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Cancers ◽  
2022 ◽  
Vol 14 (2) ◽  
pp. 408
Author(s):  
Noemí Muñoz-García ◽  
F. Morán-Plata ◽  
Neus Villamor ◽  
Margarida Lima ◽  
Susana Barrena ◽  
...  

Flow cytometric (FCM) analysis of the constant region 1 of the T-cell receptor β chain (TRBC1) expression for assessing Tαβ-cell clonality has been recently validated. However, its utility for the diagnosis of clonality of T-large granular lymphocytic leukemia (T-LGLL) needs to be confirmed, since more mature Tαβ cells (i.e., T-LGL normal-counterpart) show broader TRBC1+/TRBC1− ratios vs. total Tαβ cells. We compared the distribution and absolute counts of TRBC1+ and TRBC1− Tαβ-LGL in blood containing polyclonal (n = 25) vs. clonal (n = 29) LGL. Overall, polyclonal TRBC1+ or TRBC1− Tαβ-LGL ranged between 0.36 and 571 cells/μL (3.2–91% TRBC1+ cells), whereas the clonal LGL cases showed between 51 and 11,678 cells/μL (<0.9% or >96% TRBC1+ cells). Among the distinct TCRVβ families, the CD28− effector-memory and terminal-effector polyclonal Tαβ cells ranged between 0 and 25 TRBC1+ or TRBC1− cells/μL and between 0 and 100% TRBC1+ cells, while clonal LGL ranged between 32 and 5515 TRBC1+ or TRBC1− cells/μL, representing <1.6% or >98% TRBC1+ cells. Our data support the utility of the TRBC1-FCM assay for detecting T-cell clonality in expansions of Tαβ-LGL suspected of T-LGLL based on altered percentages of TRBC1+ Tαβ cells. However, in the absence of lymphocytosis or in the case of TαβCD4-LGL expansion, the detection of increased absolute cell counts by the TRBC1-FCM assay for more accurately defined subpopulations of Tαβ-LGL-expressing individual TCRVβ families, allows the detection of T-cell clonality, even in the absence of phenotypic aberrations.


Cells ◽  
2022 ◽  
Vol 11 (2) ◽  
pp. 268
Author(s):  
Parivash Nouri ◽  
Anja Zimmer ◽  
Stefanie Brüggemann ◽  
Robin Friedrich ◽  
Ralf Kühn ◽  
...  

Advances in the regenerative stem cell field have propelled the generation of tissue-specific cells in the culture dish for subsequent transplantation, drug screening purposes, or the elucidation of disease mechanisms. One major obstacle is the heterogeneity of these cultures, in which the tissue-specific cells of interest usually represent only a fraction of all generated cells. Direct identification of the cells of interest and the ability to specifically isolate these cells in vitro is, thus, highly desirable for these applications. The type VI intermediate filament protein NESTIN is widely used as a marker for neural stem/progenitor cells (NSCs/NPCs) in the developing and adult central and peripheral nervous systems. Applying CRISPR-Cas9 technology, we have introduced a red fluorescent reporter (mScarlet) into the NESTIN (NES) locus of a human induced pluripotent stem cell (hiPSC) line. We describe the generation and characterization of NES-mScarlet reporter hiPSCs and demonstrate that this line is an accurate reporter of NSCs/NPCs during their directed differentiation into human midbrain dopaminergic (mDA) neurons. Furthermore, NES-mScarlet hiPSCs can be used for direct identification during live cell imaging and for flow cytometric analysis and sorting of red fluorescent NSCs/NPCs in this paradigm.


Author(s):  
Aakash Chandran Chidambaram ◽  
Kiruthiga Sugumar ◽  
Selvamanojkumar Sundaravel ◽  
Jaikumar Govindaswamy Ramamoorthy ◽  
Siddardha Bathula ◽  
...  

AbstractProlidase deficiency (PD) is a rare inborn error of metabolism causing ulcers and other skin disorders, splenomegaly, developmental delay, and recurrent infections. Most of the literature is constituted of isolated case reports. It occurs due to the mutations in the prolidase gene (PEPD) that result in loss of prolidase activity. We reported here a child who had presented with features compatible with hyper-immunoglobulin E syndrome (HIES) like recurrent skin ulcers, recurrent infections, facial dysmorphism, retained primary teeth, and elevated levels of immunoglobulin E levels but with normal flow cytometric assays, which was later diagnosed as PD.


Author(s):  
Fatima Redah Alassaif ◽  
Eman Redah Alassaif ◽  
Amit Kumar Kaushik ◽  
Jeevitha Dhanapal

Objective: The aim of the present article was to enhance the therapeutic efficacy of carboplatin (CP) using the formulation of chitosan – poly (lactic glycolic acid) nanoparticles (CS-PLGA NPs). Methods: Nanoparticles were synthesized by an ionic gelation method and were characterized for their morphology, particle size, and surface potential measurements by TEM and zeta sizer. This study was highlighted for the evaluation of drug entrapment, loading and in vitro drug release capabilities of the prepared nanoparticles by spectrophotometric analysis. The stability study was also conducted after 3 months for their particle size, zeta potential, drug loading and encapsulation efficiencies. Further, ovarian cancer cell line PEO1 were used to evaluate the toxicity and efficacy of nano-formulation by MTT assay. Further, the study was evaluated for apoptosis using flow cytometric analysis. Result: The CS-PLGA-CP NPs were uniform and spherical in shape. The particle size and zeta potential of CS-PLGA-CP NPs were measured 156 ± 6.8 nm and +52 ± 2.4 mV, respectively. High encapsulation (87.4 ± 4.5 %) and controlled retention capacities confirmed the efficiency of the prepared nanoparticles in a time and dose dependant manner. The cytotoxicity assay results also showed that CS-PLGA-CP NPs has high efficiency on PEO1 cells compared to the free drug. The flow cytometric result showed 64.25 % of the PEO1 cells were apoptotic and 8.42 % were necrotic when treated with CS-PLGA-CP NPs. Conclusion: Chitosan-PLGA combinational polymeric nanoparticles were not only steady but also non-toxic. Our experiments revealed that the chitosan- PLGA nanoparticles could be used as a challenging vehicle candidate for drug delivery for the therapeutic treatment of ovarian cancer.


2022 ◽  
Vol 12 ◽  
Author(s):  
Bruce K. Patterson ◽  
Edgar B. Francisco ◽  
Ram Yogendra ◽  
Emily Long ◽  
Amruta Pise ◽  
...  

The recent COVID-19 pandemic is a treatment challenge in the acute infection stage but the recognition of chronic COVID-19 symptoms termed post-acute sequelae SARS-CoV-2 infection (PASC) may affect up to 30% of all infected individuals. The underlying mechanism and source of this distinct immunologic condition three months or more after initial infection remains elusive. Here, we investigated the presence of SARS-CoV-2 S1 protein in 46 individuals. We analyzed T-cell, B-cell, and monocytic subsets in both severe COVID-19 patients and in patients with post-acute sequelae of COVID-19 (PASC). The levels of both intermediate (CD14+, CD16+) and non-classical monocyte (CD14Lo, CD16+) were significantly elevated in PASC patients up to 15 months post-acute infection compared to healthy controls (P=0.002 and P=0.01, respectively). A statistically significant number of non-classical monocytes contained SARS-CoV-2 S1 protein in both severe (P=0.004) and PASC patients (P=0.02) out to 15 months post-infection. Non-classical monocytes were sorted from PASC patients using flow cytometric sorting and the SARS-CoV-2 S1 protein was confirmed by mass spectrometry. Cells from 4 out of 11 severe COVID-19 patients and 1 out of 26 PASC patients contained ddPCR+ peripheral blood mononuclear cells, however, only fragmented SARS-CoV-2 RNA was found in PASC patients. No full length sequences were identified, and no sequences that could account for the observed S1 protein were identified in any patient. That non-classical monocytes may be a source of inflammation in PASC warrants further study.


2022 ◽  
pp. 107815522110738
Author(s):  
Burcu Aslan Candır ◽  
Tuğçe Nur Yiğenoğlu ◽  
Merih Kızıl Çakar ◽  
Mehmet Sinan Dal ◽  
Fevzi Altuntaş

Introduction The most common kind of leukemia in adults is chronic lymphocytic leukemia (CLL). CLL is treated with ibrutinib. During the course of ibrutinib therapy, bleeding and cardiac arrhythmias may occur. Non-hemorrhagic adverse events are extremely infrequent in individuals using ibrutinib. Case report A 64 year-old man was diagnosed with CLL in June 2016. He was treated with 6 courses of FCR, he stayed in remission for 3 years and then relapsed. He achieved partial remission after two months of therapy with ibrutinib. The patient was admitted to the hospital with fever and shortness of breath. Pericardial tamponade and effusion was diagnosed during his evaluation. Management & outcome Non-hemorrhagic exudative effusion was drained by pericardiocentesis and a pericardial catheter was inserted to drain pericardial effusion. In all pleural and pericardial effusion samples, pathological and flow cytometric examination revealed no atypical malignant cells for malignancy, including CLL. Infections, both bacterial and viral, were also undetectable in the samples, as were rheumatological markers of collagen vascular disease. Ibrutinib therapy was discontinued. The pericardial effusion and tamponade were linked to ibrutinib treatment after evaluating the adverse drug reaction probability scale with a total score of 6. Colchicine was administered to reduce the pericardial effusion. The catheter was removed; pericardial effusion did not reoccur during follow up visits. Discussion Serious adverse events of ibrutinib are seen when treating CLL patients. This group of individuals should be closely monitored for potentially serious complications such as pericardial effusion and cardiac tamponade.


2022 ◽  
Vol 308 (1) ◽  
Author(s):  
Josef Greimler ◽  
Eva M. Temsch ◽  
Zhiqing Xue ◽  
Hanna Weiss-Schneeweiss ◽  
Polina Volkova ◽  
...  

AbstractThe grass Deschampsia cespitosa is a variable taxon out of which many varieties, subspecies and endemic species have been separated. In this paper, the variation in genome size (GS) and ploidy of this grass including several of its subspecies and two related species in Eurasia was investigated by flow cytometric (FCM) measurements. GS and ploidy data were also related to specific environments and reproduction mode. Ploidy levels found by FCM were confirmed by chromosome counts of diploid (2n = 28) and tetraploid (2n = 52) samples. Seminiferous (seed bearing) D. cespitosa was mainly diploid (GS between 3.754 and 5.438 pg/1C). GS variation in diploids showed a geographic pattern with a significant difference (H = 41,441, P < 0.001) between European (median = 4.377 pg) and Asian (median = 4.881 pg) accessions. Genome size (1C) in tetraploids ranged from 7.9426 to 9.0399 pg. Tetraploid seminiferous D. cespitosa was found mostly in disturbed habitats in western and southern Europe, while tetraploids in Asia were registered in wet Arctic habitats. Genome size (1C between 8.3278 and 8.8603 pg) of the pseudoviviparous plants (spikelets produce plantlets asexually) of wet habitats in central and northern Europe indicated tetraploidy. A putative triploid (GS 6.6817 pg) was detected in Iceland. Summing up, we found a high variation in GS on the geographic scale with significant regional differences in diploid D. cespitosa. Among the tetraploids, the asexually reproducing plants were bound to specific habitats, while the seminiferous plants showed a habitat preference similar to the diploids.


Blood ◽  
2022 ◽  
Author(s):  
Gabrielle Paras ◽  
Linde M. Morsink ◽  
Megan Othus ◽  
Filippo Milano ◽  
Brenda M. Sandmaier ◽  
...  

In acute myeloid leukemia (AML), measurable residual disease (MRD) before or after allogeneic hematopoietic cell transplantation (HCT) is an established, independent indicator of poor outcome. To address how peri-HCT MRD dynamics could refine risk assessment across different conditioning intensities, we analyzed 810 adults transplanted in remission after myeloablative conditioning (MAC; n=515) or non-MAC (n=295) who underwent multiparameter flow cytometry-based MRD testing before and 20-40 days after allografting. Patients without pre- and post-HCT MRD (MRDneg/MRDneg) had the lowest risks of relapse and highest relapse-free survival (RFS) and overall survival (OS). Relative to those patients, outcomes for MRDpos/MRDpos and MRDneg/MRDpos patients were poor regardless of conditioning intensity. Outcomes for MRDpos/MRDneg patients were intermediate. Among 161 patients with MRD before HCT, MRD was cleared more commonly with a MAC (85/104 [81.7%]) than non-MAC (33/57 [57.9%]) regimen (P=0.002). Although non-MAC regimens were less likely to clear MRD, if they did the impact on outcome was greater. Thus, there was a significant interaction between conditioning intensity and "MRD conversion" for relapse (P=0.020), RFS (P=0.002), and OS (P=0.001). Similar findings were obtained in the subset of 590 patients receiving HLA-matched allografts. C-statistic values were higher (indicating higher predictive accuracy) for peri-HCT MRD dynamics compared to the isolated use of pre-HCT MRD status and post-HCT MRD status for prediction of relapse, RFS, and OS. Across conditioning intensities, peri-HCT MRD dynamics improve risk assessment over isolated pre- or post-HCT MRD assessments.


2022 ◽  
Vol 15 (1) ◽  
pp. 57
Author(s):  
Katalin Szabó ◽  
Ágnes Kemény ◽  
Noémi Balázs ◽  
Esam Khanfar ◽  
Zoltán Sándor ◽  
...  

Transient Receptor Potential Ankyrin 1 (TRPA1) has been reported to influence neuroinflammation and lymphocyte function. We analysed the immune phenotype and activation characteristics of TRPA1-deficient mice (knockout—KO) generated by targeted deletion of the pore-loop domain of the ion channel. We compared TRPA1 mRNA and protein expression in monocyte and lymphocyte subpopulations isolated from primary and secondary lymphatic organs of wild type (WT) and KO mice. qRT-PCR and flow cytometric studies indicated a higher level of TRPA1 in monocytes than in lymphocytes, but both were orders of magnitude lower than in sensory neurons. We found lower CD4+/CD8+ thymocyte ratios, diminished CD4/CD8 rates, and B cell numbers in the KO mice. Early activation marker CD69 was lower in CD4+ T cells of KO, while the level of CD8+/CD25+ cells was higher. In vitro TcR-mediated activation did not result in significant differences in CD69 level between WT and KO splenocytes, but lower cytokine (IL-1β, IL-6, TNF-α, IL-17A, IL-22, and RANTES) secretion was observed in KO splenocytes. Basal intracellular Ca2+ level and TcR-induced Ca2+ signal in T lymphocytes did not differ significantly, but interestingly, imiquimod-induced Ca2+ level in KO thymocytes was higher. Our results support the role of TRPA1 in the regulation of activation, cytokine production, and T and B lymphocytes composition in mice.


2021 ◽  
Vol 15 (12) ◽  
pp. 531-539
Author(s):  
Kandaiah Vidhyaini ◽  
Singaram Nallammai ◽  
Isparan Kandasamy Kodi

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