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Objective:To evaluate the performance of an artificial intelligent (AI)-based automated digital cell morphology analyzer (hereinafter referred as AI morphology analyzer) in detecting peripheral white blood cells (WBCs).Methods:A multi-center study. 1. A total of 3010 venous blood samples were collected from 11 tertiary hospitals nationwide, and 14 types of WBCs were analyzed with the AI morphology analyzers. The pre-classification results were compared with the post-classification results reviewed by senior morphological experts in evaluate the accuracy, sensitivity, specificity, and agreement of the AI morphology analyzers on the WBC pre-classification. 2. 400 blood samples (no less than 50% of the samples with abnormal WBCs after pre-classification and manual review) were selected from 3 010 samples, and the morphologists conducted manual microscopic examinations to differentiate different types of WBCs. The correlation between the post-classification and the manual microscopic examination results was analyzed. 3. Blood samples of patients diagnosed with lymphoma, acute lymphoblastic leukemia, acute myeloid leukemia, myelodysplastic syndrome, or myeloproliferative neoplasms were selected from the 3 010 blood samples. The performance of the AI morphology analyzers in these five hematological malignancies was evaluated by comparing the pre-classification and post-classification results. Cohen′s kappa test was used to analyze the consistency of WBC pre-classification and expert audit results, and Passing-Bablock regression analysis was used for comparison test, and accuracy, sensitivity, specificity, and agreement were calculated according to the formula.Results:1. AI morphology analyzers can pre-classify 14 types of WBCs and nucleated red blood cells. Compared with the post-classification results reviewed by senior morphological experts, the pre-classification accuracy of total WBCs reached 97.97%, of which the pre-classification accuracies of normal WBCs and abnormal WBCs were more than 96% and 87%, respectively. 2. The post-classification results reviewed by senior morphological experts correlated well with the manual differential results for all types of WBCs and nucleated red blood cells (neutrophils, lymphocytes, monocytes, eosinophils, basophils, immature granulocytes, blast cells, nucleated erythrocytes and malignant cells r>0.90 respectively, reactive lymphocytes r=0.85). With reference, the positive smear of abnormal cell types defined by The International Consensus Group for Hematology, the AI morphology analyzer has the similar screening ability for abnormal WBC samples as the manual microscopic examination. 3. For the blood samples with malignant hematologic diseases, the AI morphology analyzers showed accuracies higher than 84% on blast cells pre-classification, and the sensitivities were higher than 94%. In acute myeloid leukemia, the sensitivity of abnormal promyelocytes pre-classification exceeded 95%. Conclusion:The AI morphology analyzer showed high pre-classification accuracies and sensitivities on all types of leukocytes in peripheral blood when comparing with the post-classification results reviewed by experts. The post-classification results also showed a good correlation with the manual differential results. The AI morphology analyzer provides an efficient adjunctive white blood cell detection method for screening malignant hematological diseases.
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The corona virus disease 2019(COVID-19) pandemic, caused by the severe acute respiratory syndrome coronavirus 2(SARS-CoV-2), has created an urgent need of scientific and effective biomarkers for the purpose of prevention and control. Currently, commonly employed viral nucleic acids, antibodies, and rapid antigen test detection technologies all exhibit a range of limitations, including restricted applicability, inadequate sensitivity and specificity. Plasma SARS-CoV-2 quantitative antigen, as an emerging biomarker, has garnered significant attention due to its potential clinical value in the diagnosis and management of COVID-19. This article comprehensively analyzes the principles and clinical applications of quantitative detection technology for plasma SARS-CoV-2 antigen. Additionally, it explores the challenges encountered in this field and provides insights into future prospects.
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Humanos , COVID-19/terapia , SARS-CoV-2 , Antígenos Virais , Teste para COVID-19RESUMO
The corona virus disease 2019(COVID-19) pandemic, caused by the severe acute respiratory syndrome coronavirus 2(SARS-CoV-2), has created an urgent need of scientific and effective biomarkers for the purpose of prevention and control. Currently, commonly employed viral nucleic acids, antibodies, and rapid antigen test detection technologies all exhibit a range of limitations, including restricted applicability, inadequate sensitivity and specificity. Plasma SARS-CoV-2 quantitative antigen, as an emerging biomarker, has garnered significant attention due to its potential clinical value in the diagnosis and management of COVID-19. This article comprehensively analyzes the principles and clinical applications of quantitative detection technology for plasma SARS-CoV-2 antigen. Additionally, it explores the challenges encountered in this field and provides insights into future prospects.
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Humanos , COVID-19/terapia , SARS-CoV-2 , Antígenos Virais , Teste para COVID-19RESUMO
Objective:To investigate the pathogenesis, selection of endovascular treatment (EVT) strategies, and efficacies of acute vertebrobasilar artery occlusion (AVBAO) of different lesion sites.Methods:One hundred and five patients with AVBAO, admitted to and accepted EVT in our hospital from February 2017 to September 2019, were chosen in our study. The data of disease onset, imaging findings, EVT status, perioperative complications, and prognoses of these patients were collected. According to DSA results, the involved lesions were divided into 4 sites: the upper segment of basilar artery (BA), the middle segment of BA, the lower segment of BA, and the intracranial segment of vertebral artery (V4 segment), and patients with tandem lesions would be recorded as distal lesions. The risk factors, EVT strategies, and prognoses 90 d after follow-up (modified Rankin scale [mRS] scores≤3: good prognosis) were compared in patients with 4 different lesion sites.Results:There were significant differences in etiological classifications and percentage of patients combined with atrial fibrillation among patients with 4 different lesion sites ( P<0.05). There was significant difference in proportion of patients accepted emergency stent implantation among patients with 4 different lesion sites ( P<0.05): those with lesions at the V4 segment had the highest proportion of patients accepted emergency stent implantation (79.55%), followed by those with lesions at the lower segment of BA (50.00%). There was significant difference in EVT time (the time from arterial puncture to successful recanalization of occluded vessels) among patients with 4 different lesion sites ( P<0.05): the EVT time in patients with lesions at the middle segment of BA was the shortest (87.5 [58.5, 130.8] min), and the EVT time in patients with lesions at the lower segment of BA was the longest (115.0 [81.0, 163.0] min). There was no statistical difference among patients with different lesion sites in good prognosis rate 90 d after follow-up ( P>0.05). Conclusion:The pathogenesis of patients with different AVBAO lesion sites is different, so different EVT strategies should be adopted.
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AIM:To analyze the correlation between optical coherence tomography(OCT)parameters and central retinal vein occlusion of macular edema secondary(CRVO-ME), and compare the clinical efficacy of ranibizumab combined with laser photocoagulation and ranibizumab alone in the treatment of CRVO-ME.METHODS:There were 43 case with 43 eyes of patients in CRVO-ME diagnosed in our hospital from January 2020 to December 2020 included in the present study and divided into two groups, namely A and B. Patients in group A were treated with ranibizumab combined with laser photocoagulation, while patients in group B were treated with ranibizumab alone. The structure of outer retina and “SAVE” scores were observed and estimated using OCT and fluorescein angiography(FFA)examination before and after the treatment at 1, 3, 6, 12mo, and then analyzed their correlation with best corrected visual acuity(BCVA, LogMAR). The BCVA, central macular thickness(CMT), intraocular pressure and average number of drug injections were also compared between the two groups before and after treatment.RESULTS:At 12mo after treatment, the BCVA in the OCT baseline external limiting membrane(ELM)intact group and baseline ellipsoid zone(EZ)intact group before and after treatment were significantly improved than those of the fracture group(0.47±0.16 vs 0.21±0.15, P=0.013; 0.44±0.20 vs 0.25±0.17, P=0.008). There was no statistically significant difference in BCVA changes between baseline RPE fracture group and RPE intact group(P>0.05). The number of patients with “S” and “A” at 1 score decreased significantly at 12mo after treatment in both groups, the BCVA of patients with “V” and “E” at 0 score before treatment was significantly improved than those patients at 1 score(all P<0.05). The BCVA and CMT of patients after treatment in groups A and B were both significant improved compared with before treatment(P<0.05). There were no significant differences in the BCVA and CMT in the number of drug injections between the two groups(P>0.05). In addition, there were no severe complications such as secondary glaucoma and endophthalmitis in both groups.CONCLUSION: Baseline status of ELM and EZ, presence or absence of vitreoretinal abnormalities(V), and focal leakage(E)could suggest the treatment efficacy of CRVO-ME. Ranibizumab in the treatment of CRVO-ME demonstrates prominent efficacy and great safety, and there was no better effect was observed when combined with laser photocoagulation.
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Central nervous system (CNS) fungal infections are challenging and difficult to diagnose and treat. This article introduces the high risk factors, pathogen spectrum and laboratory indicators that cause CNS fungal infection. As patients with CNS fungal infections are often accompanied by immunodeficiency, it is especially necessary for clinical early detection, early prevention, and early diagnosis, and timely and effective implementation of optimized diagnosis and treatment programs to prevent further deterioration of the disease.
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Humanos , Sistema Nervoso Central , Infecções Fúngicas do Sistema Nervoso Central/microbiologia , Infecções do Sistema Nervoso Central , Fungos , Fatores de RiscoRESUMO
Cerebrospinal fluid (CSF) is a colorless and transparent fluid filling the ventricular system, subarachnoid space, and central canal of the spinal cord, which could directly reflect diseases from the human central nervous system. As advanced technologies were used widely in CSF-based tests, they provided a new perspective for diagnosing, prognosis and treatment monitoring of central nervous system diseases based on CSF. The main challenges currently are resolving the biological variation of CSF markers, selecting appropriate technologies, and standardizing CSF-based analysis.
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Objective:To establish a disk (CD) microfluidic chip detection platform for the rapid detection of CALR-1 and CALR-2 mutations in patients with cerebral infarction, and summarize its clinical application value.Methods:Based on microfluidic technology and loop mediated isothermal amplification technology, a CD microfluidic chip detection platform for simultaneous detection of CALR-1 and CALR-2 gene mutations were established, and the sensitivity, specificity, repeatability and accuracy of the platform were verified. A total of 124 patients with cerebral infarction treated in Huashan Hospital, Shanghai Medical College, Fudan University from November 2019 to March 2021 were prospectively selected into the experimental group; and 80 healthy subjects were included in the control group. The CALR-1 and CALR-2 gene mutations in anticoagulant peripheral blood samples were detected by the CD microfluidic chip. Each chip could detect 4 samples at the same time and synchronously detect 3 indexes of each sample. The detection results could be obtained after isothermal amplification for 40 min. At the same time, sequencing method was used to verify the test results, and the consistency of the results of the two detection methods was compared.Results:Using this CD microfluidic chip platform, the synchronous amplification of 3 indexes in the sample could be completed within 40 min without the need of thermal circulation, and the whole detection process of the sample could be completed within 60 min. For samples with a high concentration of target nucleic acid, typical positive signals could be visualized after amplification for 10 min, and the test results would be available within 30 minutes after receiving the samples. The detection sensitivity of CD microfluidic chip method for CALR-1 and CALR-2 mutation load concentration was 1.0% and 0.5% respectively. Nonspecific amplification was not observed for the non-target nucleic acid samples, indicating the high specificity of this method. The coincidence rates of intra and inter batch repeatability were 100% (20/20) respectively. Two samples with CALR gene mutation were found in the cerebral infarction group, both of which were CALR-1 mutations (L367fs*46). There was no CALR-1 or CALR-2 mutation in the control group. The detection results of CD microfluidic chip method were completely consistent with the sequencing verification results (100% [204/204]).Conclusions:The CD microfluidic chip method could be used for the detection of CALR-1 and CALR-2 gene mutations in clinical samples of patients with cerebral infarction. This method has the advantages of high detection sensitivity, good detection specificity, fast detection speed and high detection flux, which is helpful to clarify the etiology of patients with cerebral infarction.
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Objective:This study has investigated the value of detecting cerebrospinal fluid (CSF) MyD88L265P mutation and interleukin-10 (IL-10) levels in the prognosis of PCNSL.Methods:We retrospectively analyzed the clinical data, CSF characteristics (including cytology, cell counting, total protein, and the level of cytokine IL-10) and treatment process of 39 PCNSL patients newly diagnosed by surgery and pathology (18 males and 21 females, aged 40-73 years) from August 2013 to December 2016 in Hua Shan Hospital North. MyD88L265P mutation was detected by digital PCR in 39 paraffin-embedded tissues and 35 cerebrospinal fluid samples. Log-rank test was used for univariate analysis and Cox regression for multivariate analysis to establish the prognosis model of PCNSL which might be related to PCNSL first progress-free survival (PFS) and overall survival (OS).Results:The median age of the 39 PCNSL patients was 59 years old, with 30.8% (12/39) intraocular involvement. The mutation rate of MyD88L265P in tissues and cerebrospinal fluid was 74.4% (29/39) and 40.0% (14/35), respectively. 51.9% (14/27) patients were observed with MyD88L265P mutation in both tissues and CFS. Univariate analysis showed that intraocular involvement, high level of IL-10 in CFS (≥45 pg/ml), and MyD88L265P mutation in CFS are factors significantly influencing median progression-free survival (mPFS) of patients ( P<0.05). Patients with intraocular involvement had shorter OS than those without involvement which was statistically significant ( HR=6.5,95% CI 1.7-47.3, P<0.05). And multivariate analysis showed that intraocular involvement ( HR=2.4, 95% CI 1.3-7.8, P<0.05) and CFS MyD88L265P mutation ( HR=2.1, 95% CI 1.1-5.7, P<0.05) were independent prognostic factors for PFS. Conclusion:The presence of intraocular involvement and MyD88L265P mutation in CFS indicated poor prognosis of PCNSL patients. High CSF IL-10 level was not an independent factor affecting prognosis.
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INTRODUCTION@#Fragile X syndrome (FXS) is the most prevalent X-linked intellectual disability (ID) and a leading genetic cause of autism, characterised by cognitive and behavioural impairments. The hyperexpansion of a CGG repeat in the fragile X mental retardation 1 (FMR1) gene leads to abnormal hypermethylation, resulting in the lack or absence of its protein. Tools for establishing the diagnosis of FXS have been extensively developed, including assays based on triplet-primed polymerase chain reaction (TP-PCR) for detection and quantification of the CGG trinucleotide repeat expansion, as well as determination of the methylation status of the alleles. This study aimed to utilise a simple, quick and affordable method for high sensitivity and specificity screening and diagnosis of FXS in institutionalised individuals with ID.@*METHODS@#A total of 109 institutionalised individuals at the Center for Social Rehabilitation of Intellectual Disability Kartini, Temanggung, Central Java, Indonesia, were screened in a three-step process using FastFrax™ Identification, Sizing and Methylation Status Kits.@*RESULTS@#Two samples that were classified as indeterminate with respect to the 41-repeat control at the identification step were subsequently determined to be non-expanded by both sizing and methylation status analyses. Two samples classified as expanded at the identification step were determined to carry full mutation expansions > 200 repeats that were fully methylated using sizing and methylation status analyses, respectively, yielding a disease prevalence of 1.83%.@*CONCLUSION@#Repeat expansion and methylation-specific TP-PCR is practical, effective and inexpensive for the diagnosis of FXS, especially in high-risk populations of individuals with ID of undetermined aetiology.
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The analysis of cerebrospinal fluid (CSF) facilitates the diagnosis and therapy of central nervous system (CNS) diseases. Compared to traditional methods, single-cell sequencing is conducive to analyze the cells composition and heterogeneity and discover scarce cells in CSF. Recent studies of single-cell sequencing in CSF has mainly focused on the neuroinfectious diseases, neurodegenerative and neuroinflammatory diseases, and leptomeningeal metastases (LM), reflecting the superiority and clinical value of single-cell sequencing in CNS diseases and providing new directions for the diagnosis and treatment of CNS diseases.
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OBJECTIVE@#To analyze the clinical data of patients undergoing intravenous sedation in oral and maxillofacial surgery, to understand the epidemiological characteristics, to evaluate the efficacy and safety of intravenous sedation for oral surgery, and to summarize our experience.@*METHODS@#We retrospectively reviewed the clinical data of patients undergoing intravenous sedation between January 2010 and December 2018 in the Department of Oral and Maxillofacial Surgery, Peking University School of Stomatology. The gender, age, source, disease types, the values of perioperative vital signs, the use of sedatives and analgesics, duration of surgery and sedation, effect of sedation during the operation and the postoperative anterograde amnesia were analyzed.@*RESULTS@#A total of 2 582 patients experienced oral surgery by intravenous sedation. The peak age was 3.5 to 10 years and between 21 to 40 years. Supernumerary teeth (38%, 981/2 582) and impacted third molars (30%, 775/2 582) were the major disease types, and other types of disease accounted for 32 percent (826/2 582). The values of heart rate(HR), mean arterial pressure(MAP), respiration rate(RR) and bispectral index(BIS) showed statistically significant differences at the time of before sedation, local anesthesia injection, surgical incision, 10 min after operation and the end of operation. In the study, 69%(1 781/2 582) cases received midazolam alone, 7%(181/2 582) cases received propofol alone, and 24% (620/2 582) cases received midazolam and propofol combined for intravenous sedation. Fentanyl (33%, 852/2 582)was the most common intravenous analgesic we used, followed by flurbiprofen axetil (23%, 594/2 582) and ketorolac tromethamine (6%, 157/2 582). Besides, 35% (907/2 582) patients didn't use any intravenous analgesic during the surgery. The average operation time was (31.2±20.8) min, and the average sedation time was (38.4±19.2) min. During the surgery procedure, most of the patients scored on a scale of 2 to 4 according to the Ramsay sedation score (RSS). The postoperative anterograde amnesia rates of local anesthesia injection, surgical incision and dental drill during surgery were 94% (2 431/2 582), 92% (2 375/2 582) and 75% (1 452/1 936).@*CONCLUSION@#Intravenous sedation on the oral and maxillofacial surgery is effective and safe, can make the patients more comfortable, and should be further promoted and applied.
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Humanos , Anestesia Dentária , Anestésicos Intravenosos , Hipnóticos e Sedativos , Midazolam , Propofol , Estudos Retrospectivos , Cirurgia BucalRESUMO
At present, the prevention and control of new coronavirus has entered a critical period. However, the use of quantitative real-time PCR (qRT-PCR)assays for the detection of viral nucleic acid, as a crucial diagnostic approach, has been doubted in clinical practice. Herein, we have reviewed the current status of epidemic prevention and control, latest development of detection technologies, disease characteristics, clinical sampling and transport. It has also discussed the factors that may affect the performance of viral nucleic acid detection, and suggested some effective methods to improve the detection performance of the assays.
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At present, the situation of COVID-19 epidemic prevention and control in China is gradually improving, but the situation of overseas import prevention and control remains difficult. The COVID-19 epidemic may exist for long due to the undetermined source of infection, the difficulty in completely cutting off the transmission route, and a large number of susceptible people. Therefore, prevention and control will be a long-term and arduous task, making it necessary to adhere to the principle of combining emergency response with regular prevention and control, coordinating the epidemic prevention and social-economic development in a balanced way. In retrospect, the epidemic has exposed the ambiguous positioning and unsatisfying hardware construction of hospital laboratory departments, and delayed intervention of laboratory experts in the public health treatment system of China. This paper reflects on the hospital laboratory departments′ problems during the anti-epidemic activities, and put forward suggestions to improve the future development of clinical laboratories in the national public health system.
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Objective By a sequencing panel consisting of 50 targeted genes, aiming at depicting the molecular landscape of ET, PV, and PMF, which are three major subtypes of MPN, to provide valuable information in the diagnosis and prognosis of MPN.Methods A retrospective study was conducted of 53 patients from Huashan hospital and Changhai hospital. All patients were diagnosed in accordance with the 2016 WHO diagnostic criteria for MPN, including 31 cases of ET(11 males, 20 females, median age 55 years), 17 cases of PV(12 males, 5 females, median age 65 years), and 5 cases of PMF(4 males, 1 females, median age 67 years), and underwent next-generation of DNA sequencing of their bone marrow or blood samples. The genetic analyses were performed on bone marrow or peripheral blood. Referring to COSMIC, dbSNP, Clinvar and other public databases, we analyzed the sequencing data, and elucidated the mutation profile of MPN patients, combining with their clinic information. Results In addition to the typical JAK2, CALR, and MPL mutations, pathogenic mutations in other 11 genes were detected, as well as 4 SNPs that confer individual susceptibility to MPNs (rs4858647, rs9376092, rs58270997, rs621940). The average rate of mutated genes was 2.3 genes per patient. In all patients (53 cases), the mutated genes detected were TET2, EZH2, ASXL1, MIR662, SF3B1, BARD1, DNMT3A, KIT, RUNX1, TP53, NRAS according to their mutational frequency. Conclusions Applying next-generation sequencing technology, multi-gene sequencing of a bunch of typical BCR-ABL-negative MPN patients can be performed at one time within 2 working days, and pathogenic mutations other than JAK2, CALR, MPL can be found, which has a bright prospection in clinic.
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Objective To evaluate the inter-bottle variations, stability in consumption and interchangeability of unassayed biochemistry serum controls. Methods Comparison between unassayed serum controls from a domestic "Pretrol?" and an international "Bio-Rad" manufacturer was conducted in department of laboratory, in May 2016, with Roche Cobas 8000, Roche Hitachi 7600 and Siemens 2400 modular analyzer. The inter-bottle variation was determined by monitoring the inter-batch variation of 10 bottles of control samples after eliminating the intra-batch variation from the same bottle. Stability in consumption was determined as the precision after 7 days storage under 2℃ to 8℃ or 30 days of storage under -20 ℃ since reconstitution. The interchangeability was determined as the precisionbetween the controls from different manufacturers for the same test. Results The inter-bottle imprecision of controls from domestic manufacturer for 13 biochemistry tests (CV concentration 1/CV concentration 2) were potassium (0.26%/0.42%), sodium (0.26%/0.21%), phosphorus (0.00%/0.62%), cholesterol (0.56%/0.54%), total protein (0.52%/0.33%), albumin (0.44%/2.00%), alanine aminotransferase (1.72%/0.57%),γ-glutamylaminotransferase (0.52%/0.62%), aspartate aminotransferase (3.10%/1.09%), lactate dehydrogenase (0.76%/0.91%), alkaline phosphatase (1.13%/0.97%), amylase (0.30%/0.39%) and glucose (0.00%/0.40%). The stability in consumption of the controls from the domestic manufacturer (CV concentration 1/CV concentration 2 under 2℃to 8℃storage;CV concentration 1/CV concentration 2 under-20℃ storage) were potassium (1.06%/0.36%; 0.74%/0.48%), sodium (0.49%/0.59%; 0.72%/0.65%), phosphorus (0.95%/0.80%;1.43%/0.84%), cholesterol (1.49%/1.58%;2.17%/1.80%), total protein (0.84%/0.75%; 1.60%/1.68%), albumin (1.33%/2.28%; 1.94%/2.43%), alanine aminotransferase (1.41%/0.51%;3.24%/1.60%) γ-glutamylaminotransferase (1.16%/1.16%; 2.85%/2.49%), aspartate aminotransferase (4.37%/2.14%; 2.99%/1.31%), lactate dehydrogenase (2.70%/2.54%; 3.84%/2.97%), alkaline phosphatase (2.63%/1.96%; 2.31%/2.10%), amylase (0.95%/2.19%; 1.58%/1.38%) and glucose (0.60%/0.48%; 1.41%/1.55%). The Inter-bottle variation and stability in consumption of biochemistry test unassayed controls from the domestic manufacturer were compatible for clinical assay according to the CV% specification from the Clinical Biochemistry Test Quality Requirement (WS/T 403-2012). The imprecision of the controls from both the domestic and international manufacturers (CVp concentration 1/CVp concentration2;CVq concentration 1/CVq concentration 2) were potassium (0.52%/0.46%; 2.39%/0.47%), sodium (0.30%/0.17%; 0.81%/0.47%), phosphorus (2.72%/1.11%;4.57%/2.07%), cholesterol (0.29%/1.38%;2.94%/1.81%), total protein (0.66%/2.46%;1.85%/2.54%), alkaline phosphatase (2.67%/4.66%;3.58%/8.55%), total bilirubin (5.71%/5.09%; 9.55%/7.41%), albumin (1.10%/2.61%; 4.79%/1.93%), alanine aminotransferase (6.42%/1.25%;5.74%/1.63%), γ-glutamylaminotransferase (2.27%/4.35%; 4.38%/0.74%), aspartate aminotransferase (0.56%/2.84%; 0.91%/2.11%) and lactate dehydrogenase (2.36%/2.47%; 3.10%/1.52%). The interchangeability of serum controls from domestic manufacturer was better than clinical serum samples. Conclusion The unassayed serum biochemistry test controls from domestic manufacturer are suitable for the intra-laboratory quality control and showed a promising compatibility for inter-laboratory quality control usage .
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OBJECTIVE@#To evaluate the synergy of the Burkholderia signaling molecule cis-2-dodecenoic acid (BDSF) and fluconazole (FLU) or itraconazole (ITRA) against two azole-resistant C. albicans clinical isolates in vitro and in vivo.@*METHODS@#Minimum inhibitory concentrations (MICs) of antibiotics against two azole-resistant C. albicans were measured by the checkerboard technique, E-test, and time-kill assay. In vivo antifungal synergy testing was performed on mice. Analysis of the relative gene expression levels of the strains was conducted by quantitative reverse-transcription polymerase chain reaction (qRT-PCR).@*RESULTS@#BDSF showed highly synergistic effects in combination with FLU or ITRA with a fractional inhibitory concentration index of ⪕ 0.08. BDSF was not cytotoxic to normal human foreskin fibroblast cells at concentrations of up to 300 μg/mL. The qRT-PCR results showed that the combination of BDSF and FLU/ITRA significantly inhibits the expression of the efflux pump genes CDR1 and MDR1 via suppression of the transcription factors TAC1 and MRR1, respectively, when compared with FLU or ITRA alone. No dramatic difference in the mRNA expression levels of ERG1, ERG11, and UPC2 was found, which indicates that the drug combinations do not significantly interfere with UPC2-mediated ergosterol levels. In vivo experiments revealed that combination therapy can be an effective therapeutic approach to treat candidiasis.@*CONCLUSION@#The synergistic effects of BDSF and azoles may be useful as an alternative approach to control azole-resistant Candida infections.
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Humanos , Antifúngicos , Farmacologia , Burkholderia cenocepacia , Química , Candida albicans , Fisiologia , Candidíase , Tratamento Farmacológico , Farmacorresistência Fúngica , Ácidos Graxos Monoinsaturados , Fluconazol , Farmacologia , Testes de Sensibilidade Microbiana , Triazóis , MetabolismoRESUMO
@#To synthesize the folic acid-alliinase conjugate(FA-Alliinase), and to study its targeting and antitumor activity on cervical cancer HeLa cells. FA-Alliinase I and FA-Alliinase II were synthesized by two methods. The couping ratios of two conjugates measured were 12 and 31, respectively. The FA-Alliinase II with high coupling ratio was selected and its structure was characterized preliminarily. The activity of alliinase retained about 50% in FA-Alliinase II determined by HPLC. The specific effect of FA-Alliinase II on HeLa cells was observed by confocal laser and flow cytometry. The antitumor activity of conjugate combined with alliin was determined by MTT, and IC50 of alliin was(127. 6±2. 3)μmol/L. This study provides a direct evaluation method for the synthesis and optimization of FA-Alliinase.
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Objective@#To evaluate the inter-bottle variations, stability in consumption and interchangeability of unassayed biochemistry serum controls.@*Methods@#Comparison between unassayed serum controls from a domestic "Pretrol®" and an international "Bio-Rad" manufacturer was conducted in department of laboratory, in May 2016, with Roche Cobas 8000, Roche Hitachi 7600 and Siemens 2400 modular analyzer. The inter-bottle variation was determined by monitoring the inter-batch variation of 10 bottles of control samples after eliminating the intra-batch variation from the same bottle. Stability in consumption was determined as the precision after 7 days storage under 2 ℃ to 8 ℃ or 30 days of storage under -20 ℃ since reconstitution. The interchangeability was determined as the precisionbetween the controls from different manufacturers for the same test.@*Results@#The inter-bottle imprecision of controls from domestic manufacturer for 13 biochemistry tests (CV concentration 1/CV concentration 2) were potassium (0.26%/0.42%), sodium (0.26%/0.21%), phosphorus (0.00%/0.62%), cholesterol (0.56%/0.54%), total protein (0.52%/0.33%), albumin (0.44%/2.00%), alanine aminotransferase (1.72%/0.57%), γ-glutamylaminotransferase (0.52%/0.62%), aspartate aminotransferase (3.10%/1.09%), lactate dehydrogenase (0.76%/0.91%), alkaline phosphatase (1.13%/0.97%), amylase (0.30%/0.39%) and glucose (0.00%/0.40%). The stability in consumption of the controls from the domestic manufacturer (CV concentration 1/CV concentration 2 under 2 ℃ to 8 ℃storage; CV concentration 1/CV concentration 2 under -20 ℃ storage) were potassium (1.06%/0.36%; 0.74%/0.48%), sodium (0.49%/0.59%; 0.72%/0.65%), phosphorus (0.95%/0.80%; 1.43%/0.84%), cholesterol (1.49%/1.58%; 2.17%/1.80%), total protein (0.84%/0.75%; 1.60%/1.68%), albumin (1.33%/2.28%; 1.94%/2.43%), alanine aminotransferase (1.41%/0.51%; 3.24%/1.60%) γ-glutamylaminotransferase (1.16%/1.16%; 2.85%/2.49%), aspartate aminotransferase (4.37%/2.14%; 2.99%/1.31%), lactate dehydrogenase (2.70%/2.54%; 3.84%/2.97%), alkaline phosphatase (2.63%/1.96%; 2.31%/2.10%), amylase (0.95%/2.19%; 1.58%/1.38%) and glucose (0.60%/0.48%; 1.41%/1.55%). The Inter-bottle variation and stability in consumption of biochemistry test unassayed controls from the domestic manufacturer were compatible for clinical assay according to the CV% specification from the Clinical Biochemistry Test Quality Requirement (WS/T 403-2012). The imprecision of the controls from both the domestic and international manufacturers (CVp concentration 1/CVp concentration2; CVq concentration 1/CVq concentration 2) were potassium (0.52%/0.46%; 2.39%/0.47%), sodium (0.30%/0.17%; 0.81%/0.47%), phosphorus (2.72%/1.11%; 4.57%/2.07%), cholesterol (0.29%/1.38%; 2.94%/1.81%), total protein (0.66%/2.46%; 1.85%/2.54%), alkaline phosphatase (2.67%/4.66%; 3.58%/8.55%), total bilirubin (5.71%/5.09%; 9.55%/7.41%), albumin (1.10%/2.61%; 4.79%/1.93%), alanine aminotransferase (6.42%/1.25%; 5.74%/1.63%), γ-glutamylaminotransferase (2.27%/4.35%; 4.38%/0.74%), aspartate aminotransferase (0.56%/2.84%; 0.91%/2.11%) and lactate dehydrogenase (2.36%/2.47%; 3.10%/1.52%). The interchangeability of serum controls from domestic manufacturer was better than clinical serum samples.@*Conclusion@#The unassayed serum biochemistry test controls from domestic manufacturer are suitable for the intra-laboratory quality control and showed a promising compatibility for inter-laboratory quality control usage.
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Objective@#To investigate the application value of cerebrospinal fluid circulating cell-free DNA (cfDNA) in the diagnosis and treatment of leptomeningeal metastases in non-small-cell lung cancer (NSCLC).@*Methods@#Twenty-five patients with leptomeningeal metastases of NCSLC from Fudan University Huashan Hospital North during the period from September 2017 to November 2018 were enrolled. All 25 patients were confirmed leptomeningeal metastases by cerebrospinal fluid cytology and immunocytochemical staining of cytokeratin(CK7), carcinoembryonic antigen(CEA), thyroid transcription factor-1(TTF-1) and Ki67. The cerebrospinal fluid cfDNA was extracted and genetic variation of 12 genes including epidermal growth factor receptor(EGFR), TP53 and anaplastic lymphoma kinase(ALK) was detected by next-generation sequencing [PlasAim TM gene non-invasive detection of lung cancer (12 gene) kit, Singlera Genomics].The application value of cerebrospinal fluid cfDNA in the diagnosis and treatment of leptomeningeal metastases of NSCLC was analyzed with the cfDNA mutation data and the clinical follow-ups.@*Results@#Morphologically typical lung cancer tumor cells with tumor immunochemistry markerCK, CK7 and CEA were found in the cerebrospinal fluid of all 25 patients. Next generation sequencing of cerebrospinal fluid showed that 96% (24/25) patients had at least one single nucleotide variation (SNV) or copy number variation (CNV). The EGFR and TP53 mutations were identified in 80% (20/25) and 48%(12/25) of the patients, respectively. In addition, patients with bone metastases had a higher rate of EGFR mutations than those without bone metastases (100% vs 64%, P<0.05). Changes in the mutant allele frequency of EGFR and TP53 in cerebrospinal fluid were consistent with patients′ disease progression parameters including neurological symptoms, imaging, and tumor biomarkers. The results indicate that genetic alteration of EGFR in cerebrospinal fluid cfDNA is an actionable biomarker for targeted therapy of leptomeningeal metastases of lung cancer.@*Conclusion@#Cerebrospinal fluid cfDNA accurately reveals the unique genetic background of leptomeningeal metastasis in NSCLC, showing great application value in the diagnosis and treatment of the leptomeningeal metastasis of NSCLC.