ME vs MO · 2014-02-10 · 2/10/2014 1 Microscopia - A Ciência “rente ao olhar”- Rui Malhó...
Transcript of ME vs MO · 2014-02-10 · 2/10/2014 1 Microscopia - A Ciência “rente ao olhar”- Rui Malhó...
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Microscopia
- A Ciência “rente ao olhar” -
Rui Malhó Dept. Biologia VegetalUniv de Lisboa
Técnicas de Microscopia:
- Microscopia óptica- campo claro- campo escuro- contraste de fase- contraste de fase- DIC
- Microscopia de Fluorescência- Vídeo Digital- Confocal- Convencional
- Microscopia Electrónicap- Transmissão- Varrimento
- Microscopia de Força Atómica- Microsondas
- Microsondas vibráteis- Patch-clamp
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Óptica
Qualidade daQualidade daimagem finalimagem final
Arquitectura do sistema
Detectores
Resoluçãodo sistema
Arquitectura do sistema
MS
SCC
DIC-O
BF-ISFLUORESCENCEMICROSCOPE
FRAME GRABBER+
A/D CONVERTERFILTERCHANGE
MICROCOMPUTER
DIGITAL IMAGING SYSTEMPMT
SourcePinhole
DetectorPinhole
EM-FC
EX-FC
EX-IS
OBJ
DM
DM
F
MS
> 700 nm
DIGITAL IMAGEPROCESSING
(e.g. CDrom)
DATAARCHIVING
High resolution
monitor
HARD COPYDEVICE
CHANGECONTROL
(FRONTVIEW)
P
GRAPHICBOARD
Laser DM
OBJ
MSFocus
CC
DC
AM
ER
A
FLUORESCENCEIMAGING
VS
monitor
FLUORESCENCEEMISSION
EXCITATION
(reflected)In-focus light rays
Out-of-focus light rays
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Câmaras Digitais 8-16 bit
CCDs
Câmaras 35 mm
CCDs (cool)
Luminómetros
Sistema confocal
Fotomultiplicadores
MET vs MEV
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NpCDPK7 localises to the generative cell membrane
52 289 555 586 732 766a.aNVsmGFPKDCV (766 a.a)
NV Kinase domain JD CaM like domain CVsmGFP
52 289 319 555 586 732 766a.a
NVsmGFPKDK82MCV (766 a.a)
NV Kinase domain JD CaM like domain CVsmGFPK82M
Capture the tissue / cell suspension on the membrane filter
Transfer the cells to solid culture media on the slide glass
Transform by biolistic gun
GFP fluorescence iimage
Bright field image
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Equação Difusão (onda de cálcio):c / t = D 2C / x2em que: D (coeficiente de difusão); t (tempo - amplificação);c (concentração do mensageiro); x (distância)
GloFish
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FRET & FRAPbRET & FRAP
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Reconstrução 3D - um exemplo com biofilmes
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Lily pollen Olho de aranhaPenicillum (Brie)
Neurónio Estoma Espermatozoide Corte de célula