In order to further improve and standardize the laboratory diagnosis methods of hemolytic disease of the fetus and newborn（HDFN）and improve the level of clinical diagnosis and treatment, we investigated the laboratory detection of HDFN in 57 medical institutions in China, performed a Meta-analysis of 26 studies including 32 932 maternal and fetal ABO incompatibility cases, and summarized the diagnosis and treatment strategies at home and abroad. We hope to put forward the existing problems of laboratory detection, and discuss new strategy for diagnosis and treatment for HDFN.

Objective We investigated the current status of immunohematology laboratory tests for hemolytic disease of the fetus and newborn（HDFN）in China, and provided the basis for the establishment of performance specifications, test standards and expert consensus to improve the ability of laboratory tests for HDFN. Methods We compiled “the questionnaire of laboratory tests for hemolytic disease of the fetus and newborn”and distributed to the domestic medical facilities. The contents of the investigation included prenatal and postpartum immunohematologic tests, methods and laboratory quality control. We collected the survey data for statistical analysis. Results Fifty-seven questionnaires were collected, and the results were as follows: The timing of prernatal immunohematologic tests were performed at different times in pregnancy, mainly at 10 to 20 weeks (38.6%); prenatal immunohematologic tests mainly included ABO and RhD blood group type (95.7%), pregnant woman's husband ABO and RhD blood group type (63.8%), irregular antibody screening (93.6%), antibody identification (83%) and antibody titers [anti-A/B antibody (83%), irregular antibody (53.2%)] and cell-free fetal DNA (cffDNA) (0%); reagents for destroying IgM antibodies were 0.1mol/L 2-mercaptoethanol (82.6%) and 0.01mol/L 2-fimbritol (23.9%); postpartum immunohematologic tests mainly included neonatal ABO and RhD blood group type (100%), direct antiglobulin test (98.1%), serum free antibody test (94.2%), red blood cell (RBC) antibody elution methods (96.2%); RBC antibody elution methods were heat elution (40%) and acid elution method (54%) ; the facilities that carried out laboratory quality control: 42.3% internal quality assessment, 50% external quality assessment, 98.1% standard operating procedures; reference standards for serum antibody (0%). Conclusion There is an urgent need to establish HDFN laboratory standard operating procedures, standardization methods, reference range and quality control monitoring, and develop expert consensus.

Objective To investigate the transfusion strategy and prognostic effectiveness of hemolytic disease of the fetus and newborn（HDFN）caused by anti-D antibody. Methods Ten patients diagnosed with HDFN due to anti-D antibody at our hospital between January 2015 and December 2019 were enrolled,all of whom were treated with compatible blood. Changes in blood routine indexes and blood biochemical indexes before and after treatment were observed. Results The values of Hb,RBC and Hct before treatment were（104.70±3.72）g/L,（3.01±0.14）×10¹²/L,（0.33±0.01）%,and these values after treatment were（136.60±2.40）g/L,（4.09±0.17）×10¹²/L,（0.40±0.01）%,respectively. The indexes after treatment was significantly better than those before treatment（P<0.05）. The values of TBL,DBL,IBL and LDH before treatment were（294.09±26.91）mol/L,（25.68±3.84）mol/L,（268.41±24.16）mol/L,（591.60±59.69）IU/L,and these values after treatment were（164.86±25.4）mol/L,（16.03±1.86）mol/L,（143.41±24.3）mol/L,（347.20±33.82）IU/L,respectively. The indexes after treatment was significantly better than those before treatment（P<0.05）. Conclusion When a transfusion is given,it is preferable for patients with HFDN to receive blood of the same ABO and RhD negative type,and its clinical effect after transfusion is good.

Objective The phenotype of 1,3-D-galactosyltransferase p.Arg187Cys mutation previously found in Chinese population is normal B type. This study aims to explore the possible molecular mechanism of AB subtype induced by this mutation. Methods We identified the blood group and serotype of a Chinese individual by immunoserology. The enhancer, promoter, all seven exons and flanking sequences of ABO gene were amplified by PCR, and the amplified products were sequenced. The 3D molecular model was constructed by chimera software to predict the effect of mutation on the structure of 1,3-D-galactosyltransferase. Results It was identified as ABx subtype by serological method. The ABO gene sequencing showed that this healthy individual was A1.02/B.03 genotype; Only c.559C>T（p.Arg187Cys）mutation was detected in exon 7 of allele B, and no mutation was found in the rest of ABO gene. The 3D molecular model analysis showed that p.Arg187Cys mutation changed the local hydrogen bonding network of 1,3-D-galactosyltransferase, which may decrease enzyme activity. When co-expressed with A gene, it showed weak B phen-otype under the competition of 1,3-N-acetylgalactosamine transferase. Conclusion The c.559C>T mutation of ABO*B gene leads to the formation of ABx subtype by weakening the activity of 1,3-D-galactosyltransferase and the competition of ABO*A gene products.

Objective Explore the molecular mechanism of the rare p phenotype in the P1Pk blood group system of the Yi family. Methods Blood groups of the proband and 33 family members were identified by serologic test. Rare blood group genotyping was determined by qRT-PCR, the α-1,4-galactosyltransferase gene and β-1,3-N-acetylgalactosyltransferase were analyzed by Sanger sequencing. Results The proband and his brother are both rare p phenotypes and the serum contains anti-PP1PK antibody that can reacts with all non-p phenotype erythrocytes and causes hemolysis, other pedigree members are the normal P1 or P2 phenotype. The gene sequencing revealed that 2 of the 34 Yi family members were homozygous 456-457 ins ACACCCC (Bank IT: MG812384) in the A4GALT gene coding region, while 6 of them were 456-457ins ACACCCC heterozygous, 7 of them were 109A>G and 987G>A heterozygous, and 3 of them were IVS+228C> T heterozygous. The 7 bp insertion caused the frame shift in reading frame to form a premature termination codon, thus generate an invalid alleles. All the family members showed the consensus B3GALNT1 genes with the standard sequence. Conclusion In Yi family, we identified two cases of p phenotype with a novel A4GALT 456-457 ins ACACCCC alleles. The homozygous mutation is the molecular basis of the p phenotype.

Objective To study the effect of temperature difference caused by different storage location ingas phase liquid nitrogen tank on the cryopreservation quality of umbilical cord blood（UCB）derived hematopoietic stem cells. Methods Twenty units of UCB hematopoietic cells in liquid nitrogen were selected as control group A, twenty units of samples preserved at the lowest temperature in the gas phase liquid nitrogen tank as experimental group B, and twenty units preserved at the highest temperature in the tank as experimental group C. After thawing and resuscitation, nucleated cell count, cell activity, CD34⁺% and CFU-GM were detected and one-way ANOVA was performed. Results There was no significant difference in the recovery rate of nucleated cells, the change rate of activity of nucleated cells, the recovery rate of CFU-GM and the recovery rate of CD34⁺ cells among the three groups.（P>0.05）Conclusion The temperature difference caused by storage position has no effect on the cryopreservation quality of UCB hematopoietic stem cells in the temperature range of no more than -188℃.

Objective In order to reduce the risk of infection, the blood samples of patients suspected or diagnosed with novel coronavirus (SARS-Cov-2) were incubated at 56℃ for 30 minutes before serological blood typing in some laboratories. The influence of this method on pre-transfusion detection has not been systematically evaluated. In this study, we investigated the hemolysis rate, erythrocyte A, B, D antigen agglutination intensity and antibody titer of blood samples after this procedure. Methods The blood samples were randomly collected from blood donors and clinical patients in Shanghai, including 20 cases of type A, 20 cases of type B, 10 cases of type AB, 20 cases of type O (all of them were RHD positive), 7 cases of ABO subtype and 6 cases of D variant. The level of free hemoglobin was measured to evaluate the hemolysis rate. The detection of blood typing, antigen-antibody neutralization and antibody titer was to evaluate the agglutination intensity of A, B and D antigens and the changes of anti-A and anti-B titers. Results After the blood samples were incubated at 56℃, the RBC hemolysis rate was 1.87%±2.02%, and the agglutination intensity of A and B antigens decreased. The D antigens in normal D positive individuals did not decrease, while in D-variant individuals decreased. The titer of anti-A and anti-B antibody in serum decreased. Conclusion It was showed that after blood samples were incubated at 56℃, there was a low degree of hemolysis which did not affect serological blood typing, the agglutination intensity of A and B antigens and anti-A and anti-B antibodies decreased which was not strong enough to affect blood typing tests and only ABO subtype antigens may be affected.

Objective IgG anti-D is the main cause of severe hemolytic disease in newborns. The continuous-flow antibody quantifier has been widely used in Europe, North America and Australia. In China it is usually tested and recorded by titer assay in pregnant women and patients' antiserum. This study mainly compares the correlation between the titer assay and the antibody quantification method, and studies whether the titer assay can represent the amount of antibody. Methods In this study, 17 plasma samples with anti-D titer greater than 64 were compared with the antibody quantification method. The titer method was performed by ORTHOVISION automatic blood type detector for automatic gel column detection. The automatic continuous flow antibody quantizer (WHS-D0120) was used for quantification. Results It was found that the correlation between antibody titer and antibody quantification was r=0.92（P<0.001）. The comparative study between the titer groups of n≥3 (256 groups, 256 groups, 2 048 groups), found that there was a statistically significant difference between the 256 and 512 groups（P<0.001）, and there was no statistical difference between the 256 and 2 048 groups, or between the 512 and 2 048 groups（P>0.05）. In the mononuclear monolayer assay, it was found that there were large phagocytosis differences between the four antibody samples of anti-plasma group with a titer of 256. Conclusion For the anti-D assessment and monitoring of Rh-negative patients and pregnant women's plasma, quantification is more accurate than the titer assay.

Objective To compare the effects of 0.2 mol/L dithiothreitol（DTT）, 0.2 mol/L 2-mercaptoethanol （2-ME） and 1% bromelin on the removal of CD38 antigen on the surface of red blood cells（RBCs）. Methods 2%~5% RBCs suspension was mixed with 0.2 mol/L DTT, 0.2 mol/L 2-ME, and 1% bromelin in a volume ratio of 1∶4 respectively, and then incubated at 37℃ for 30 minutes. Untreated RBCs were used as positive control. Indirect anti-human globulin test（IAT）was performed with the RBCs suspension and the plasma of patients with Daratumumab（Anti-CD38）. Results The results showed that no agglutination was found in DTT and 2-ME treated RBCs, that weak agglutination was observed under microscope in bromelin treated RBCs, and that 1+ agglutination was in positive control. It was found that hemolysis occurred in all treated RBCs during the IAT procedure, and that bromelin treated RBCs were shown most serious hemolysis. Conclusion All three reagents can remove CD38 antigen on the surface of RBCs, and the removal efficiency of 0.2 mol/L DTT and 0.2 mol/L 2-ME is better than that of 1% bromelin.

Objective To investigate the clinical effect and safety of autologous blood donation before spinal surgery. Methods A restropective analysis was performed on 134 cases of benign spinal diseases in Guangzhou First People's Hospital. They were divided into autologous blood group（n=77）and allogeneic transfusion group（n=57）.The length of hospital stay, hemoglobin (Hb) changes and the transfusion related adverse reactions of two groups were recorded and compared. Results All the patients in autologous blood group had no related adverse reactions during preoperative blood donation and transfusion. Compared to allogeneic transfusion group, blood transfusion volume [（347.85±66.72）mL v.s. （478.88±194.23）mL,P<0.001）], the total length of hospital stay [（19.3±8.8）d v.s.（25.4±11.7）d, P=0.002）]and the length of postoperative hospital stay（10.77±5.41d v.s. 16.43±8.41d,P<0.001）were significantly reduced in autologous blood group. There were no significant differences in hemoglobin [（115.30±12.94）g/L v.s.（110.35±16.71）g/L,P=0.075] and hematocrit（0.34±0.03 v.s. 0.33±0.05,P=0.088）one day after operation between two groups. Conclusion Preoperative autologous donation is safe and economical in spinal surgery, which could effectively decrease the volume of allogeneic transfusion and the length of hospital stays.

Objective To analyze the characteristics of thromboelastography in children with Kawasaki disease （KD）and provide laboratory evidence for its diagnosis and treatment. Methods A total of 110 children's KD cases in Beijing Children's Hospital from January 1,2016 to September 30,2016 were enrolled as the experimental group,and 60 children undergoing selective operation with normal coagulation function were recruited as the control group. The baseline results of TEG, blood coagulation and platelet count were retrospectively analyzed. TEG parameters include reaction time（R value）,clotting time（K value）,α angle,maximum amplitude（MA value）,estimated percentage of dissolution（EPL）,percentage of dissolution 30 minutes after MA（LY30）and comprehensive coagulation index（CI）. Results The levels of R and K value in the experimental group were significantly lower than those in control group（P<0.01）,and the levels of αangle,MA,EPL,LY30 and CI in the experimental group were significantly higher than those in the control group（P<0.01）. Conclusion Children with KD had increased coagulation factor activity,high fibrinogen content,high platelet number or hyperfunction,and fibrinolytic system was out of balance,which showing a hypercoagulable state overall.

Objective To ensure the safety of blood transfusion , we established an information system management model for the precise distribution of blood transfusion samples. Methods The patient information system and the clinical blood transfusion information management system were used to dynamically track the average delivery time of blood transfusion samples in ordinary and emergency (rescue) departments monthly, and the number of sample delivery errors and missed cases were observed. Results In 2016~2017, information network distribution was not implemented. In 2018~2019, information network dynamic tracking and distribution was implemented. The monthly average blood transfusion sample time for ordinary consultation was 45.0±3.1 min, 40.0±4.2 min, 20.0±3.5 min, 19.0±3.3 min, respectively. There is a statistical difference with the year (F=170.95, P<0.05); while the information network distribution was not implemented in 2016-2017, and the information network dynamic tracking and distribution was implemented in 2018~2019. The monthly average emergency (rescue) blood transfusion sample time was 16.0±2.4 minutes, 15.0±2.1 minutes, 10.0±3.1 minutes, 9.0±1.8 minutes, respectively, there were also statistical differences with the year (F=25.44, P<0.05). In 2016 and 2017, the information network was not implemented to dynamically track the number of delivery errors, missed deliveries and detentions of blood transfusion samples were 286 cases (accounting for 7.77‰) and 269 cases (accounting for 6.90‰) respectively; the establishment was applied in 2018 and 2019, The accurate distribution of blood transfusion samples in the information system did not cause any delivery error, missed delivery and detention, and there were statistical differences between them (χ²=613.49, P<0.05). Conclusion The establishment of dynamic tracking management of blood transfusion sample information network can improve the accuracy of distribution and work efficiency, and also realize the quality process control of clinical blood transfusion work.

Objective We conducted co-word analysis on transfusion-related adverse events in foreign literature to form a visualization map, and analyzed research hotspots and found out risks. Methods A total of 765 foreign literature studies on transfusion-related adverse events were statistically analyzed, and statistical analysis tools and knowledge visualization software used for literature bibliographic information were imported for research. Results Research in blood transfusion focused on four major areas, including haemovigilance, transfusion reactions, infection prevention and control, and complications. Conclusion Transfusion reaction, viral infections and other complications are still the current hotspots. It is necessary to establish a blood early warning system in China.

Objective To investigate the drug resistance phenotype of carbapenems resistance in Serratia marcescens strains. Methods From June 2018 to September 2019, 235 non-repetitive strains of Serratia marcescens were isolated from patients in Gulou Hospital affiliated to Medical College of Nanjing University. The drug sensitivity was detected by Vitek 2 Compact matched GN13 drug sensitivity board and disk diffusion method. The carbapenem phenotype was screened by modified carbapenem inactivation test（mCIM）and EDTA- modified carbapenem inactivation test（eCIM）. The carrying status of carbapenem gene was detected by PCR and sequencing analysis. Results Among 235 strains of Serratia marcescens, 53 strains（22.6%）were resistant to both meropenem and imipenem. Among the 53 carbapenem resistant strains, 23（43.4%）were sensitive and 21（39.6%）were intermediate to ceftazidine. The resistance rates to ceftriaxone, amtronan, ciprofloxacin and levofloxacin were more than 98%, and the resistance rates to amikacin and cotrimoxacin were 1.7% and 1.6%, respectively. All of them were positive for mCIM and 3 strains were positive for eCIM. Among the 53 strains, 51 strains had bla_KPC-2 gene, 1 strain had bla_NDM-1 gene, 1 strain carried both bla_KPC-2 andbla_NDM-1 genes, and no bla_IMI, bla_OXA48, bla_FOX, bla_IMP and bla_VIM genes were detected. Conclusion The detection rate of carbapenem-resistant Serratia marcescens was high in our hospital. Bla_kpc-2 gene is the main prevalent genotype in carbapenem-resistance in Serratia marcescens strains.

Objective To investigate the difference of serum urea（UREA）,creatinine （CREA）and urea removal rate（URR）between urease rate method and dilution mode and its effect on the evaluation of dialysis adequacy. Methods one hundred and forty-eight inpatients（UREA level basically uniform covering linear range） in our hospital were enrolled as group A and one hundred and thirty-four dialytic patients were enrolled as group B. The level of UREA and CREA levels in group A were detected in original concentration and 5-fold dilution mode. The levels of UREA in serum of group B before and after dialysis were detected by original concentration,3-fold dilution mode and 5-fold dilution mode respectively. We calculated the URR of each patient and compared the difference of urea,URR and dialysis adequacy evaluation. Results The results of full-range UREA detected by dilution mode were significantly different from those of original mode,and the relative error was positively correlated with the level of UREA（r=0.57,P<0.05）,while the CREA level has no effect（P>0.05）. In group B,the levels of UREA and URR detected by dilution modes were significantly higher than those of original mode （P<0.05）.The percentage of dialysis adequacy evaluated by dilution modes were significantly higher than that of original mode（P<0.01）,and there was no significant difference among different dilution modes（P>0.05）. Conclusion Urease rate method in dilution mode affects the detection results of UREA,which leads to the increase of URR in dialysis patients and reduces the accuracy of dialysis adequacy evaluation.

Objective To investigate the expression levels and clinical significance of serum miR-205 and miR-944 in patients diagnosed with cervical cancer. Methods One hundred sixty two patients with cervical cancer（cervical cancer group）, 85 patients with cervical intraepithelial neoplasia（CIN group）and 65 healthy women（control group）in our hospital were selected from January 2017 to December 2019. The expression levels of serum miR-205 and miR-944 in each group were detected, and the relationship between expression levels and clinicopathological characteristics was analyzed. The diagnostic value of miR-205 and miR-944 in cervical cancer was analyzed using receiver operating characteristic (ROC) curve. Pearson correlation analysis was used to analyze the correlation between miR-205 and miR-944. Results The serum levels of miR-205（1.25±0.61 vs 0.71±0.13, 0.14±0.10）and miR-944（0.82±0.38 vs 0.46±0.02, 0.03±0.01）in cervical cancer group were significantly higher than those in CIN group and control group (P<0.01). In cervical cancer patients, the expression levels of miR-205 and miR-944 were correlated to clinical stage, depth of invasion and lymph node metastasis ( P<0.05). The ROC curves showed that the optimal cut-off values of miR-205 and miR-944 for diagnostic tests of cervical cancer were 0.94 and 0.53, respectively. The area under the curve (AUC) of the two combined diagnosis of cervical cancer was the largest（0.926, 95% CI: 0.864-0.985）, with 92.0% sensitivity, 88.7% specificity. The correlation analysis showed that there was a positive relationship between miR-205 and miR-944 in cervical cancer patients（r=0.824, P<0.01）. Conclusion The expression levels of miR-205 and miR-944 in cervical cancer patients were significantly higher, and a combination of the two tests can help guide the diagnosis of cervical cancer.

Objective To determine the clinical significance of serum inflammatory markers and 25-hydroxyvitamin D in diagnosing and treating depression in patients with Alzheimer's disease （AD）. Methods Serum levels of inflammatory markers were measured by ELISA and 25 hydroxyvitamin D（25-OH-VD）by chemiluminescent immunoassay （CLIA） in 20 AD patients with depression,26 AD patients without depression and 24 eligible healthy volunteers. Participants were evaluated for cognitive function by the Mini-Mental State Examination（MMSE）scores. Results Serum levels of IL-6 and TNF-α in AD with depression group were significantly increased,and（25-OH-VD）was significantly decreased. There were significant differences between the three groups （P<0.01）. Pearson correlation analysis showed that MMSE score in AD with depression group was negatively correlated with the levels of IL-1β、IL-6 and TNF-α（r=-0.437、-0.371、-0.532, P<0.05）,and positively correlated with 25-OH-VD（r=0.512,P<0.05）. （25-OH-VD）was negatively correlated with IL-1β,IL-6 and TNF-α（r=-0.452、-0.441、-0.471, P<0.05）. Conclusion The marker levels of peripheral inflammatory IL-6 and TNF-α in AD patients with depression were significantly increased, and the level of（25-OH-VD）was significantly decreased. This may be closely related to cognitive impairment in AD patients.

Objective To explore the necessity and the value of Case-Based Learning (CBL) based on research study which applied in the teaching process of clinical blood transfusion laboratory medicine. Methods Traditional teaching was used in the control group, CBL was used in clinical case analysis in the experimental group. Questionnaire was used to test the effect of teaching class. Formative evaluation and final paper was also used to analysis the difference between the two groups. Investigation was applied to acquire the effect of practical sessions. Results Questionnaire showed that clinical case analysis was beneficial, and CBL was widely approved. Formative evaluation, final paper and investigation presented that the effect of teaching class in the experimental group was better than that in the control group. Conclusion CBL based on research study, used in the clinical case analysis may greatly promote undergraduates to study actively and help them grasp the basic theoretical knowledge in class.