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Resumen Antecedentes: Las alteraciones cromosómicas están presentes en 50 a 60 % de los abortos espontáneos y en 6 a 19 % de los mortinatos. Aunque se prefieren los microarreglos para estudiarlos, numerosos hospitales no pueden ofrecerlos. Objetivo: Presentar los resultados del estudio citogenético de 303 productos de la concepción (POC), 184 se obtuvieron de abortos espontáneos, 49 fueron mortinatos y en 17 no se identificó la de edad gestacional. Material y métodos: Se empleó cariotipo, hibridación in situ con fluorescencia, secuencias cortas repetidas en tándem y microarreglos, según el tipo de pérdida y la muestra disponible. Resultados: En 29 POC se encontró tejido materno, por lo que fueron eliminados de los análisis. En 250 (91.2 %)/274 casos se obtuvieron resultados informativos; la tasa de éxito del cariotipo fue de 80.7 %; la de los microarreglos de SNP, de 94.5 %; y la de la hibridación fluorescente in situ y la repetición corta en tándem, de 100 %. Se observaron anomalías citogenéticas en 57.6 % de los abortos espontáneos y en 24.5 % de los mortinatos; 94 % de las anomalías fueron numéricas y 6 %, submicroscópicas. Conclusiones: El cariotipo en conjunto con el estudio de secuencias cortas repetidas en tándem para descartar contaminación de células maternas es efectivo para estudiar abortos espontáneos; los microarreglos se recomiendan en los mortinatos.
Abstract Background: Chromosomal abnormalities are present in 50 to 60 % of miscarriages and in 6 to 19 % of stillbirths. Although microarrays are preferred for studying chromosomal abnormalities, many hospitals cannot offer this methodology. Objective: To present the results of the cytogenetic analysis of 303 products of conception (POC), which included 184 miscarriages, 49 stillbirths and 17 cases of undefined age. Material and methods: Karyotyping, fluorescence in situ hybridization, short tandem repeats and microarrays were used, depending on the type of loss and available sample. Results: In 29 POCs we found maternal tissue and were eliminated from the analyses. Informative results were obtained in 250 (91.2 %)/274 cases; the karyotyping success rate was 80.7 %; that of single nucleotide polymorphism microarrays, 94.5 %; and that of fluorescence in situ hybridization and short tandem repeat, 100 %. Cytogenetic abnormalities were observed in 57.6 % of miscarriages and in 24.5 % of stillbirths; 94 % of total anomalies were numerical and 6 % were submicroscopic. Conclusions: Karyotyping with simultaneous short tandem repeat study to rule out contamination of maternal cells is effective for studying miscarriages; in stillbirths, microarrays are recommended.
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ObjectiveTo screen out the transcriptomes related to the intervention of Wuzi Yanzongwan on the spermatogenic function of semi-castrated male mice， and to explore its potential mechanism in the intervention of the progress of low spermatogenic function. MethodBalb/c mice were randomly divided into sham-operated group， model group， testosterone propionate group（0.2 mg·kg-1·d-1， intramuscular injection） and Wuzi Yanzongwan group（1.56 g·kg-1·d-1， intragastric administration） according to body weight， with 12 mice in each group. The right testicle and epididymis were extracted from the model group and the drug administration group to construct the semi-castrated model of low spermatogenic function， while the fur and the right scrotum of the sham-operated group were only cut and immediately sterilized and sutured. At the end of the intervention， hematoxylin-eosin（HE） staining was used to observe the histopathology of testis， enzyme-linked immunosorbent assay（ELISA） was used to detect the levels of serum testosterone（T）， luteinizing hormone（LH） and follicle stimulating hormone（FSH）. The sperm count and motility of epididymis were measured by automatic sperm detector of small animal. Transcriptomic microarray technology was used to detect the mRNA expression level of testicular tissue in each group， the transcriptome of genes related to the regulation of Wuzi Yanzongwan was screened， and three mRNAs were selected for Real-time fluorescence quantitative polymerase chain reaction（Real-time PCR） to verify the transcriptome data. Through the annotation analysis of Gene Ontology（GO） and the signaling pathway analysis of Kyoto Encyclopedia of Genes and Genomes（KEGG）， the related functions of drugs regulating transcriptome were analyzed. ResultCompared with the sham-operated group， the testicular tissue of mice in the model group showed spermatogenic injury， contraction and vacuolization of the seminiferous tubules， reduction of spermatogenic cells at all levels， widening of the interstitial space， obstruction of spermatogonial cell development and other morphological abnormalities， and serum T significantly decreased， LH significantly increased（P<0.01）， and FSH elevated but no statistically significant difference， the count and vitality of epididymal sperm significantly decreased（P<0.01）. There were 882 differentially expressed mRNAs in the testicular tissues， of which 565 were up-regulated and 317 were down-regulated. Cluster analysis showed that these differentially expressed mRNA could effectively distinguish between the sham-operated group and the model group. Compared with the model group， the damage to testicular tissue in the Wuzi Yanzongwan group was reduced， the structure of the seminiferous tubules was intact， vacuolization was reduced， and the number of spermatogenic cells at all levels was significantly increased and arranged tightly. The serum T significantly increased， LH significantly decreased（P<0.01）， and FSH decreased but the difference was not statistically significant. The count and vitality of sperm in the epididymis were significantly increased（P<0.01）. Moreover， Wuzi Yanzongwan could regulate 159 mRNA levels in the testes of semi-castrated mice， of which 32 were up-regulated and 127 were down-regulated， and the data of the transcriptome assay was verified to be reliable by Real-time PCR. GO and KEGG analysis showed that the transcriptome functions regulated by Wuzi Yanzongwan were involved in the whole cell cycle process of sperm development such as sex hormone production of interstitial cells in testis， renewal， differentiation， metabolism， apoptosis and signal transduction of spermatogenic cells， and were closely related to the biological behaviors of signaling pathways such as spermatogenic stem cell function， endoplasmic reticulum protein processing and metabolic program. ConclusionWuzi Yanzongwan can effectively improve the low spermatogenic function of semi-castrated male mice， and its mechanism may be related to the regulation of testicular transcriptional regulatory network， the synthesis of sex hormones in testicular interstitial cells， the function of spermatogenic stem cells， the whole cell cycle process of spermatogenesis， as well as the expression of endoplasmic reticulum protein processing and metabolic program related genes transcription.
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Three carboline fluorescent probes F1-F3 were designed and synthesized, based on lead compound JYJ-19, an antifungal compound discovered previously by our group. The antifungal activity in vitro results showed that compound F1 had moderate antifungal activity (MIC80 = 32 μg·mL-1). The stokes shift of F1 is 70 nm. The fluorescent probe F1 has good optical properties and can be used for fluorescence imaging research. Subcellular localization experiments results showed that F1 was enriched in the mitochondria of fungal cells. The detection of intracellular reactive oxygen species levels shows that JYJ-19 enhances intracellular reactive oxygen species levels. The above results indicated that carboline compounds could exert antifungal effects by acting on fungal mitochondria.
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Biosensor analysis technology is a kind of technology with high specificity that can convert biological reactions into optical and electrical signals. In the development of drugs for Alzheimer's disease (AD), according to different disease hypotheses and targets, this technology plays an important role in confirming targets and screening active compounds. This paper briefly describes the pathogenesis of AD and the current situation of therapeutic drugs, introduces three biosensor analysis techniques commonly used in the discovery of AD drugs, such as surface plasmon resonance (SPR), biolayer interferometry (BLI) and fluorescence analysis technology, explains its basic principle and application progress, and summarizes their advantages and limitations respectively.
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@#his consensus aims to introduce the applications of 4K high-definition technology and fluorescence technology in thoracic surgery, summarize and categorize the technical support for pulmonary segment surgery, and innovatively propose technical support for precise sleeve resection of pulmonary segments. It provides a reference for clinical use, points out the direction for the research and innovation of domestically produced high-end endoscopes, promotes the widespread application of excellent domestically produced medical endoscopes, and facilitates the development of domestically produced medical equipment.
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@#Objective To prepare a national reference standard for the quantification of HEK293 cell DNA content,so as to provide a support for the determination of residual DNA in HEK293 cells in the industry.Methods HEK293 cell DNA prepared using Genomic-tip 500/G and genomic DNA purification reagents was used as source materials,and the purity and content were assessed using ultraviolet spectrophotometry and agarose gel electrophoresis.After dilution to approximately 100 ng/μL,the DNA was aliquoted at 160 μL/tube.Five different laboratories were organized for collaborative calibration by using ultraviolet spectrophotometry, and the stability and applicability were evaluated.Results The HEK293 cell DNA national reference standard exhibited A_(260)/A_(280) ratios between 1.8 and 2.0 and displayed a single band on electrophoresis,meeting the specified criteria.Collaborative calibration across five laboratories yielded 78 valid data points with an average content of 104.8 ng/μL,a relative standard deviation(RSD) of 4.2%.The 95% confidence interval for the mean was 103.8—105.8 ng/μL,and the 95% reference range for single measurements was 96.0—113.6 ng/μL.The average confidence limit rate was 1.0%,and the recommended storage condition was-80 ℃.Applicability studies were conducted using two different models of fluorescence quantitative PCR instruments.The reference standard exhibited good applicability within the range of 0.3—3 000 pg/reaction,with amplification efficiencies of 101% and 95%,and R~2 values of 0.999 2 and 0.999 5 for the standard curves,respectively.Conclusion This batch of HEK293 cell DNA national reference standard meets all required specifications and can be utilized as a national reference standard for fluorescence quantitative PCR detection,with a certified content of 104.8 ng/μL,assigned batch number 270039-202301.
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Abstract A simple, rapid, precise, accurate and sustainable spectrofluorimetric method (SFM) was developed, validated and applied for the determination of 4-aminobenzoic acid and aromatic amino acids (phenylalanine, tryptophan and tyrosine). These compounds are used in biopharmaceutical formulations and therefore must be analyzed by quality control laboratories to meet the criteria established in pharmacopoeias. In general, potentiometric titration (PT) is described in the compendia as the official analytical technique. However, this method showed low sensitivity and selectivity, and moreover was performed with a non-aqueous solvent (acetic acid), which led to higher consumption of reagents and consequently to the formation of residues. Therefore, the SFM was developed in aqueous medium at pH 7.2 using phosphate buffer. It was successfully validated according to the ICH guidelines and showed good linearity range (r>0.999), specificity, accuracy and precision (within and between days) and robustness. The test results were compared between the SFM and PT using raw material samples, while according to the F- and t-tests at 95% confidence level, no statistical difference was found between the methods
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Quality Control , Biological Products/classification , Spectrometry, Fluorescence/methods , 4-Aminobenzoic Acid/agonists , Amino Acids, Aromatic/adverse effectsABSTRACT
ABSTRACT Background: Recipients of a related haploidentical stem cell transplant (haplo-SCT) can have preformed antibodies to HLA donor's antigens. Objective: The aim of the study was to evaluate the engraftment rate and major clinical associations of anti-HLA donor-specific antibodies (DSA) at two mean fluorescence intensity (MFI) thresholds in recipients of an outpatient haplo-SCT. Methods: Seventy haplo-HCT recipients were analyzed. A virtual crossmatch was performed using the donor HLA typing and the recipient's anti-HLA DSA test results. Data for anti-HLA-A, -B, -C, and -DR were analyzed. Recipients with DSA ≥ 500 MFI were considered positive, and those with < 500 were considered negative; the same was adopted for MFI ≥ 1000. Results: Post-transplant infection was higher in recipients with DSA ≥ 500 MFI (84.6%, p = 0.041). First-year mortality was higher in DSA-positive patients ≥ 500 MFI, p = 0.004, and DSA ≥ 1000 MFI, p = 0.022, than in DSA-negative recipients. Graft failure in the first 100 days was not associated with DSA ≥ 500 or ≥ 1000 MFI. There was no difference in acute (a-GVHD) or chronic (c-GVHD) graft versus host disease between DSA-positive and negative patients. Conclusions: There was no association of anti-HLA DSA at MFI ≥ 500 and ≥ 1000 with graft failure, however, increased infection and 1st-year mortality were documented in related haplo-HCT at the MFI cutoffs studied. (REV INVEST CLIN. 2023;75(5):249-58)
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Saliva is a colourless, watery and complex fluid secreted by the salivary glands into the oral cavity. It plays an important role in maintaining the moisture and well being of the oral cavity. In recent years, Saliva has been used by the researches, as it is non invasive technique, limited training required, potentially valuable for children and elderly patients, cost effective, eliminates the risk of infection and screening in large population. Collecting dried salivary sample is still being a greater challenge in the forensic. The process of identifying a disease, condition, or injury from its signs and symptoms is referred as diagnosis and diagnostic tool helps in simplifying the clinical findings to obtain a reasonable and relevant differential diagnosis. Recently, Dried salivary spot has been most commonly used among Forensic Odontology which can be used as an efficient diagnostic aid. This article aims to review the updates and significance of dried saliva spot and sampling in forensic odontology.
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Introducción: El blanqueamiento global coralino refleja la fragilidad del coral a condiciones ambientales adversas y al cambio climático. Porites lobata es la especie de coral hermatípico más abundante actualmente en El Salvador y ha experimentado eventos de blanqueamiento recientes en el área natural protegida Complejo Los Cóbanos. Objetivo: Describir los episodios de blanqueamiento que colonias de P. lobata experimentaron en Los Cóbanos, El Salvador, durante los últimos 18 años, y analizar los esfuerzos de restauración de coral. Métodos: Aproximadamente 2 100 imágenes (2006 - 2022) tomadas sin periodicidad específica de dos sitios fueron revisadas. Parámetros del agua desde 2014 fueron registradas sin una periodicidad temporal definida. Las áreas de estudio fueron son playa El Faro y colonias submareales cerca de un importante resort de playa. La primera área presentó colonias dispersas en una zona intermareal, la otra, colonias más grandes a 2 m de profundidad en marea baja. También, revisar literatura acerca de actividades para restaurar arrecifes dentro del país. Resultados: Varios eventos de blanqueamiento ocurrieron durante este período de observación. Algunos de ellos podrían estar asociados a temperaturas altas del agua de mar (> 30 o C) de mayo a octubre (estación lluviosa), floraciones algales nocivas, así como causas desconocidas. Varios blanqueamientos que causaron pérdida de cobertura de coral en las colonias intermareales ocurrieron durante el evento global (2015-2016). Conclusiones: Blanqueamiento en el Complejo los Cóbanos ocurre en diferentes patrones de intensidad durante la estación lluviosa que comprende niveles de alta temperatura y turbidez en el agua, así como floraciones algales. El episodio de 2015-2016 causó pérdida de cobertura de coral en las colonias intermareales y las submareales experimentaron pequeños daños permanentes (< 5 % mortalidad). Las políticas de restauración nacionales requieren más esfuerzos estratégicos considerando la hidrodinámica y las capacidades institucionales locales.
Introduction: Global coral bleaching reflects coral fragility to adverse environmental conditions and climate change. Porites lobata is the most abundant hermatypic coral species currently in El Salvador and has experienced recent bleaching events at natural protected area Complejo Los Cóbanos. Objective: To describe the bleaching episodes that colonies of P. lobata experienced at Los Cóbanos, El Salvador, during the last 18 years, and analyze coral restoration efforts. Methods: Approximately 2 100 images (2006 - 2022) taken without specific periodicity from two sites were reviewed. Water parameters since 2014 were registered without defined frequency. The areas sampled were El Faro beach and subtidal colonies near an important beach resort. The first area presented dispersed colonies on an intertidal zone, the other, larger colonies at 2 m deep in low tide. Also, literature about activities to restore reefs within the country was reviewed. Results: Several bleaching events occurred during this observation period. Some of them could be associated to high sea water temperatures (> 30 o C) from May to October (rainy season), others to harmful algae blooms, as well as unknown causes. Severe bleaching that caused loss of coral coverage on intertidal colonies occurred during the global event (2015-2016). Conclusion: Bleaching at Los Cóbanos occurs in different intensity patterns usually during the rainy season that encompasses high temperature and turbidity levels in water, as well as algal blooms. The 2015-2016 episode caused coral coverage loss on the intertidal colonies and subtidal ones experienced little permanent damages (< 5% mortality). National restoration policies for coral colonies require more strategic efforts considering the local littoral hydrodynamics and institutional capabilities.
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This article for the first time presents the results of the study of qualitative and quantitative elemental and amino acid composition of the aboveground part of the plant Nepeta olgae Regel (L.) taken in the territory of Chust and Kosonsai districts (from the slopes of Gova and Kosonsai mountains) of Namangan region during the period before and during flowering (May-June, 2021-2022). The use of instrumental analysis of high-throughput energy dispersive X-ray fluorescence spectrometry, allowed to establish 20 mineral elements in the plant Nepeta olgae Regel (L.), among which to vital 9 elements and 3 to conditionally necessary. The amino acid composition of the plant Nepeta olgae Regel (L.) was studied by high performance liquid chromatography (HPLC) and 17 compounds were identified. Of these, 8 were substitutable and 9 essential amino acids.
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@#Objective To develop and validate the real-time fluorescent quantitative PCR(Q-PCR)method for the detection of 8 murine viruses. Methods The specificity,sensitivity and precision of the Q-PCR method were verified by four laboratories,and the virus simulated contamination test and blind sample detection were carried out simultaneously,of which the detection results were compared. The Q-PCR method was used to detect 26 batches of monoclonal antibody cell lines for SARS-CoV-2 vaccine production and 15 batches of other products of murine origin. Results The Q-PCR method used for detecting 8 kinds of murine viruses had no cross reaction with the same family and genus or other murine viruses. Except the sensitivity of laboratory 2 to ectromelia virus(EctV/Mouse Pox,MPV)was 2 × 10~2copies/μL,the sensitivity of laboratory 2 to other 7 viruses and 3 other laboratories to 8 murine viruses was 2 × 10~1copies/μL. Except the inter-assay CV of the copy number of mouse adenovirus(MAdV)detected by laboratory 3 was 37. 58%,the intra-assay and inter-assay CVs of the Ct and copy number of other 7 viruses detected by laboratory 3 and those of 8 viruses detected by other 3 laboratories were all less than 25%.The sensitivity of virus simulated contamination test met the parameter requirements. The coincidence rate of blind sample detection results by 4 laboratories was 100%. All the 26 batches of monoclonal antibody cell lines for SARS-CoV-2 vaccine production and 15 batches of other murine derived products were negative for 8 murine viruses. Conclusion Q-PCR method for murine virus has good specificity,sensitivity and precision,and can be used for the detection of murine derived biological products.
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@#Objective To develop a method for determination of activity and viability of Hansenula polymorpha by dual fluorescent staining with carboxyfluorescein diacetate(CFDA)and propidium iodide(PI).Methods The time durations(5,10,20,40,60 and 120 min for CFDA,5,10,15,20 and 25 min for PI)and working solution concentrations(50,100,200and 300 μg for CFDA,2,10,20 and 30 μmol/L for PI)for the dual fluorescent staining were optimized by single factor test. Under the optimal condition,the H.polymorpha samples at theoretical survival rates of 0,25%,50%,75% and 100%were determined,of which the fluorescent intensity was observed under fluorescent microscope,and the gray value was analyzed by ImageJ software. The live and dead cells were counted,based on which the actual survival and death rates were calculated. Meanwhile,the relationships of actual gray value,actual survival rate and actual death rate to the corresponding theoretical values were analyzed. Activity and viability of three batches of cultured H.polymorpha were detected by CFDA-PI dual fluorescence staining.Results The optimal time durations for staining with CFDA and PI were 60 and 5 min,while the optimal working solution concentrations were 200 and 2 μmol/L,respectively. The actual gray value,actual survival rate and actual death rate of H.polymorpha samples at various theoretical survival rates were significantly correlated to the corresponding theoretical values(R~2=0. 998 3～0. 999 2,P < 0. 05). The CVs of activity and viability values in three detections of three batches of H.polymorpha culture were 3. 20%～4. 03% and 1. 10%～2. 27%, respectively.Conclusion The CFDA-PI dual fluorescent staining was successfully developed,which may be used for determination of activity and vitality of H.polymorpha.
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@#Objective To express dengue virus(DENV)NS2B-NS3 protease in E.coli,optimize the expression conditions and determine the enzyme activity,so as to lay a foundation of screening and discovering of lead compounds targeting DENV.Methods Codon-optimized NS2B-NS3 gene was inserted into pET-28a vector to construct recombinant prokaryotic expression plasmid pET-28a-NS2B-NS3,which was transformed E.coli Rosetta(DE3)competent cells and induced by IPTG to express NS2B-NS3 protease. The optimal expression conditions of NS2B-NS3 protease in E.coli were determined by optimizing induction length,induction temperature and IPTG concentration. NS2B-NS3 protease was isolated and purified by HisTrap~(TM) affinity chromatography column and measured for the protease activity by fluorescence resonance energy transfer(FRET)assay.Results The recombinant prokaryotic expression plasmid pET-28a-NS2B-NS3 was constructed correctly as identified by restriction analysis(NheⅠ/XhoⅠ)and sequencing. The optimal expression conditions of NS2BNS3 protease in E.coli were as follows:induction temperature of 20 ℃,induction length of 10 h and IPTG concentration of0. 2 mmol/L. The purified NS2B-NS3 protease showed a purity of more than 90% with a exhibited a of 20 mg/L,which bound to mouse monoclonal antibody against His-tag specifically and had good hydrolytic activity with a specific activity of 16. 111 U/mg,a K_m of 16. 46 μmol/L and a k_(cat) of 0. 028/s.Conclusion DENV NS2B-NS3 protease with high purity and activity was successfully prepared,which laid an experimental foundation of the establishment of high-throughput screening model for inhibitors targeting NS2B-NS3 protease.
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@#Objective To develop and verify a multiplex fluorescence quantitative PCR(Taqman probe) method for the detection of telomerase activity.Methods Specific reverse transcription primers,two pairs of quantitative primers and probes were designed for the CDS sequence of telomerase catalytic subunit telomerase reverse transcriptase(TERT).After optimization of the reverse transcription primers(specific reverse transcription primers and random primers) and quantitative primers(two pairs of quantitative primer probes used alone or in combination) in the reaction system,with the primer probe of internal reference gene GAPDH,multiplex fluorescence quantitative PCR was performed in a single tube.In addition,telomerase positive standard and negative standard were prepared with 293T and MRC-5 cells respectively,and the stability and precision of the method were verified.The telomerase activity in 19 normal mesenchymal cell samples and 32 breast cancer cell samples were detected by the developed method.Results The optimum reaction system was as follows:using cDNA synthesized with specific reverse transcription primers as the template,2 pairs of quantitative primer probes of TERT gene were mixed with internal reference gene GAPDH primer probes for multiplex fluorescence quantitative PCR reaction in a single tube.After optimization,the sensitivity and TERT fluorescence signal quantity of the system were greatly improved,and the ΔRn was enhanced by 3 times.The amplification curve of positive standard TERT gene was normal,and the ΔCt between TERT gene and GAPDH gene remained stable.The amplification curve of GAPDH gene in negative standard was normal,while there was no amplification curve of TERT gene.There was a little difference in ΔCt between TERT and GAPDH genes in the positive standard frozen and thawed for 3 and 5 times repeatedly and the positive standard without freezing and thawing,and the CVs of precision in intra-and inter-groups were all less than 1%.Telomerase activity was negative in 19 normal mesenchymal cell samples and positive in 32 breast cancer cell samples,and significant difference in Ct value of TERT gene between them was observed(t=4.236,P <0.001).Conclusion The developed multiplex fluorescence quantitative PCR(Taqman probe) method for the detection of telomerase activity has good stability and precision,which is expected to be used in early diagnosis and gene therapy of tumors.
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@#Upconversion nanoparticles(UCNPs)doped with rare earth elements have advantages in biose-nsing because of their good fluorescence stability,biocompatibility and avoidance of background fluorescence. Therefore,fluorescence resonance energy transfer(FRET)system based on upconversion particles(UCNPs based FRET)has been widely used in biological detection. This paper reviews the application and prospect of UCNPs based FRET in biological detection of biotoxins,hormones,proteins,nucleic acids,bacteria,and so on.
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@#Objective To develop and verify a qPCR method for the qualitative and quantitative analysis of residual host DNA in human rabies vaccine(Vero cells) stock solution.Methods The qPCR standard curve was established by using the Vero cell DNA quantitative national standard,and the residual host DNA was extracted using magnetic beads.The specificity,repeatability,intermediate precision,accuracy and durability of the method were verified,and the linear range and limit of quantification were determined.The residual DNA of three batches of human rabies vaccine(Vero cells) stock solution was quantitatively analyzed and the fragment size was qualitatively analyzed by using this method.Results The correlation coefficients(R~2) of Vero cell DNA quantitative national standard amplification standard curve were all more than 0.99 by qPCR,and the quantitative range was 0.3 pg/mL～30 ng/mL.The method showed good specificity and repeatability.In the verification of intermediate precision,accuracy and durability,the relative standard deviations(RSD)of detection results of the samples were all less than 10%.The residual DNA content of Vero cells in three batches of stock solution was 0.20～0.77 ng/dose,which met the relevant standard of Chinese Pharmacopoeia(Volume Ⅲ,2020edition).The residual DNA fragments greater than 154 bp accounted for 52%～63%.Conclusion The developed qPCR method for the detection of residual DNA in human rabies vaccine(Vero cells) stock solution had good specificity,repeatability,intermediate precision and durability,and qualitatively and quantitatively analyzed the residual DNA rapidly and accurately,which was of great significance for improving the detection and control of residual DNA content in the production process and final product of human rabies vaccine(Vero cells).
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Objective:To investigate the application value of fluorescence imaging in single-port thoracoscopic anatomic segmentectomy.Methods:The clinical data of 280 patients (145 patients with fluorescence method and 135 patients with modified inflation-deflation method) who underwent thoracoscopic anatomic segmentectomy were retrospectively studied in the Anhui Chest Hospital from June 2020 to June 2021. There were 113 patients in the simple segmentectomy group and 167 patients in the complex segmentectomy group. The baseline data of the fluorescence method and the modified inflation-deflation method in the complex segmentectomy group were corrected by propensity score matching, and the perioperative results were compared between the groups.Results:There were no significant differences in segmental resection time, intraoperative blood loss, postoperative drainage, postoperative pain, postoperative extubation time, length of hospital stay, incidence of complications and cost of hand-holding between the fluorescence method and the modified method of the simple segmentectomy group.In the complex segmentectomy group, the time of segmental resection with the fluorescence method was significantly shorter than that with the modified inflation-deflation method( P<0.05), and other indexes had no significant difference. Conclusion:Fluorescence method single-port thoracoscopic anatomic segmentectomy has the same perioperative safety and short-term efficacy as modified inflation-deflation method, which can significantly shorten the operative time and improve the operative efficiency in complex anatomic segmentectomy.
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Objective:To investigate the performance of chromosome karyotype, chromosomal microarray analysis (CMA) and fluorescence in situ hybridization (FISH) in prenatal diagnosis of true fetal chromosome mosaicism. Methods:This retrospective study enrolled 40 women with true fetal chromosome mosaicism from 4 071 singleton pregnant women who were indicated for and underwent amniocentesis or/and cordocentesis in the the First Affiliated Hospital of Sun Yat-sen University from April 2018 to August 2021. The results of chromosome karyotyping, CMA and FISH, the types of chromosomal mosaicism, mosaicism ratio and pregnancy outcomes were analyzed using Chi-square test. Results:(1) The detection rate of true fetal mosaicism was 0.98% (40/4 071). (2) Sex chromosome mosaicism accounted for 42.5% (17/40). Other chromosomal mosaicism involved chromosomes 21, 22, 18, 16, 7, 12, 15, 17 and 20, as well as balanced chromosomal translocation. (3) The detection rate of true fetal mosaicism by chromosome karyotyping was 77.4% (24/31) from amniotic fluid samples and 10/19 from umbilical cord blood samples, while that data by CMA was 76.7% (23/30) and 7/11,respectively. (4) Of the 40 pregnant women with fetal chromosome mosaicism, FISH test was performed on 20 cases (14 cases were verified with both amniotic fluid and umbilical cord blood samples, five with amniotic fluid samples and one with umbilical cord blood sample), and all of the diagnosis of mosaicism were confirmed. For those with mosaicism ratio <30%, the detection rate by FISH was higher than that by CMA among amniotic fluid samples [14/19 vs 43.5% (10/23), χ2=3.88, P=0.049]. (5) Among the 40 pregnant women, five were lost to follow-up; 18 chose to terminate the pregnancy; and 17 continued the pregnancy to delivery. No abnormalities in mental or physical development were reported in the 17 neonates after birth or during on-line follow-up between 6 to 24 months old. Of the 14 pregnant women with mosaicism ratio <30% which confirmed by FISH, eight chose to continue the pregnancy, and no abnormalities in mental development or growth were found in the neonates. Conclusions:In prenatal diagnosis of true fetal choromosome mosaicism, the incidence of sex chromosome mosaicism is the highest. FISH may improve the prenatal diagnosis rate of mosaicism and is more accurate in determining the mosaicism ratio. The combination of FISH, CMA and chromosome karyotyping would significantly improve the detection rate of chromosomal mosaicism and assess the mosaicism ratio more accurately, which is of great value in clinical consultation and evaluation of fetal prognosis.
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Objective:To explore the effects of fluorescence method and improved expansion and collapse method on cardiopulmonary function, immune function and safety during segmental pneumonectomy.Methods:One hundred and fifty-six patients with lung segmental resection operation in Yucheng People′s Hospital from March 2018 to August 2020 were enrolled. They were divided into two groups according to the random number table method, each group with 78 patients. The patients in the observation group were treated with fluorescence method and the patients in the control group were treated with modified expansion collapse method. The operation indexes, immune function and cardiopulmonary function of the two groups were compared, and the incidence of postoperative complications were counted.Results:The intraoperative time and the appearance time of intersegmental plane in the observation group were shorter than those in the control group: (105.33 ± 10.62) min vs. (139.46 ± 12.58) min, (15.46 ± 1.13) s vs. (724.56 ± 65.65) s, there were statistical differences ( P<0.05). After operative for 1, 7 d, the levels of 6 min walking distance, peak expiratory flow rate, percentage of forced vital capacity and percentage forced expiratory volume in 1 second in the observation group were higher than those in the control group: after operative for 1 d: (475.36 ± 10.29) m vs. (412.11 ± 13.33) m, (284.52 ± 10.33) L/min vs. (251.13 ± 12.89) L/min, (85.65 ± 3.21)% vs. (81.13 ± 1.43)%, (83.25 ± 2.55)% vs. (74.49 ± 2.53)%; after operative for 7 d: (510.23 ± 16.66) m vs. (488.33 ± 15.42) m, (302.13 ± 15.58) L/min vs. (285.12 ± 10.22) L/min, (93.46 ± 5.79)% vs. (88.44 ± 5.44)%, (92.25 ± 2.32)% vs. (85.54 ± 2.13)%, there were statistical differences ( P<0.05). After operative for 1, 7 d, the levels of CD 4+/CD 8+, CD 8+, CD 4+ in the observation group were higher than those in the control group: after operative for 1 d: 0.85 ± 0.10 vs. 0.52 ± 0.04, 0.305 ± 0.025 vs. 0.285 ± 0.012, 0.325 ± 0.021 vs. 0.304 ± 0.025; after operative for 7 d: 1.13 ± 0.10 vs. 1.02 ± 0.07, 0.324 ± 0.029 vs. 0.306 ± 0.023, 0.359 ± 0.024 vs. 0.332 ± 0.025, there were statistical differences ( P<0.05). The rate of postoperative complications in the observation group was lower than that in the control group: 2.56%(2/78) vs. 11.54%(9/78), there was statistical difference ( χ2 = 4.79; P = 0.029). Conclusions:Both the fluorescence method and the modified inflation-collapse method have certain therapeutic effects, but the combination of fluorescence method in segmentectomy can provide reliable technical support and reduce the impact on cardiopulmonary function.