Ppt 7 12-2015
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The document describes the construction of an annexin V-EGFP fusion protein to use as a probe for detecting phosphatidylserine exposure on apoptotic cell membranes. Flow cytometry analysis was used to quantitatively analyze the interaction between annexin V and membranes under varying calcium and protein concentrations. Comparison of annexin V-EGFP to annexin V-FITC demonstrated that annexin V-EGFP is a suitable probe for labeling externalized phosphatidylserine during apoptosis.
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The Milky Way’s (MW) inner stellar halo contains an [Fe/H]-rich component with highly eccentric orbits, often referred to as the
‘last major merger.’ Hypotheses for the origin of this component include Gaia-Sausage/Enceladus (GSE), where the progenitor
collided with the MW proto-disc 8–11 Gyr ago, and the Virgo Radial Merger (VRM), where the progenitor collided with the
MW disc within the last 3 Gyr. These two scenarios make different predictions about observable structure in local phase space,
because the morphology of debris depends on how long it has had to phase mix. The recently identified phase-space folds in Gaia
DR3 have positive caustic velocities, making them fundamentally different than the phase-mixed chevrons found in simulations
at late times. Roughly 20 per cent of the stars in the prograde local stellar halo are associated with the observed caustics. Based
on a simple phase-mixing model, the observed number of caustics are consistent with a merger that occurred 1–2 Gyr ago.
We also compare the observed phase-space distribution to FIRE-2 Latte simulations of GSE-like mergers, using a quantitative
measurement of phase mixing (2D causticality). The observed local phase-space distribution best matches the simulated data
1–2 Gyr after collision, and certainly not later than 3 Gyr. This is further evidence that the progenitor of the ‘last major merger’
did not collide with the MW proto-disc at early times, as is thought for the GSE, but instead collided with the MW disc within
the last few Gyr, consistent with the body of work surrounding the VRM.
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Ppt 7 12-2015
- 1. Quantitative analysis of annexin V–membrane interaction by flow cytometry Presented by Muhammad Younis
- 2. Background • Annexin A5 (or annexin V) is a cellular protein in the annexin group. • AnxV can be used as an apoptosis marker. • In the presence of calcium, annexin V (anxV) specifically binds to PS with high affinity, which can be used to detect PS externalization occurring on the membrane of apoptotic cells. • To Distinguish early apoptotic cells from viable cells as well as late apoptotic and necrotic cells through flow cytometry.
- 4. Expression, purification, and characterization of anxV-EGFP
- 5. Preliminary characterization of anxV-EGFP protein.
- 6. Dotplot of R1 region of apoptotic cells
- 7. Calcium dependence of anxV-EGFP binding to apoptotic cell membrane P: anxV-EGFP concentration , PBI: Protein binding capacity index,
- 8. Comparison between experimental and theoretical curves
- 9. Verification and further analyses of the mathematical expression.
- 10. Evaluation of the expression with anxV- FITC Comparison between experimental curve and theoretical curve estimated under various concentrations of calcium and protein.
- 11. Summary • Constructed a green fluorescent phosphatidylserine (PS)-binding probe, • With this probe,investigated anxV–membrane interaction under different calcium and anxV-EGFP concentrations through flow cytometry (FCM). • Comparison between the performance of anxV-EGFP and anxV-FITC provided further evidence in support of anxV-EGFP being an eligible and convenient probe for labeling externalized PS during apoptosis process.