Abstract
Introduction The validation of patient derived cell lines and other biologics has gained importance due to more frequent laboratory collaborations. Polychromatic flow cytometry has been the ...gold standard for detection and analysis of biochemical, structural complexity and characterization of cellular particles in biological systems. Despite the tremendous advancement in flow cytometry from the earlier simple, slow version of 1960s to more recent complex, fast spectral analyzers with comprehensive structural and functional immune profiling capability, this development has not matched the ever-increasing need for high throughput output to phenotype cells. Mass cytometry (CyToF) uniquely combines time-of-flight mass spectrometry with metal-labeling technology which can theoretically but simultaneously analyze 50+ parameters on a single cell. Despite the functional similarities in utility, mass cytometers and flow cytometers have different operating platforms that may impact their functional readouts . The goal of this study was to investigate and compare phenotypic readouts between flow cytometry and Cytof platforms using human neuroblastoma cell lines.
Methods To test this question, we utilized custom designed staining panels for both flow cytometry (fluorochromes-PerCp Cy 5.5, PE and Coralite 488 respectively) and Cytof (lanthanides: 155Gd, 151Eu, and 116Cd respectively) targeting CD56, Nestin and Synaptophysin, markers that are consistent with neuroblastoma using both five established neuroblastoma cell lines (BE2C, CHLA90, SMS-KCNR, SHSY5Y) and NGP) as well as four novel cells lines established in our laboratory derived from patient specimens (SL01277, SL01404, SL01255 and SL01287). We compared the percent expression of respective markers across cell lines between the two platforms.
Results Using both techniques we demonstrated that there was no difference in detection of viability in cells when validated under flow cytometry or CyToF (p<0.05). We also show extracellular staining of CD56 between the two platforms are comparable(p<0.05) across tested cell lines. Finally, we demonstrate that intracellular structures can be detected using both platforms with no significant difference (p<0.05).
Discussion and Conclusion The congruence and reproducibility in readouts between flow cytometry and CyToF analysis indicates that either assay can be used to study biological systems. Given the high dimensionality and versatility, then CyToF offers a robust platform that can be leveraged for immunophenotypic and functional studies of cellular material. Furthermore, CyToF due to its wide breadth of multiparametric measurements available will be an instrument of choice where the study of multiple parameters is desired.
Citation Format: David Mulama, Roman J. Riveria, Kimberly McKinney, Divya Gandra, Kaitlyn Smith, Nicholas Tastet, Giselle L. Saulnier Sholler. Comparative validation of neuroblastoma cell lines using flow cytometry and CyToF abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3053.
Abstract
Background
The development of myocardial fibrosis is a key mechanism in the transition from compensated hypertrophy to heart failure in aortic stenosis (AS). Focal and diffuse fibrosis can ...be quantified using cardiac magnetic resonance (CMR) imaging late gadolinium-enhanced (LGE) and T1 mapping techniques.
Purpose
To assess T1 mapping measures of fibrosis in patients with severe AS referred for aortic valve intervention, and determine their associations with clinical characteristics, disease severity and long-term clinical outcome.
Methods
In this international prospective cohort study, patients with severe AS underwent contrast enhanced CMR with T1 mapping and LGE prior to aortic valve intervention. Image analysis was performed by a single core laboratory and the extracellular volume fraction ECV% calculated from T1 mapping images. The presence of LGE was determined visually and quantified using the full-width-at-half-maximum technique.
Results
Four-hundred and forty patients (70±10 years, 59% male) from ten international centres were enrolled. Aortic valve intervention was performed 15 4 to 58 days following CMR. Within a follow-up of 3.8 2.8 to 4.6 years, 52 patients died.
ECV% (mean 27.7±3.6%) correlated with increasing age, Society of Thoracic Surgeons Predicted Risk of Mortality score, known coronary artery disease, lower peak aortic-jet velocity, larger left ventricular (LV) mass, lower LV ejection fraction, and presence of LGE (P<0.05 for all). Following adjustment for all demographic and clinical variables, ECV% remained associated with age (P=0.028), LV ejection fraction (P<0.001) and presence of LGE (P=0.035).
Univariable predictors of all-cause mortality included age, male sex, impaired LV ejection fraction and presence of LGE (all P<0.05). A progressive increase in all-cause mortality was seen across tertiles of ECV% (17.3, 31.6 and 52.7 deaths per 1000 patient-years; log-rank test, P=0.009). ECV% was independently associated with all-cause mortality following adjustment for age, sex, impaired LV ejection fraction and presence of LGE (HR per unit increase in ECV: 1.10, 95%, (1.02–1.19), P=0.013).
ECV440 abstract iamge
Conclusion
In patients with severe aortic stenosis scheduled for aortic valve intervention, extracellular volume-based T1 mapping correlates with LV decompensation. ECV% is a strong independent predictor of late all-cause mortality and is a potential therapeutic target.
Holocene forested coastal dunes of different morphology fringe the Atlantic coast of southwest Aquitaine. Infra-red stimulated luminescence (IRSL) dating has been applied to sands from these dunes in ...the Aquitaine region in order to test the validity of dune-classification theories. The ages obtained from the dunes show three phases of sand invasion and dune development during the late Holocene: 3000-4000 years ago; 900-1300 years ago; 250-550 years ago. The timing of the most recent phase of sand mobilization. as dated by IRSL, is supported by historical maps and records from the fifteenth to the eighteenth centuries AD, showing problems for human settlement in the region and village abandonment due to dune drift. Sand invasion is driven by an increase in frequency of severe storms in the North Atlantic associated with the cooler periods of the ‘Little Ice Age’ and early 'Mediaeval Warm Period'. The dunes emplaced 900-1300 years ago were naturally fixed by a mixed deciduous and maritime pine forest during the latter part of the 'Mediaeval Warm Period'.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, OILJ, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
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