Objective To acquire glucose transporter 4 (GLUT4) cDNA from tissues of human muscle by RT-PCR, and to clone and express GLUT4 cDNA in E.coli. Methods GLUT4 cDNA primers were designed first and ascertained by detecting through Genbank on NCBI and DNA Star, then by using specific primers, the subjective cDNA segment was acquired by RT-PCR from human tissue. After that, it was cloned into cloning vector of pGEM-3zf(-) and sequenced automatically. Finally the subjective cDNA was cloned into vector of pBV220 and expressed in E.coli. Results The desired DNA could be acquired from tissues of human muscle by RT-PCR using special primers, the acquired cDNA fragment could be cloned into vector of pGEM-3zf(-) and sequenced. The GLUT4 cDNA sequence was highly conserved. GLUT4 cDNA could be expressed in E.coli, too. Conclusion The entire GLUT4 cDNA can be acquired from human muscular tissue by RT-PCR, and it can be expressed in E.coli.

Objective To investigate the surgery method for early spontaneous posterior interosseous nerve entrapment,to observe the postoperative efficacy in treatment,to evaluate the surgical outcomes,and to provide a foundation for clinical choice of reasonable operation scheme.Methods 21 cases of early spontaneous posterior interosseous nerve entrapment received operation. 1 3 cases (1 8 arms) were treated by neurolysis, and 8 cases (8 arms )were treated by nerve grafts with small vessels wrapping operation;all the patients were followed up for 10 to 20 months;the finger extensor muscle strength and metacarpophalangeal joint activity were evaluated. Results The lesion extensor muscles were part of the denervation changes.Neurolysis muscle strength:14 arms were excellent(82.35%),2 arms were good,1 arm was fair,1 case was lost;extensor function:15 arms were excellent(88.24%),1 arm was good,1 arm was fair,1 case was lost.Nerve graft muscle strength:6 arms were excellent(75.00%),1 arm was good,1 arm was fair;extensor function:7 arms were excellent(87.50%),0 arm was good,1 arm was fair.Conclusion Neurolysis can have a good efficacy in treatment of early spontaneous posterior interosseous nerve entrapment without obvious degeneration.When the severe degeneration of entrapment nerve happens,a nerve graft surgical treatment is needed.

Objective: To establish a candidate reference method for serum calcium determination based on inductively coupled plasma mass spectrometry(ICP-MS). Methods: Establishment of methodology: serum samples collected from the hospital were diluted 100 times by 0.3% ultrapure nitric acid. Calcium standard solutions with different concentrations were added to serum samples matrix solutions. Serum calcium concentration was calculated according to the standard addition method using germanium (Ge) as internal standard. The linearity, precision, trueness of the performance of the established candidate reference method were evaluated, and a comparison with the reference method was made. Results: The linearity was good in serum calcium concentrations from 0.000 mmol/L to 20.400 mmol/L (the concentration after dilution from 0.000 mmol/L to 0.204 mmol/L) (R²>0.999 9);the within-run precision was from 0.22% to 0.47%, and the between-run precision was from 0.64% to 0.77%; the total precision was from 0.98% to 1.09%. The determination results of 3 concentrations of SRM 956d were all in the uncertainty range required bythe certificate. The biases were -0.16%, 0.04% and 0.23%, respectively. The established candidate reference method participated in and passed the 2017 External Quality Assessment Scheme for Reference Laboratories in Laboratory Medicine (RELA) comparison. The results of the established candidate reference method were consistent well with those of the reference method in the Joint Committee for Traceability in Laboratory Medicine (JCTLM) list. There was a good correlation between the established candidate reference method and the clinical routine electrode method. Conclusions The established candidate reference method for serum calcium determination based on ICP-MS is precise, accurate, with wide range of linearity.It is expected to play a role in tracing the quantities of serum calcium in clinical laboratories.

At present, nucleic acid testing technology has been widely used in clinical laboratory diagnosis. Conventional detection technique such as real-time PCR is complicated, time consuming, and dependent on specific instruments. Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR associated protein (Cas) system is an adaptive immune defense system against viruses in bacteria and archaea, which has been developed into a powerful technology for genome editing. Recently, the leading groups engaged in CRISPR have set up new tools for nucleic acid detection based on Cas13a, Cas12a and newly discovered protein-Cas14, which plays an important role in rapid diagnosis of infectious diseases, detection of gene mutations in cancer and genotyping. Since they are ultrasensitive, specific, rapid and cost-effective, it is expected to bring great potential for molecular diagnosis. In this review, the mechanism of CRISPR/Cas system and the principle, the applications of the newly-developed diagnostic platforms are introduced. What′s more, the advantages, limitations and prospects of the technologies are summarized.

Objective To establish an isotope dilution liquid chromatography tandem mass spectrometry method (ID-LC-MS) for quantification of serum apolipoprotein E and phenotyping. Methods Method establishment. Samples underwent denaturing, alkylation and trypsin digestion with addition of internal standards as isotope labelling arginine. SB-C18 column was used for the liquid chromatographic separation and mass spectrometry positive ion mode and multiple reaction monitoring were employed for quantification and phenotyping. Precision, accuracy and linearity were investigated for method evaluation. 40 serum samples from Shanghai Dongfang Hospital during Oct. to Dec., 2018 were used for method comparison between ID-LC-MS and immunoassay. Deming regression and Bland-Altman were used for method comparison analysis and SPSS 24 for linearity. Results Target peptides reached their releasing maximum within 4 hours and SE did at 3 hours. 3 phenotyping of ApoE were observed, such as E3/E3, E2/E3 and E3 / E4. The imprecision of IQC was 5.2 ％ . The relative bias for low and high levels of accuracy-based samples was 7.6 ％ and 3.6 ％, respectively. Deming regression showed the intercept with 95 ％ confidence interval (CI) was 6.44-11.44 (P<0.05 and the 95％ confidence interval for the slopewas 0.77-0.89 (P<0.05). The coefficient was r=0.97. The mean difference was - 2.95 mg / L with 95 ％ CI-4.26--1.65 mg/L. The linearity covered from 16.9 to 58.5 mg/L. Conclusion ID-LC-MS can be used to quantify serum apolipoprotein E and simultaneously detect its phenotyping.

Objective:To explore the differences of immune reconstitution between peripheral blood stem cell transplantation and umbilical cord blood transplantation.Methods:A total of 300 patients (aged 18 (8, 33), 163 males and 137 females) with malignant hematological diseases who received allogeneic hematopoietic stem cell transplantation in the First Affiliated Hospital of University of Science and Technology of China from January 2018 to March 2020 were enrolled in this study, including 255 cases of umbilical cord blood transplantation and 45 cases of peripheral blood stem cell transplantation. Multicolor flow cytometry was applied to analyze lymphocyte subsets of the percentages and absolute counts in the two donor types and peripheral blood of patients after receiving hematopoietic stem cell transplantation. The differences between the two grafts were compared, and the lymphocyte subsets results were evaluated at 1, 2, 3, 4, 6, 9, 12, and 18 months after transplantation. 18-month disease-free survival (DFS) within the 300 patients under the two transplantation types were retrospectively analyzed.Results:1. The proportion of NKT cells in peripheral blood group was significantly higher than that in cord blood group (2.79% vs 0.24%, P<0.001). 2. The proportion of helper T cells in the UCBT group was higher than that in the PBSCT group, as well as the counts 6 months after transplantation ( P<0.05). 3. The proportion of NK1 cells (3 rd to 9 th month) and count (4 th to 12 th month) in UCBT group were significantly higher than those in PBSCT group ( P<0.05). 4. NKT cells in the UCBT group were lower than those in the PBSCT group (proportion and count) throughout the monitoring process ( P≤0.001). 5. The proportion of DNT cells (within 1 year) and count (within 6 months) in the UCBT group were significantly lower than those in the PBSCT group ( P<0.05). Conclusions:Compared with the peripheral blood stem cell transplantation group, the umbilical cord blood transplantation patients had a faster rate of lymphocyte reconstitution, and patients received umbilical cord blood transplantation had a stronger ability of immune reconstitution and could achieve long-term hematopoiesis.

OBJECTIVE To establish the project approval evaluation system for traditional Chinese medicine （TCM） preparations in medical institutions guided by new drug conversion， to improve the success rate of approval for TCM preparations in medical institutions and lay the foundation for the later drug conversion. METHODS Research and development team used the literature research method and brainstorming method to list and organize relevant elements of project evaluation and determine the initial indicator system. Experts were consulted using the Delphi method to confirm the evaluation index. The weights were calculated based on the proportion of importance scores for each indicator and assigned specific scores to each item. The indicator system was used to evaluate 31 TCM preparations applied for filing by various departments of our hospital from April to July 2023. RESULTS After two rounds of 17 experts’ consultation， the final TCM preparation system included five primary indicators， i.e. theoretical basis， clinical research foundation， pharmaceutical foundation， prescription， and clinical value， as well as 17 secondary indicators including prescription source， traditional Chinese medicine theory， clinical positioning and so on. Human experience was considered as the item which would be rejected as one vote. Based on the above indicator system， our hospital further improved the filing and project approval process for TCM preparations in medical institutions. Among the 31 TCM preparations applied for filing by various departments from April to July 2023， 8 TCM preparations with a score ≥65 were selected for development. CONCLUSIONS The evaluation system is objective， comprehensive， and highly operable. It is suitable for the selection of TCM preparations in medical institutions before research and development.