The Proposal for the Standardization of Urinary Sediments Examination and the expert consensus have provided effective and timely instruction on urinalysis normalization,but there are still some puzzles about the methods and requirements on quality improvement for urinalysis normalization.This paper is discussing about some issues referring to American and European urinalysis guidelines,which include establishment of examination procedure,specifications and methods of testing system evaluation,specimen stability,the influence of sample centrifugation,the usage of sample staining,confirmation tests,requirements on result reporting,internal quality control and external quality assessment.These discussions will contribute to the quality improvement of urinalysis in clinical laboratories.

Objective To investigate current status and problems of coagulation factor Ⅷ and Ⅸ assay in domestic laboratories so as to provide the reference for implementing the standardization and quality improvement.Methods A questionnaire survey was carried out in 76 laboratories,and quality control materials were distributed to 54 laboratories for activity assay.The questionnaire information was analyzed statistically.Test results of quality control materials were classified into three groups according to the reagents and the ranked grading analysis were used to evaluate the performance.Results This research was investigative study.The amount of sample was less than 30 per month in 72％ (52/72)of laboratories.The frequencies of calibration were different,and 33％ (24/72) of laboratories did not perform calibration in a different assay batch.39％ (28/72)of laboratories did not run internal quality control,and about 21％ (15/ 72) of laboratories just performed the normal level quality control.Individual laboratories showed a high cumulative CV (＞ 30％) of intemal quality control.For normal FⅧ and FⅨ control materials,the CV of results were 11.3％-18.2％ and 11.3％-17.9％ respectively as well as 15.3％-20.3％ and 19.5％-21％ for abnormal.Of the three groups,the proportions of laboratories which the FⅧ test results out with consensus were18％,24％ and 22％ as well as 20％,24％ and 28％ for FⅨ.Conclusions The key requirements for quality control of coagulation factors active assay remain to be addressed and implemented.The repeatability and comparability in some laboratories are not satisfactory to meet the clinical needs.With the purpose of promoting quality improvement,we need to develop guidelines,organize related training and establish a national external quality assessment scheme.

BACKGROUND: Cord blood stem cells are one of ideal target cells for gene therapy, but low gene transferring rate is the main difficulty at recent. Janus kinase tyrosine 2 (JAK2) plays an important role in self-renewing of cord blood stem/progenitor cell12s. Therefore, cord blood CD34+ cell line modified by target-amplified JAK2 genes has been developed yet by using gene regulating expression technique in order to overcome low transferring rate of cord blood genes.OBJECTIVE: To investigate the feasibility and reliability of a long-term amplified regulation for cord blood stem/progenitor cells mediated by transgene JAK2. SETTING: Department of Hematology, Beijing Hospital, Ministry of Health.MATERIALS: The experiment was carried out in the Laboratory of Hematological Department, Beijing Hospital, Ministry of Health from June 2003 to April 2006. Cord blood was derived from umbilical cord which was immediately cut from healthy, full-term and natural-parturition infants and was provided by Department of Obstetrics & Gynecology, Beijing Hospital. The experiment was approved by the local ethical committee, and informed consent was obtained from expectant mothers and their relatives for the use of cord blood cells. MiniMACS magnetic separation apparatus and immunomagnetic beads adsorbing CD34 single antibody were provided by Miltenyi Biotec Company, Germany; flow cytometer by FACScalibur, USA; recombinant human stem cell factor (rhSCF), Flt3 ligand (FL), human interleukin-6 (hIL-6), granulocyte macrophage colony stimulating factor (GM-CSF), granulocyte colony-stimulating factor (G-CSF) and thrombopoeitin (TPO) by PeproTec Company; nude mice of the SPF level by Animal Center of Beijing Medical University.METHODS: Retroviral vector MGI-F2JAK2, which was composed of functional catalytic domain of JAK2 genes and two site proteins (2xF36v, F2) combined with synthetic drug (AP20187) of target gene of small molecules, was constructed. AP20187 might specially combine with F36v to cause dimerization of JAK2 so as to activate signal conduction in cells. In addition, the vector included green fluorescence protein reporter gene, which was regarded as a label to detect proliferation. MiniMACS magnetic separation apparatus was used to purify and separate cord blood CD34+ cells. While, retrovirus supernatant including JAK2 was used to transfer cord blood CD34+ cells. After transduction, CD34+ cells were cultured with stem cell factor (SCF), Flt3 ligand, TPO and IL-6 and divided into control group (not adding AP20187) and experimental group (AP20187).MAIN OUTCOME MEASURES: ① Flow cytometer was used to detect percentage of green fluorescence protein reporter gene in the CD34+ cells and to determine gene transferring rate. ② Colony culture results of cord blood stem/progenitor cells after amplification. ③ Nude mice were given subcutaneous injection of ten-week cultured cord blood CD34+ cells at costa and neoplasia was observed after 30 days. RESULTS: ① Plentiful amplification of CD34+ cells was observed in both experimental group and control group. With the culture time passing by, positive rate of gel-filtered platelet of amplified CD34+ cells in the experimental group was gradually increased based on the basic level and more than 95% in the 11th week; however, positive rate of green fluorescence protein reporter gene in the control group was gradually decreased below the basic level and disappeared finally. ② Transgenic CD34+ cells in the experimental group still could generate brust forming unit-erythroid (BFU-E), colony-forming units granulocute/monocyte (CFU-GM) and multipotential hematopoietic progenitors (CFU-Mix); especially, CFU-GM was the main cell in hemopoietic progenitor cell (HPC). ③ Nude mice did not have neoplasia. CONCLUSION: Human cord blood CD34+ cells of transferring JAK2 genes may cooperate with other cytokines to amplify cord blood stem/progenitor cells in vitro for long. Therefore, this is potentially valuable for stem cells to treat some hereditary hematologic disease.

Objective To prepare and evaluate the reference materials for plasma von Willebrand Factor antigen testing with fresh frozen plasma.Methods The candidates were prepared by low temperature centrifugation in 5 different concentration levels.The homogeneity and stability of the preparation was evaluated according to the ISO Guide35 and CNAS-GL03.The comparability between STAGO and IL system was evaluated according to the WS/T 356-2011.Then the preparations were characterized by six laboratories with the Secondary Coagulation Standard established by NIBSC(SSCLOT4).Results Homogeneity evaluation of the preparation showed that there was no statistically significant difference between the groups (P >0.05),the F values of factor analysis of variance were 0.317~0.844,the uncertainty range was 1.01% ~2.06%.A linear regression based on stability evaluation indicated that the linear trend (within 24 weeks)was insignificant (P >0.05). The uncertainty range of long-term (within 24 weeks)stability was 0.79% ~ 1.20%.The results of the preparations on STAGO and IL system were comparable.The certificated values of the candidates were range from 12.2% to 138.9% with uncertainties were 0.06%~0.09%,respectively.The range of combined standard uncertainty was 0.03% ~ 0.16% while the expanded uncertainty was 2.2%~6.7%.Conclusion The reference materials for von Willebrand Factor antigen testing were stable and homogenous with comparability between STAGO and IL.The method of characterization was accurate and reliable.

Objective To establish the national standard materials for haemiglobincyanide (HiCN) for the traceability assays of hemoglobin. Methods HiCN national standard materials were established according to the document of International Committee for Standardization in Haematology (ICSH). The standard materials were certificated according to ISO Guide 35, including homogeneity and stability. Then they were characterized by the calibrated spectrophotometer which can be traceable to National Institute of Standards and Technology (NIST). The international reference materials of HiCN were compared with the result of the WHO reference laboratory to confirm the reliability. Results The uncertainty of the HiCN standard materials was 0.000 4 g/L and the variation coefficient (CV) was 0.09%. The uncertainty of long-term stability was 0.000 6 g/L; the certificated value of the standard materials was 0.615 9 g/L with uncertainty of 0.000 4 g/L. The combined uncertainty was 0.000 9 g/L and the expanded uncertainty was 0.001 8 g/L when the cover factor was 2. The relative error was 0.08% between the result of the standard materials and the international certificated value. Conclusion The homogeneity and stability of the standard material is acceptable and the method of characterization is accurate and reliable.

A reference system for the completed blood count (CBC) have been established in National Center for Clinical Laboratory (NCCL) according to the standards published by International Council for Standardization in Hematology (ICSH) and International Organization for Standardization (ISO) in order to calibrate hematology analyzer.The contents of our study mainly include:(1)Establishment of calibration laboratory for CBC, which is the first calibration laboratory accepted by China National Accreditation Board for Laboratories in all medical laboratories.(2)We firstly set up the reference method for CBC in China. In addition, the data between NCCL and a foreign reference laboratory have been compared. (3)We have calibrated some instruments from routine laboratories by the fresh blood or calibrator valuated by the reference system, which acts as a new way to calibrate hematology analyzer.(4) A secondary standard assay system has been established and the data between it and the foreign reference laboratory have been compared chronically. The experience has been introduced to local laboratories in 26 provinces.(5) We have drafted out several documents about technical standard for laboratory medicine. The main institution of applications includes: local center for clinical laboratory, clinical laboratories for routine examination, institutions for identification to instruments and reagents, centers for disease control and prevention, and so on.

Objective To evaluate the accuracy and comparability of secondary hematology analyzer for platelet enumeration in order to determine the accuracy and reliability of assigned value of fresh blood.Methods The results between secondary standard hematology analyzer and the reference method of platelet enumeration of 40 specimens were compared according to the document from CLSI EP9-A2.The correlation and bias were calculated.At the same time,the results of secondary standard hematology analyzer between our laboratory and Japan reference laboratory were compared.The fresh blood from normal people was prepared to be used as calibrator after assigned value by secondary standard hematology analyzer.And 36 hematology analyzers were performed correctness validation and calibrated by 36 fresh bloods.Results The results of 40 specimens by secondary standard hematology analyzer and the reference method were ( 108 -326) × 109/L and( 110 -327 ) × 109/L respectively.Correlation coefficient between the secondary standard hematology analyzer and the reference method was 0.993.The bias between two methods was from -3.8％to 3.4％.The results of NCCL and Japan reference laboratory from 2009 to 2010 were( 185 -203) × 109/L and (185 - 198) × 109/L The bias range between our laboratory and reference laboratory in Japan was from - 1.4％ to 3.7％.The ranges of coefficient variations of two laboratories were from 2.0％ to 3.0％ and from 2.6％ to 3.4％,respectively.The biases of 20 hematology analyzers were from - 2.6％ to 2.1％ and they passed the correctness validation.The biases of 16 hematology analyzers were decreased from 3.4％ - 12.6％of pre-calibration to 0％ - 2.8％ of post-calibration.Conclusions The results of secondary standard hematology analyzer are assured to be accurate and comparable by the comparison of reference laboratories.It is feasible that fresh blood assigned value by secondary standard hematology analyzer can be used as calibrator for the hematology analyzer.

Objective To verify the reference interval for D-dimer assay and analyze the influence of age and gender on the ref-erence interval.Methods Inclusion criteria for reference individuals were established.60 healthy males and 63 females were enrolled and divided to three groups by age,including 20 to 39 years old group (20 males and 20 females),40 to 59 years old group (20 males and 23 females)and above 60 years old group (20 males and 20 females).Blood samples were drawn in cit-rate sodium anticoagulated tubes and D-dimer concentration was determined by three different coagulation analyzers using o-riginal reagents.According to CLSI guideline C-28-A3,the reference interval for each measurement system from reagent manufacturer was verified and the difference of D-dimer concentration between different age-group and sex-group was ana-lyzed using non-parameters tests.Results All reference intervals were verified for people under age 40,while one reference interval cannot be verified for people from 40 to 59 years old as same as one for people above 60 years old.D-dimer concen-tration increased with age and there was significantly different between 20~39 years old group and 40~59 years old group or above 60 years old group(P<0.05).There was only a significant difference between sex-group for people under age 60(P<0.05).Conclusion D-dimer concentration was associated with age and sex.For people under age 40,the reference inter-val from reagent manufacture can be verified and directly used in laboratory,while for people above age 60,the reference in-terval from reagent manufacture cannot be verified.The cause should be investigated and a new reference interval should be established separately when necessary.

The pediatric reference intervals in clinical laboratory play an important role in diagnosis of illness,therapeutic monitoring,prediction of prognosis and health evaluation.Compared with establishing reference interval for adults,there are more challenges to establish pediatric reference intervals.Therefore,the procedure and key technologies of direct method and indirect method are stated based on the characteristics of children population and pediatric,by which to define,transfer and validate pediatric reference intervals.This study will provide systematically methodological ideas for clinical laboratories to establish pediatric reference intervals.

Objective:To investigate the analytical performance verification protocols and performance specifications of CD34+cell enumeration by flow cytometry for clinical laboratories.Methods:According to international guidelines and National Health Standard of China, we designed the performance verification protocols of CD34 +cell enumeration (including percent count and absolute count) by flow cytometry. Four quality assessment materials, three leukapheresis products and three samples of peripheral blood were selected to verify the precision, linearity, carryover, trueness and accuracy of FACSCanto Ⅱ measurement system, and the assessment criterion was set according to the detection technologies of clinical laboratories. Results:The CVs of intra-run precision of percent count and absolute count were 2.5% to 8.9% and 3.0% to 9.0%; the CVs of inter-run precision were 2.8% to 10.5% and 3.8% to 9.9%, respectively. The slopes of linearity regression equation of low range (3.6/μl to 123.6/μl) and high range (113.2/μl to 1196.3/μl) were 0.993 2 and 0.965 2, and R2 were 0.999 6 and 0.993 9, and the biases were -8.67% to 0.22%. The carryover of percent and absolute count were 0.07% and 0.00%. When percent count≤0.2% or absolute count≤20/μl, the absolute biases of trueness were in the range of ±0.006% or ±0.5/μl, and the absolute biases of accuracy were in the range of ±0.02% or ±0.9/μl; when percent count>0.2% or absolute count>20/μl, the relative biases of trueness were in the range of ±5.65%, and the relative biases of accuracy were in the range of ±8.19%. The verification results met the assessment criterion set in this study. Conclusions:The performance verification protocols and assessment criterion formulated in this study not only conform to the recommendations of domestic and foreign guidelines, but also conform to state of the detection technologies of native clinical laboratories, which can be taken as a reference of performance verification for clinical laboratories.