silver slips were 3. 9/hundred women year and 4. 8/hundred women year, which did not show significant difference ( all P ＞ 0. 05 ). There was no significant difference in acceptability and side effects of all women between two methods (P ＞ 0. 05). The training of service providers could influence acceptability of women (P ＜ 0. 05). Conclusions Clinical efficacy was not influenced by those two methods. The operative time and acceptability were improved by training surgeons in silver clips method.

BACKGROUND: The special genetic law of short tandem repeat on chromosome X (X-STR) makes it incomparable with autosome markers in forensic identification. However, the population genetics data is far less than that of the autosome STR, and especially the haplotype data are rarely reported.OBJECTIVE: To study the genetic polymorphism of 12 X-STR loci in Shenzhen area by pedigree analysis, aiming to provide scientific and effective data for the application of X-STR in forensic medicine and genetics. METHODS: The blood samples of 118 families were taken to extract DNA by Chelex-100, followed by PCR amplification using Investigator Argus X-12 kit. The frequency of alleles of 231 unrelated individuals was counted by direct counting method and Excel software. Hardy-Weinberg equilibrium test was performed on 12 X-STR loci of female samples by chi-square test. Discrimination power and mean exclusion chance were calculated according to the formula. Pedigree analysis was done to identify haplotypes of female samples and the haplotype frequencies of 4 linkage groups in 111 fathers and 119 mothers were calculated using direct counting method and Excel software.RESULTS AND CONCLUSION: In this study, 349 haplotypes were obtained. There were 238, 139, 153 and 157 haplotypes in linkage groups X1-X4, respectively. The polymorphism of DXS10135 locus was the highest with 21 alleles,while the polymorphism of DXS7423 locus was the worst with only 4 alleles. The combined discrimination power was 0.99999999 in males and 0.99999999 in females. The combined mean exclusion chance was 0.99999999 in trio cases,and 0.99999811 in duo cases. These findings indicate that the X-12 detection system has high polymorphism in Shenzhen Han population, and has important application value in forensic individual identification and paternity testing.

0.05).Conclusion Donepezil hydrochloride do not improve the learning and memory function of normal under age rats.

Objective To explore the effect of Acanthopanax senticosus injection (ASI) on brain multi-infract dementia rats. Methods Thirty-six male Wistar rats were randomly divided into 3 groups:control group, model group and ASI group. The multi-infract dementia rat models were set up by injecting mini-sludged blood in carotis internal arteries, learning and memory function of rats were determined by Morris water maze and Step-down test, the section of rat cerebral cortex and hippocampus were stained with haematoxylin eosin (HE). Results Compared with model group, latency in ASI group shortened significantly at 2nd,5th and 6th day, the distance shortened at 1st,2nd,5th and 6th day. The seeking tactics of ASI trgated rats improved in the Morris water maze test. The error times of ASI treated rats decreasd at 1st and 2nd day in Step-down test; ASI did not reduce significantly pathological changes of vascular dementia rats. Conclusion ASI has effect of treatment on multi-infract dementia rats.