As the gold-standard method for single-cell analysis,flow cytometry enables high-throughput and multiple-parameter characterization of individual biological cells.This review highlights the demands for clinical flow c...As the gold-standard method for single-cell analysis,flow cytometry enables high-throughput and multiple-parameter characterization of individual biological cells.This review highlights the demands for clinical flow cytometry in laboratory hematology(e.g.,diagnoses of minimal residual disease and various types of leukemia),summarizes state-of-the-art clinical flow cytometers(e.g.,FACSLyricTMby Becton Dickinson,DxFLEX by Beckman Coulter),then considers innovative technical improvements in flow cytometry(including quantitative,spectral,and imaging approaches)to address the limitations of clinical flow cytometry in hematology diagnosis.Finally,driven by these clinical demands,future developments in clinical flow cytometry are suggested.展开更多
Objective:To investigate the effect of combined detection of serum carcinoembryonic antigen(CEA),cytokeratin 19 fragment(CYFRA21-1),cancer antigen 125(CA125),and neuron-specific enolase(NSE)in patients with lung cance...Objective:To investigate the effect of combined detection of serum carcinoembryonic antigen(CEA),cytokeratin 19 fragment(CYFRA21-1),cancer antigen 125(CA125),and neuron-specific enolase(NSE)in patients with lung cancer by fluorescence flow cytometry.Methods:From August 2019 to July 2022,200 patients with lung cancer diagnosed by pathology in our hospital were retrospectively analyzed.2 mL venous blood was collected in a fasting state and centrifuged to separate the serum(containing human chorionic gonadotropin antibody[anti-hCG antibody],hepatitis B surface antibody[anti-HBs antibody],and CEA).Results:The sensitivities of CEA and CYFRA21-1 detected via enzyme-linked immunosorbent assay(ELISA)were 100%,and the detection limits were 0.5 ng/mL and 0.1 ng/mL,respectively;the sensitivities of CA125 and NSE detected via flow cytometry were 100%,and the detection limits were 10 U/mL and 2 ng/mL,respectively.Compared with ELISA,the sensitivities of CA125 and NSE detected via flow cytometry were higher.When the concentration of CEA was 10-40 ng/mL,the sensitivities of the three markers CYFRA21-1,CA125,and NSE showed no significant changes(P>0.05);when the concentration of CEA was 40-80 ng/mL,the sensitivity of CEA significantly decreased(P<0.01),but the sensitivities of the three markers CYFRA21-1,CA125,and NSE showed no significant changes(P>0.05);when the concentration of CEA was 80-200 ng/mL,the sensitivities of all four markers showed no significant changes(P>0.05).Conclusion:Compared with the double-antibody sandwich ELISA,fluorescence flow cytometry has certain advantages,including high sensitivity,good precision,short detection time,low sample usage,and low medical cost;thus,it is worthy of clinical promotion.展开更多
Objective:To explore the value of flow cytometry(FCM)in detecting the level of exfoliated cells in pleural effusion in the differential diagnosis of non-small cell lung cancer and benign lung diseases.Methods:Clinical...Objective:To explore the value of flow cytometry(FCM)in detecting the level of exfoliated cells in pleural effusion in the differential diagnosis of non-small cell lung cancer and benign lung diseases.Methods:Clinical data of patients with non-small cell lung cancer who were hospitalized in Hebei hospital from June 2019 to March 2022 were collected.A total of 98 patients were included,and 63 patients with alveolar lung disease were screened during the same period,and the two groups of patients were analyzed.Results:Compared with alveolar lung disease group,FCM detection and analysis showed that the level of exfoliated cells in the pleural effusion of non-small cell lung cancer(NSCLC)patients was 99(3-969)/100,000,and patients with alveolar lung disease was 4(0~19)/100,000.Additionally,compared with the alveolar lung disease group,the level of exfoliated cells in the pleural effusion of patients with non-small cell lung cancer(NSCLC)was significantly increased(P<0.001).The diagnostic efficacy of FCM for detecting pleural fluid exfoliated cells in non-small cell lung cancer was assessed using ROC curves and using 95%CI(-11.1,-13.2)with a sensitivity of 0.75 and specificity of 0.94,and the diagnostic efficacy of FCM for detecting pleural fluid exfoliated cells in alveolar lung disease was assessed using 95%CI(-11.1,-13.2)with a sensitivity of 0.71 and specificity of 0.87.Conclusion:Flow cytometry has a wider range of clinical applications,simple operation,low cost,and high sensitivity,which makes it of great significance in disease diagnosis.展开更多
Flow cytometry(FCM) technique has been widely applied to estimating the genome size of various higher plants. However, there is few report about its application in algae. In this study, an optimized procedure of FCM w...Flow cytometry(FCM) technique has been widely applied to estimating the genome size of various higher plants. However, there is few report about its application in algae. In this study, an optimized procedure of FCM was exploited to estimate the genome size of two eukaryotic algae. For analyzing Alexandrium catenella, an important red tide species, the whole cell instead of isolated nucleus was studied, and chicken erythrocytes were used as an internal reference. The genome size of A. catenella was estimated to be 56.48 ± 4.14 Gb(1C), approximately nineteen times larger than that of human genome. For analyzing Gracilariopsis lemaneiformis, an important economical red alga, the purified nucleus was employed, and Arabidopsis thaliana and Chondrus crispus were used as internal references, respectively. The genome size of Gp. lemaneiformis was 97.35 ± 2.58 Mb(1C) and 112.73 ± 14.00 Mb(1C), respectively, depending on the different internal references. The results of this research will promote the related studies on the genomics and evolution of these two species.展开更多
Gaining a full understanding of the mechanisms of action of natural products as therapeutic agents includes observing the effects of natural products on cellular morphology,because abnormal cellular morphology is an i...Gaining a full understanding of the mechanisms of action of natural products as therapeutic agents includes observing the effects of natural products on cellular morphology,because abnormal cellular morphology is an important aspect of cellular transformations that occur as part of disease states.In this study a set of natural products was examined in search of small molecules that influence the cylindrical morphology of fission yeast Schizosaccharomyces pombe.Imaging flow cytometry of large populations of S.pombe exposed to natural products captured cell images and revealed changes in mean length and aspect ratio of cells.Several natural products were found to alter S.pombe’s morphology relative to control,in terms of elongating cells,shrinking them,or making them more round.These results may facilitate future investigations into methods by which cells establish and maintain specific shapes.展开更多
Background:Genetic selection in cattle has been directed to increase milk production.This,coupled to the fact that the vast majority of bovine artificial inseminations(AI)are performed using cryopreserved sperm,have l...Background:Genetic selection in cattle has been directed to increase milk production.This,coupled to the fact that the vast majority of bovine artificial inseminations(AI)are performed using cryopreserved sperm,have led to a reduction of fertility rates over the years.Thus,seeking sensitive and specific sperm biomarkers able to predict fertility rates is of vital importance to improve cattle reproductive efficiency.In humans,sperm chromatin condensation evaluated through chromomycin A3(CMA3)has recently been purported to be a powerful biomarker for sperm functional status and male infertility.The objectives of the present study were:a)to set up a flow cytometry method for simultaneously evaluating chromatin condensation and sperm viability,and b)to test whether this parameter could be used as a predictor of in vivo fertility in bulls.The study included pools of three independent cryopreserved ejaculates per bull from 25 Holstein males.Reproductive outcomes of each sire were determined by non-return rates,which were used to classify bulls into two groups(highly fertile and subfertile).Results:Chromatin condensation status of bovine sperm was evaluated through the combination of CMA3 and Yo-Pro-1 staining and flow cytometry.Sperm quality parameters(morphology,viability,total and progressive motility)were also assessed.Pearson correlation coefficients and ROC curves were calculated to assess their capacity to predict in vivo fertility.Sperm morphology,viability and total motility presented an area under the ROC curve(AUC)of 0.54,0.64 and 0.68,respectively(P>0.05),and thus were not able to discriminate between fertile and subfertile individuals.Alternatively,while the percentage of progressively motile sperm showed a significant predictive value,with an AUC of 0.73(P=0.05),CMA3/Yo-Pro-1 staining even depicted superior results for the prediction of in vivo fertility in bulls.Specifically,the percentage of viable sperm with poor chromatin condensation showed better accuracy and precision to predict in vivo fertility,with an AUC of 0.78(P=0.02).Conclusions:Chromatin condensation evaluated through CMA3/Yo-Pro-1 and flow cytometry is defined here as a more powerful tool than conventional sperm parameters to predict bull in vivo fertility,with a potential ability to maximising the efficiency of dairy breeding industry.展开更多
[Objectives]This study was conducted to establish a method for detecting the immunophenotypes of venous blood CIK cells in healthy donors and patients with triple-negative breast cancer or lung cancer by flow cytometr...[Objectives]This study was conducted to establish a method for detecting the immunophenotypes of venous blood CIK cells in healthy donors and patients with triple-negative breast cancer or lung cancer by flow cytometry,and to further analyze and discuss the proportion of cellular immunophenotypes such as CD3^(+),CD4^(+),CD8^(+),CD3^(+)CD8^(+),CD3^(+)CD56^(+)in CIK cells.[Methods]Human venous blood was drawn,then anticoagulated with heparin and isolated with lymphocyte isolation solution,and the relevant immunophenotypes were detected by flow cytometry after 21 d of culture.[Results]The expression of CD3^(+)CD8^(+)and CD3^(+)CD56^(+)in the venous blood CIK cells was significantly higher in healthy donors than that in triple-negative breast and lung cancer patients.[Conclusions]CD3^(+)CD8^(+)and CD3^(+)CD56^(+)CIK cells have high anti-tumor activity.展开更多
<strong>Objective:</strong> <span><span><span style="font-family:;" "=""><span style="font-family:Verdana;">To investigate the value of cell morph...<strong>Objective:</strong> <span><span><span style="font-family:;" "=""><span style="font-family:Verdana;">To investigate the value of cell morphology and flow cytometry in the diagnosis of Multiple myeloma (MM). </span><b><span style="font-family:Verdana;">Methods:</span></b><span style="font-family:Verdana;"> 166 newly diagnosed MM patients were retrospectively analyzed, and the proportion of plasma cells detected by flow cytometry and cell morphology in these patients was statistically analyzed. </span><b><span style="font-family:Verdana;">Results:</span></b><span style="font-family:Verdana;"> All patients were divided into three groups according to the proportion of plasma cells detected by cell morphology: group A (≥30%), group B (≥10% and <30%), and group C (<10%). 1) In all patients and group A, B and C, the proportion of plasma cells detected by cell morphology was all correlated with that detected by flow cytometry. 2) The proportion of plasma cells detected by cell morphology in group A and group B was higher than that by flow cytometry, while there was no significant difference in group C. 3) The sensitivity of plasma cells detected by flow cytometry in group A, B and C was 88.7%, 51.2% and 10%, respectively. 4) According to the immunophenotypic analysis, the immunophenotypic characteristics were similar, all of them were CDl9 </span></span></span></span><span><span><span style="font-family:;" "=""><span style="font-family:Verdana;">- </span><span style="font-family:Verdana;">CD38 + CDl38 + CD56 +/</span><span style="font-family:Verdana;">- </span><span style="font-family:Verdana;">CD45dim ~ </span><span style="font-family:Verdana;">-</span><span><span style="font-family:Verdana;">. </span><b><span style="font-family:Verdana;">Conclusion:</span></b><span style="font-family:Verdana;"> There is a certain correlation between the two detection methods. The quantitative detection of myeloma cells by cell morphology is better, and the qualitative detection by flow cytometry is better. Combined with the two detection methods, the detection rate of multiple myeloma can be significantly improved.</span></span></span></span></span>展开更多
Sperm capacitation involves functional changes, such as the removal or appearance of specific molecules and changes in the plasma membrane;the acrosome reaction (AR) is an exocytotic event induced by calcium influx, e...Sperm capacitation involves functional changes, such as the removal or appearance of specific molecules and changes in the plasma membrane;the acrosome reaction (AR) is an exocytotic event induced by calcium influx, enabling the spermatozoa to penetrate the zona pellucida. These processes can be achieved only if the spermatozoa have good viability;indeed, determination of sperm viability is used for the assessment of semen quality. Membrane integrity and mitochondrial activity are important viability parameters of spermatozoa and fluorescent techniques based on membrane permeability to dyes have been developed to determine these parameters. The aim of this work was to determine the viability of boar sperm (fresh, one hour of capacitation induction and 20 min of AR induction) by flow cytometry using propidium iodide (PI) (1.25 μg/mL) and rhodamine 123 (R123) (0.20 μg/mL). Aliquots of 5 × 105 sperm were incubated with each fluorochrome separately and simultaneously for 10 or 20 min, respectively, at 38℃. The proportion of labeled spermatozoa and their fluorescence intensities were measured using a flow cytometer. The fluorescence index (FI) with PI gradually increased during the incubation and we found significant differences between all the groups. With R123, the FI increased in the capacitated sperm but decreased in the acrosome-reacted sperm, with significant differences between the fresh and capacitated spermatozoa. Our results suggest that the increase in the R123 fluorescence intensity in capacitated spermatozoa is due to changes in the mitochondrial membrane activity because the spermatozoa experienced changes in membrane fluidity and flagellar activation during capacitation. The use of fluorochromes and flow cytometry is a good tool for monitoring many markers of sperm function. Although capacitation and AR processes have been well studied, there is still much information to be elucidated with regard to these complex processes.展开更多
Immunophenotyping is proving crucial to understanding the role of the immune system in health and disease.High-through-put flow cytometry has been used extensively to reveal changes in immune cell composition and func...Immunophenotyping is proving crucial to understanding the role of the immune system in health and disease.High-through-put flow cytometry has been used extensively to reveal changes in immune cell composition and function at the single-cell level.Here,we describe six optimized 11-color flow cytometry panels for deep immunophenotyping of human whole blood.A total of 51 surface antibodies,which are readily available and validated,were selected to identify the key immune cell populations and evaluate their functional state in a single assay.The gating strategies for effective flow cytometry data analysis are included in the protocol.To ensure data reproducibility,we provide detailed procedures in three parts,including(1)instrument characterization and detector gain optimization,(2)antibody titration and sample staining,and(3)data acquisition and quality checks.This standardized approach has been applied to a variety of donors for a better understanding of the complexity of the human immune system.展开更多
Nowadays,doctors and nutritionists recommend individuals incorporate selenium-rich foods such as nuts,cereals,and mushrooms into their regular diet to maintain fitness and overall health.Selenium nanoparticles(SeNPs)e...Nowadays,doctors and nutritionists recommend individuals incorporate selenium-rich foods such as nuts,cereals,and mushrooms into their regular diet to maintain fitness and overall health.Selenium nanoparticles(SeNPs)exhibit strong chemopreventive capabilities.The anticipations for SeNPs with enhanced and tunable bioactive activities have led to a keen interest in phytofabrication.In this study,the aqueous extract of Clerodendron phlomidis plant leaves was utilized for the synthesis of SeNPs.In traditional Indian medicine,this plant extract is recognized as a significant anti-diabetic agent.The flavonoids tetrahydroxylflavone,7-hydroxyflavanone,and 6,4’-dimethyl-7-acetoxy-scutellarein present in this plant leaf extract demonstrate excellent anticancer activity.These secondary metabolites exhibit the ability to reduce sodium selenite into SeNPs.At a concentration of 13μg/mL,the synthesized SeNPs effectively inhibited the proliferation of the HepG2 cell line.The results suggest that the SeNPs possess promising anti-cancer potential against liver cancer and can be considered as a therapeutic agent for liver cancer treatment.Additionally,the cell cycle arrest induced by SeNPs was further confirmed by the fluorescence-activated cell sorting(FACS)method,indicating that SeNPs could efficiently differentiate cancer cells from normal cells.Notably,it showed a significant improvement in diethylnitrosamine(DEN)-induced Swiss Wistar rat groups.This scientific investigation highlights the high anti-cancer potential of SeNPs,positioning them as a promising therapeutic agent for liver cancer treatment.展开更多
Early and accurate diagnosis and treatment of cancer depend on rapid,sensitive,and selective detection of tumor cells.Current diagnosis of cancers,especially leukemia,relies on histology and fl ow cytometry using sing...Early and accurate diagnosis and treatment of cancer depend on rapid,sensitive,and selective detection of tumor cells.Current diagnosis of cancers,especially leukemia,relies on histology and fl ow cytometry using single dye-labeled antibodies.However,this combination may not lead to high signal output,which can hinder detection,especially when the probes have relatively weak affi nities or when the receptor is expressed in a low concentration on the target cell surface.To solve these problems,we have developed a novel method for sensitive and rapid detection of cancer cells using dye-doped silica nanoparticles(NPs)which increases detection sensitivity in fl ow cytometry analyses between 10-and 100-fold compared to standard methods.Our NPs are~60 nm in size and can encapsulate thousands of individual dye molecules within their matrix.We have extensively investigated surface modifi cation strategies in order to make the NPs suitable for selective detection of cancer cells using fl ow cytometry.The NPs are functionalized with polyethylene glycol(PEG)to prevent nonspecifi c interactions and with neutravidin to allow universal binding with biotinylated molecules.By virtue of their reliable and selective detection of target cancer cells,these NPs have demonstrated their promising usefulness in conventional fl ow cytometry.Moreover,they have shown low background signal,high signal enhancement,and effi cient functionalization,either with antibody-or aptamer-targeting moieties.展开更多
This study aims to demonstrate the validity of fluorescence-based methods,together with flow cytometry,as a complementary tool to conventional physicochemical analyses carried out in wastewater treatment plants(WWTPs)...This study aims to demonstrate the validity of fluorescence-based methods,together with flow cytometry,as a complementary tool to conventional physicochemical analyses carried out in wastewater treatment plants(WWTPs),for the control of the currently largely unknown activated sludge process.Staining with SYTO 9,propidium iodide and 5-(and 6)-carboxy-2’,7’-difluorodihydrofluorescein diacetate(carboxy-H2 DFFDA)was used for cell viability and oxidative stress monitoring of the bacterial population forming the activated sludge of a WWTP.Throughout the period of research,several unstable periods were detected,where the non-viable bacteria exceeded the 75%of the total bacterial population in the activated sludge,but only in one case the cells with oxidative stress grew to 9%,exceeding the typical values of2%-5%of this plant.These periods coincided in two cases with high values of total suspended solids(SST)and chemical oxygen demand(COD)in the effluent,and with an excess of ammonia in other case.A correlation between flow cytometric and physicochemical data was found,which enabled to clarify the possible origin of each case of instability in the biological system.This experience supports the application of bacterial fluorescence staining,together with flow cytometric analysis,as a simple,rapid and reliable tool for the control and better understanding of the bacteria dynamics in a biological wastewater treatment process.展开更多
The assessment of neutralization activity is an important step in the evaluation of neutralizing antibodies (NAbs).The traditional methods for measuring the antibody neutralization of human adenovirus type 3 (HAd V-3)...The assessment of neutralization activity is an important step in the evaluation of neutralizing antibodies (NAbs).The traditional methods for measuring the antibody neutralization of human adenovirus type 3 (HAd V-3) are the microneutralization (MN) assay,which has insufficient sensitivity,and the plaque reduction neutralization test (PRNT),which is not suitable for high-throughput screening.Herein,we describe the development of a flow cytometry-based neutralization(FCN) assay for measuring the neutralization of sera,cell culture supernatants,and chimeric antibodies against HAd V-3 on the basis of a recombinant HAd V-3 (r HAd V-3) construct expressing the enhanced green fluorescent protein (EGFP).For flow cytometry-based assays,the optimal cell confluence was determined as 90%,and the virus was titrated using the assay.The established FCN assay follows the percentage law and an optimal MOI of not less than 5 9 10~(-4)was determined by using a purified chimeric antibody.In addition,comparison of the anti-HAd V-3 NAb titers of 72 human serum samples by the MN and FCN assays,showed that both assays correlated strongly with each other.Our FCN assay was an improvement over the MN assay because the observation period was reduced from 3 to 1 days and data analysis could be performed objectively and robotically.Importantly,the newly established FCN assay allows measurement of the neutralization activity of chimeric antibodies expressed in cell culture supernatants.Thus,this sensitive and highthroughput FCN assay is a useful alternative to the MN assay for measuring the antibody neutralization of HAd V-3 and for screening anti-HAd V-3 NAbs in cell culture supernatants.展开更多
Soil microhabitats and their heterogeneity are often considered to be among the most important factors affecting soil biotic communities.The microbial commu-nity has become one of the most important links in soil nutr...Soil microhabitats and their heterogeneity are often considered to be among the most important factors affecting soil biotic communities.The microbial commu-nity has become one of the most important links in soil nutrient cycles and trophic components due to its role in biological processes,spatial and temporal dynamics,and physiological adaptation.Sandy-soil desert systems are characterized by fast water infiltration during the rainy season,high salinity,and low moisture availability in the upper soil layers.Plants have developed different ecophy-siological adaptations in order to cope with this harsh environment.The Tamarix aphylla is known to be one of the most commonly adapted plants,exhibiting a mechan-ism for secretion of excess salts as aggregates through its leaves.These leaves aggregate beneath the plant,creating'islands of salinity'.Soil biotic components are,therefore,exposed to extreme abiotic stress conditions in this niche.The goal of this study was to examine the effect of T.aphylla on the live/dead bacterial population ratio on a spatial and temporal scale.The results emphasize the effect of abiotic factors,which changed on temporal as well as spatial scales,and also on the size of the active soil bacterial community,which fluctuated between 1.44%and 25.4%in summer and winter,respectively.The results of this study elucidate the importance of moisture availability and the'island-of-salinity'effect on the active microbial community in a sandy desert system.展开更多
Objective:Dianjixueteng is a geoherb in Yunnan Province,the source plant of which is Kadsura interior.However,the formation of this geoherb is not clear in genetic mechanism,in which genome size is the first step that...Objective:Dianjixueteng is a geoherb in Yunnan Province,the source plant of which is Kadsura interior.However,the formation of this geoherb is not clear in genetic mechanism,in which genome size is the first step that should be known on the genomic level.In this study we aimed to estimate the genome sizes of source plants of K.interior and three related herbs K.heteroclita,K.longipedunculata,and K.coccinea by flow cytometry(FCM)and make a comparison.Methods:The genome sizes of K.interior,K.heteroclita,K.longipedunculata and K.coccinea,i.e.,the source plants of Dianjixueteng and its relative medicinal materials,were estimated by FCM.The nuclei of K.interior were isolated using modified LB01 buffer,for the rest species,by the Galbraith’s buffer.Results:The genome sizes of K.interior,K.heteroclita,K.longipedunculata,and K.coccinea were 7.36,7.12,7.01,and 5.15 pg/1 C,respectively.Genome size of K.interior had no significant variation with those of K.heteroclita and K.longipedunculata(P=0.296),which was significantly larger than that of K.coccinea.Conclusion:Genome size can not distinguish K.interior from K.heteroclita and K.longipedunculata,but could distinguish them from K.coccinea,which lays the foundation for future studies on genetic mechanism of the geoherb formation.展开更多
A laminar flow bioelectrochemical systems(BES)was designed and benchmarked using microbial anodes dominated with Geobacter spp.The reactor architecture was based on modeled flow fields,the resulting structure was 3D p...A laminar flow bioelectrochemical systems(BES)was designed and benchmarked using microbial anodes dominated with Geobacter spp.The reactor architecture was based on modeled flow fields,the resulting structure was 3D printed and used for BES manufacturing.Stratification of the substrate availability within the reactor channels led to heterogeneous biomass distribution,with the maximum biomass found mainly in the initial/middle channels.The anode performance was assessed for different hydraulic retention times while coulombic efficiencies of up to 100%(including also hydrogen recycling from the cathode)and current densities of up to 75 μA cm^(-2) at an anode surface to volume ratio of 1770 cm^(2) L^(-1) after 35 days were achieved.This low current density can be clearly attributed to the heterogeneous distributions of biomass and the stratification of the microbial community structure.Further,it was shown that time and space resolved analysis of the reactor microbiomes per channel is feasible using flow cytometry.展开更多
基金financial support of the National Natural Science Foundation of China(Grant Nos.61922079,61825107,and 62121003)the Chinese Academy of Sciences(Grant Nos.GJJSTD20210004 and Y201927)the National Key Research and Development Program of China(Grant No.2021YFC2500300).
文摘As the gold-standard method for single-cell analysis,flow cytometry enables high-throughput and multiple-parameter characterization of individual biological cells.This review highlights the demands for clinical flow cytometry in laboratory hematology(e.g.,diagnoses of minimal residual disease and various types of leukemia),summarizes state-of-the-art clinical flow cytometers(e.g.,FACSLyricTMby Becton Dickinson,DxFLEX by Beckman Coulter),then considers innovative technical improvements in flow cytometry(including quantitative,spectral,and imaging approaches)to address the limitations of clinical flow cytometry in hematology diagnosis.Finally,driven by these clinical demands,future developments in clinical flow cytometry are suggested.
文摘Objective:To investigate the effect of combined detection of serum carcinoembryonic antigen(CEA),cytokeratin 19 fragment(CYFRA21-1),cancer antigen 125(CA125),and neuron-specific enolase(NSE)in patients with lung cancer by fluorescence flow cytometry.Methods:From August 2019 to July 2022,200 patients with lung cancer diagnosed by pathology in our hospital were retrospectively analyzed.2 mL venous blood was collected in a fasting state and centrifuged to separate the serum(containing human chorionic gonadotropin antibody[anti-hCG antibody],hepatitis B surface antibody[anti-HBs antibody],and CEA).Results:The sensitivities of CEA and CYFRA21-1 detected via enzyme-linked immunosorbent assay(ELISA)were 100%,and the detection limits were 0.5 ng/mL and 0.1 ng/mL,respectively;the sensitivities of CA125 and NSE detected via flow cytometry were 100%,and the detection limits were 10 U/mL and 2 ng/mL,respectively.Compared with ELISA,the sensitivities of CA125 and NSE detected via flow cytometry were higher.When the concentration of CEA was 10-40 ng/mL,the sensitivities of the three markers CYFRA21-1,CA125,and NSE showed no significant changes(P>0.05);when the concentration of CEA was 40-80 ng/mL,the sensitivity of CEA significantly decreased(P<0.01),but the sensitivities of the three markers CYFRA21-1,CA125,and NSE showed no significant changes(P>0.05);when the concentration of CEA was 80-200 ng/mL,the sensitivities of all four markers showed no significant changes(P>0.05).Conclusion:Compared with the double-antibody sandwich ELISA,fluorescence flow cytometry has certain advantages,including high sensitivity,good precision,short detection time,low sample usage,and low medical cost;thus,it is worthy of clinical promotion.
文摘Objective:To explore the value of flow cytometry(FCM)in detecting the level of exfoliated cells in pleural effusion in the differential diagnosis of non-small cell lung cancer and benign lung diseases.Methods:Clinical data of patients with non-small cell lung cancer who were hospitalized in Hebei hospital from June 2019 to March 2022 were collected.A total of 98 patients were included,and 63 patients with alveolar lung disease were screened during the same period,and the two groups of patients were analyzed.Results:Compared with alveolar lung disease group,FCM detection and analysis showed that the level of exfoliated cells in the pleural effusion of non-small cell lung cancer(NSCLC)patients was 99(3-969)/100,000,and patients with alveolar lung disease was 4(0~19)/100,000.Additionally,compared with the alveolar lung disease group,the level of exfoliated cells in the pleural effusion of patients with non-small cell lung cancer(NSCLC)was significantly increased(P<0.001).The diagnostic efficacy of FCM for detecting pleural fluid exfoliated cells in non-small cell lung cancer was assessed using ROC curves and using 95%CI(-11.1,-13.2)with a sensitivity of 0.75 and specificity of 0.94,and the diagnostic efficacy of FCM for detecting pleural fluid exfoliated cells in alveolar lung disease was assessed using 95%CI(-11.1,-13.2)with a sensitivity of 0.71 and specificity of 0.87.Conclusion:Flow cytometry has a wider range of clinical applications,simple operation,low cost,and high sensitivity,which makes it of great significance in disease diagnosis.
基金supported by the National Natural Science Foundation of China (Nos.41176098 and 31372529)
文摘Flow cytometry(FCM) technique has been widely applied to estimating the genome size of various higher plants. However, there is few report about its application in algae. In this study, an optimized procedure of FCM was exploited to estimate the genome size of two eukaryotic algae. For analyzing Alexandrium catenella, an important red tide species, the whole cell instead of isolated nucleus was studied, and chicken erythrocytes were used as an internal reference. The genome size of A. catenella was estimated to be 56.48 ± 4.14 Gb(1C), approximately nineteen times larger than that of human genome. For analyzing Gracilariopsis lemaneiformis, an important economical red alga, the purified nucleus was employed, and Arabidopsis thaliana and Chondrus crispus were used as internal references, respectively. The genome size of Gp. lemaneiformis was 97.35 ± 2.58 Mb(1C) and 112.73 ± 14.00 Mb(1C), respectively, depending on the different internal references. The results of this research will promote the related studies on the genomics and evolution of these two species.
基金financial support from the University of North Florida.
文摘Gaining a full understanding of the mechanisms of action of natural products as therapeutic agents includes observing the effects of natural products on cellular morphology,because abnormal cellular morphology is an important aspect of cellular transformations that occur as part of disease states.In this study a set of natural products was examined in search of small molecules that influence the cylindrical morphology of fission yeast Schizosaccharomyces pombe.Imaging flow cytometry of large populations of S.pombe exposed to natural products captured cell images and revealed changes in mean length and aspect ratio of cells.Several natural products were found to alter S.pombe’s morphology relative to control,in terms of elongating cells,shrinking them,or making them more round.These results may facilitate future investigations into methods by which cells establish and maintain specific shapes.
基金the support from the Ministry of ScienceInnovation and Universities,Spain (AGL2017–88329-R and FPU18/00666)+1 种基金Regional Government of Catalonia,Spain (2017-SGR-1229)University of Girona (Postdoc-Ud G2020)。
文摘Background:Genetic selection in cattle has been directed to increase milk production.This,coupled to the fact that the vast majority of bovine artificial inseminations(AI)are performed using cryopreserved sperm,have led to a reduction of fertility rates over the years.Thus,seeking sensitive and specific sperm biomarkers able to predict fertility rates is of vital importance to improve cattle reproductive efficiency.In humans,sperm chromatin condensation evaluated through chromomycin A3(CMA3)has recently been purported to be a powerful biomarker for sperm functional status and male infertility.The objectives of the present study were:a)to set up a flow cytometry method for simultaneously evaluating chromatin condensation and sperm viability,and b)to test whether this parameter could be used as a predictor of in vivo fertility in bulls.The study included pools of three independent cryopreserved ejaculates per bull from 25 Holstein males.Reproductive outcomes of each sire were determined by non-return rates,which were used to classify bulls into two groups(highly fertile and subfertile).Results:Chromatin condensation status of bovine sperm was evaluated through the combination of CMA3 and Yo-Pro-1 staining and flow cytometry.Sperm quality parameters(morphology,viability,total and progressive motility)were also assessed.Pearson correlation coefficients and ROC curves were calculated to assess their capacity to predict in vivo fertility.Sperm morphology,viability and total motility presented an area under the ROC curve(AUC)of 0.54,0.64 and 0.68,respectively(P>0.05),and thus were not able to discriminate between fertile and subfertile individuals.Alternatively,while the percentage of progressively motile sperm showed a significant predictive value,with an AUC of 0.73(P=0.05),CMA3/Yo-Pro-1 staining even depicted superior results for the prediction of in vivo fertility in bulls.Specifically,the percentage of viable sperm with poor chromatin condensation showed better accuracy and precision to predict in vivo fertility,with an AUC of 0.78(P=0.02).Conclusions:Chromatin condensation evaluated through CMA3/Yo-Pro-1 and flow cytometry is defined here as a more powerful tool than conventional sperm parameters to predict bull in vivo fertility,with a potential ability to maximising the efficiency of dairy breeding industry.
基金Supported by The 2021 Undergraduate Innovation and Entrepreneurship Training Program of Guangdong Province(S202110564094)Horizontal Project of South China Agricultural University(h20220138).
文摘[Objectives]This study was conducted to establish a method for detecting the immunophenotypes of venous blood CIK cells in healthy donors and patients with triple-negative breast cancer or lung cancer by flow cytometry,and to further analyze and discuss the proportion of cellular immunophenotypes such as CD3^(+),CD4^(+),CD8^(+),CD3^(+)CD8^(+),CD3^(+)CD56^(+)in CIK cells.[Methods]Human venous blood was drawn,then anticoagulated with heparin and isolated with lymphocyte isolation solution,and the relevant immunophenotypes were detected by flow cytometry after 21 d of culture.[Results]The expression of CD3^(+)CD8^(+)and CD3^(+)CD56^(+)in the venous blood CIK cells was significantly higher in healthy donors than that in triple-negative breast and lung cancer patients.[Conclusions]CD3^(+)CD8^(+)and CD3^(+)CD56^(+)CIK cells have high anti-tumor activity.
文摘<strong>Objective:</strong> <span><span><span style="font-family:;" "=""><span style="font-family:Verdana;">To investigate the value of cell morphology and flow cytometry in the diagnosis of Multiple myeloma (MM). </span><b><span style="font-family:Verdana;">Methods:</span></b><span style="font-family:Verdana;"> 166 newly diagnosed MM patients were retrospectively analyzed, and the proportion of plasma cells detected by flow cytometry and cell morphology in these patients was statistically analyzed. </span><b><span style="font-family:Verdana;">Results:</span></b><span style="font-family:Verdana;"> All patients were divided into three groups according to the proportion of plasma cells detected by cell morphology: group A (≥30%), group B (≥10% and <30%), and group C (<10%). 1) In all patients and group A, B and C, the proportion of plasma cells detected by cell morphology was all correlated with that detected by flow cytometry. 2) The proportion of plasma cells detected by cell morphology in group A and group B was higher than that by flow cytometry, while there was no significant difference in group C. 3) The sensitivity of plasma cells detected by flow cytometry in group A, B and C was 88.7%, 51.2% and 10%, respectively. 4) According to the immunophenotypic analysis, the immunophenotypic characteristics were similar, all of them were CDl9 </span></span></span></span><span><span><span style="font-family:;" "=""><span style="font-family:Verdana;">- </span><span style="font-family:Verdana;">CD38 + CDl38 + CD56 +/</span><span style="font-family:Verdana;">- </span><span style="font-family:Verdana;">CD45dim ~ </span><span style="font-family:Verdana;">-</span><span><span style="font-family:Verdana;">. </span><b><span style="font-family:Verdana;">Conclusion:</span></b><span style="font-family:Verdana;"> There is a certain correlation between the two detection methods. The quantitative detection of myeloma cells by cell morphology is better, and the qualitative detection by flow cytometry is better. Combined with the two detection methods, the detection rate of multiple myeloma can be significantly improved.</span></span></span></span></span>
基金partially supported by the CONACYT(Mexico)grant 0105961/10110/194/09.
文摘Sperm capacitation involves functional changes, such as the removal or appearance of specific molecules and changes in the plasma membrane;the acrosome reaction (AR) is an exocytotic event induced by calcium influx, enabling the spermatozoa to penetrate the zona pellucida. These processes can be achieved only if the spermatozoa have good viability;indeed, determination of sperm viability is used for the assessment of semen quality. Membrane integrity and mitochondrial activity are important viability parameters of spermatozoa and fluorescent techniques based on membrane permeability to dyes have been developed to determine these parameters. The aim of this work was to determine the viability of boar sperm (fresh, one hour of capacitation induction and 20 min of AR induction) by flow cytometry using propidium iodide (PI) (1.25 μg/mL) and rhodamine 123 (R123) (0.20 μg/mL). Aliquots of 5 × 105 sperm were incubated with each fluorochrome separately and simultaneously for 10 or 20 min, respectively, at 38℃. The proportion of labeled spermatozoa and their fluorescence intensities were measured using a flow cytometer. The fluorescence index (FI) with PI gradually increased during the incubation and we found significant differences between all the groups. With R123, the FI increased in the capacitated sperm but decreased in the acrosome-reacted sperm, with significant differences between the fresh and capacitated spermatozoa. Our results suggest that the increase in the R123 fluorescence intensity in capacitated spermatozoa is due to changes in the mitochondrial membrane activity because the spermatozoa experienced changes in membrane fluidity and flagellar activation during capacitation. The use of fluorochromes and flow cytometry is a good tool for monitoring many markers of sperm function. Although capacitation and AR processes have been well studied, there is still much information to be elucidated with regard to these complex processes.
基金supported by the National Key Research and Development Program of China(2021YFA1301000)Shanghai Municipal Science and Technology Major Project(Grant No.2017SHZDZX01).
文摘Immunophenotyping is proving crucial to understanding the role of the immune system in health and disease.High-through-put flow cytometry has been used extensively to reveal changes in immune cell composition and function at the single-cell level.Here,we describe six optimized 11-color flow cytometry panels for deep immunophenotyping of human whole blood.A total of 51 surface antibodies,which are readily available and validated,were selected to identify the key immune cell populations and evaluate their functional state in a single assay.The gating strategies for effective flow cytometry data analysis are included in the protocol.To ensure data reproducibility,we provide detailed procedures in three parts,including(1)instrument characterization and detector gain optimization,(2)antibody titration and sample staining,and(3)data acquisition and quality checks.This standardized approach has been applied to a variety of donors for a better understanding of the complexity of the human immune system.
文摘Nowadays,doctors and nutritionists recommend individuals incorporate selenium-rich foods such as nuts,cereals,and mushrooms into their regular diet to maintain fitness and overall health.Selenium nanoparticles(SeNPs)exhibit strong chemopreventive capabilities.The anticipations for SeNPs with enhanced and tunable bioactive activities have led to a keen interest in phytofabrication.In this study,the aqueous extract of Clerodendron phlomidis plant leaves was utilized for the synthesis of SeNPs.In traditional Indian medicine,this plant extract is recognized as a significant anti-diabetic agent.The flavonoids tetrahydroxylflavone,7-hydroxyflavanone,and 6,4’-dimethyl-7-acetoxy-scutellarein present in this plant leaf extract demonstrate excellent anticancer activity.These secondary metabolites exhibit the ability to reduce sodium selenite into SeNPs.At a concentration of 13μg/mL,the synthesized SeNPs effectively inhibited the proliferation of the HepG2 cell line.The results suggest that the SeNPs possess promising anti-cancer potential against liver cancer and can be considered as a therapeutic agent for liver cancer treatment.Additionally,the cell cycle arrest induced by SeNPs was further confirmed by the fluorescence-activated cell sorting(FACS)method,indicating that SeNPs could efficiently differentiate cancer cells from normal cells.Notably,it showed a significant improvement in diethylnitrosamine(DEN)-induced Swiss Wistar rat groups.This scientific investigation highlights the high anti-cancer potential of SeNPs,positioning them as a promising therapeutic agent for liver cancer treatment.
基金by the NIH National Cancer Institute,R21CA122648,ONR N00014-07-1-0982,and the State of Florida Center of Excellence.M.-C.E.acknowledges fi nancial support from the Department d’Universitats,Recerca i Societat de la Informacióde la Generalitat de Catalunya,Spain.
文摘Early and accurate diagnosis and treatment of cancer depend on rapid,sensitive,and selective detection of tumor cells.Current diagnosis of cancers,especially leukemia,relies on histology and fl ow cytometry using single dye-labeled antibodies.However,this combination may not lead to high signal output,which can hinder detection,especially when the probes have relatively weak affi nities or when the receptor is expressed in a low concentration on the target cell surface.To solve these problems,we have developed a novel method for sensitive and rapid detection of cancer cells using dye-doped silica nanoparticles(NPs)which increases detection sensitivity in fl ow cytometry analyses between 10-and 100-fold compared to standard methods.Our NPs are~60 nm in size and can encapsulate thousands of individual dye molecules within their matrix.We have extensively investigated surface modifi cation strategies in order to make the NPs suitable for selective detection of cancer cells using fl ow cytometry.The NPs are functionalized with polyethylene glycol(PEG)to prevent nonspecifi c interactions and with neutravidin to allow universal binding with biotinylated molecules.By virtue of their reliable and selective detection of target cancer cells,these NPs have demonstrated their promising usefulness in conventional fl ow cytometry.Moreover,they have shown low background signal,high signal enhancement,and effi cient functionalization,either with antibody-or aptamer-targeting moieties.
基金supported by the Bilbao Bizkaia Water Consortium
文摘This study aims to demonstrate the validity of fluorescence-based methods,together with flow cytometry,as a complementary tool to conventional physicochemical analyses carried out in wastewater treatment plants(WWTPs),for the control of the currently largely unknown activated sludge process.Staining with SYTO 9,propidium iodide and 5-(and 6)-carboxy-2’,7’-difluorodihydrofluorescein diacetate(carboxy-H2 DFFDA)was used for cell viability and oxidative stress monitoring of the bacterial population forming the activated sludge of a WWTP.Throughout the period of research,several unstable periods were detected,where the non-viable bacteria exceeded the 75%of the total bacterial population in the activated sludge,but only in one case the cells with oxidative stress grew to 9%,exceeding the typical values of2%-5%of this plant.These periods coincided in two cases with high values of total suspended solids(SST)and chemical oxygen demand(COD)in the effluent,and with an excess of ammonia in other case.A correlation between flow cytometric and physicochemical data was found,which enabled to clarify the possible origin of each case of instability in the biological system.This experience supports the application of bacterial fluorescence staining,together with flow cytometric analysis,as a simple,rapid and reliable tool for the control and better understanding of the bacteria dynamics in a biological wastewater treatment process.
基金supported by the National Science and Technology Major Project of China (2017ZX10103011-003, 2018ZX10102001)the National Natural Science Foundation of China (31570163)Guangzhou Science and Technology Program key projects (201803040004)。
文摘The assessment of neutralization activity is an important step in the evaluation of neutralizing antibodies (NAbs).The traditional methods for measuring the antibody neutralization of human adenovirus type 3 (HAd V-3) are the microneutralization (MN) assay,which has insufficient sensitivity,and the plaque reduction neutralization test (PRNT),which is not suitable for high-throughput screening.Herein,we describe the development of a flow cytometry-based neutralization(FCN) assay for measuring the neutralization of sera,cell culture supernatants,and chimeric antibodies against HAd V-3 on the basis of a recombinant HAd V-3 (r HAd V-3) construct expressing the enhanced green fluorescent protein (EGFP).For flow cytometry-based assays,the optimal cell confluence was determined as 90%,and the virus was titrated using the assay.The established FCN assay follows the percentage law and an optimal MOI of not less than 5 9 10~(-4)was determined by using a purified chimeric antibody.In addition,comparison of the anti-HAd V-3 NAb titers of 72 human serum samples by the MN and FCN assays,showed that both assays correlated strongly with each other.Our FCN assay was an improvement over the MN assay because the observation period was reduced from 3 to 1 days and data analysis could be performed objectively and robotically.Importantly,the newly established FCN assay allows measurement of the neutralization activity of chimeric antibodies expressed in cell culture supernatants.Thus,this sensitive and highthroughput FCN assay is a useful alternative to the MN assay for measuring the antibody neutralization of HAd V-3 and for screening anti-HAd V-3 NAbs in cell culture supernatants.
文摘Soil microhabitats and their heterogeneity are often considered to be among the most important factors affecting soil biotic communities.The microbial commu-nity has become one of the most important links in soil nutrient cycles and trophic components due to its role in biological processes,spatial and temporal dynamics,and physiological adaptation.Sandy-soil desert systems are characterized by fast water infiltration during the rainy season,high salinity,and low moisture availability in the upper soil layers.Plants have developed different ecophy-siological adaptations in order to cope with this harsh environment.The Tamarix aphylla is known to be one of the most commonly adapted plants,exhibiting a mechan-ism for secretion of excess salts as aggregates through its leaves.These leaves aggregate beneath the plant,creating'islands of salinity'.Soil biotic components are,therefore,exposed to extreme abiotic stress conditions in this niche.The goal of this study was to examine the effect of T.aphylla on the live/dead bacterial population ratio on a spatial and temporal scale.The results emphasize the effect of abiotic factors,which changed on temporal as well as spatial scales,and also on the size of the active soil bacterial community,which fluctuated between 1.44%and 25.4%in summer and winter,respectively.The results of this study elucidate the importance of moisture availability and the'island-of-salinity'effect on the active microbial community in a sandy desert system.
基金financially supported by the National Natural Science Foundation of China(Grant No.81872965,81703650)。
文摘Objective:Dianjixueteng is a geoherb in Yunnan Province,the source plant of which is Kadsura interior.However,the formation of this geoherb is not clear in genetic mechanism,in which genome size is the first step that should be known on the genomic level.In this study we aimed to estimate the genome sizes of source plants of K.interior and three related herbs K.heteroclita,K.longipedunculata,and K.coccinea by flow cytometry(FCM)and make a comparison.Methods:The genome sizes of K.interior,K.heteroclita,K.longipedunculata and K.coccinea,i.e.,the source plants of Dianjixueteng and its relative medicinal materials,were estimated by FCM.The nuclei of K.interior were isolated using modified LB01 buffer,for the rest species,by the Galbraith’s buffer.Results:The genome sizes of K.interior,K.heteroclita,K.longipedunculata,and K.coccinea were 7.36,7.12,7.01,and 5.15 pg/1 C,respectively.Genome size of K.interior had no significant variation with those of K.heteroclita and K.longipedunculata(P=0.296),which was significantly larger than that of K.coccinea.Conclusion:Genome size can not distinguish K.interior from K.heteroclita and K.longipedunculata,but could distinguish them from K.coccinea,which lays the foundation for future studies on genetic mechanism of the geoherb formation.
基金financed by the German Federal Ministry of Education and Research(BMBF)under the ElektroPapier project(Grant nr:03XP0041G)supported by the Helmholtz-Association within the Research Programme Renewable Energies.
文摘A laminar flow bioelectrochemical systems(BES)was designed and benchmarked using microbial anodes dominated with Geobacter spp.The reactor architecture was based on modeled flow fields,the resulting structure was 3D printed and used for BES manufacturing.Stratification of the substrate availability within the reactor channels led to heterogeneous biomass distribution,with the maximum biomass found mainly in the initial/middle channels.The anode performance was assessed for different hydraulic retention times while coulombic efficiencies of up to 100%(including also hydrogen recycling from the cathode)and current densities of up to 75 μA cm^(-2) at an anode surface to volume ratio of 1770 cm^(2) L^(-1) after 35 days were achieved.This low current density can be clearly attributed to the heterogeneous distributions of biomass and the stratification of the microbial community structure.Further,it was shown that time and space resolved analysis of the reactor microbiomes per channel is feasible using flow cytometry.
