httpwwwglassbottomdishescom Medium aging where fluorescent molecules are Information regarding protein mobilitymeasuring the rate of fluorescenc Why Does Fluorescence Recover PrebleachPartia ID: 831932
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Why Live Imaging?1.Confocal and Multi-ph
Why Live Imaging?1.Confocal and Multi-photon Fluorescence Mi
croscopy 2.Green Fluorescent Protein (GF
croscopy 2.Green Fluorescent Protein (GFPs)http://www.glass
-bottom-dishes.com/Mediumaging where f
-bottom-dishes.com/Mediumaging where fluorescent molecules
are Information regarding protein mobil
are Information regarding protein mobilitymeasuring the rat
e of fluorescencWhy Does Fluorescence R
e of fluorescencWhy Does Fluorescence Recover?Pre-bleachPa
rtial recoveryFull recovery(Pure Diffus
rtial recoveryFull recovery(Pure Diffusion FRAP)(first pos
tbleachintensity)) / 2)halfZ2/(4 ât1/
tbleachintensity)) / 2)halfZ2/(4 ât1/2 )where Zis the rad
ius of bleached areaMore accurate estima
ius of bleached areaMore accurate estimates of D can be FRA
P Experiments at DBCChromatin structur
P Experiments at DBCChromatin structure Transcription mR
NA mobility Protein recycling Signal t
NA mobility Protein recycling Signal transduction Cytoske
letaldynamics Vesicle transport Cell a
letaldynamics Vesicle transport Cell adhesion Mitosisf~
50 kDaGFP-histoneH2BMolecular massto cro
50 kDaGFP-histoneH2BMolecular massto cross the nucleusDiffu
sion of proteins in the nucleusConclusio
sion of proteins in the nucleusConclusionGFP-HMG-17,GFP-SF2
/ASF and GFP-fibrillarin) mobility in th
/ASF and GFP-fibrillarin) mobility in the nucleoplasmof livi
ng cells. The lower mobility of the fus
ng cells. The lower mobility of the fusion with the Splici
ng Factor Compartments (SFC)1.Associati
ng Factor Compartments (SFC)1.Association of GFP-SF2/AS2.Im
morbilefraction ~ 10% indicating virtual
morbilefraction ~ 10% indicating virtuall3.Inhibition of tra
nscription by -amanitinaccelerates the r
nscription by -amanitinaccelerates the reassociationof GFP-t
he absence of nucleoplasmicbinding Quan
he absence of nucleoplasmicbinding Quantitative analysis of
binding reactions by FRAPa. Diffusion-u
binding reactions by FRAPa. Diffusion-uncoupled FRAP recove
riesThe diffusion time is much shorter t
riesThe diffusion time is much shorter than the time to begi
n binding. Consequently, recovery due t
n binding. Consequently, recovery due to diffusion occurs f
irst and recovery due to binding follows
irst and recovery due to binding follows later, resulting in
a FRAP curve that can be separated into
a FRAP curve that can be separated into two phases. Fluor
escent moleculeBleached moleculeBindin
escent moleculeBleached moleculeBinding sitermixed throug
hout the recovery, the FRAP Distinguish
hout the recovery, the FRAP Distinguishing diffusion-couple
d from diffusion-uncoupled FRAPDifferent
d from diffusion-uncoupled FRAPDifferent behavior arise depe
nding on whether the FRAP recovery is di
nding on whether the FRAP recovery is diffusion coupled or u
ncoupled, therefore, it is esstwo situat
ncoupled, therefore, it is esstwo situations for correctly e
xtracting binding information from FRAP
xtracting binding information from FRAP data.ent size and c
ompare recoDiffusion coupledDiffusion
ompare recoDiffusion coupledDiffusion uncoupledDoing FRAP
with ZeissLSM510 Meta SystemClick to s
with ZeissLSM510 Meta SystemClick to start to define the b
leach areaDoing FRAP with ZeissLSM510 M
leach areaDoing FRAP with ZeissLSM510 Meta SystemChoose th
e shape of Locate the bleaching area on
e shape of Locate the bleaching area on the pre-acquired ima