????? ???????????????? Electron Microscopy technique - PowerPoint PPT Presentation

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????? ???????????????? Electron Microscopy technique

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Electron Microscopy technique / - ... – PowerPoint PPT presentation

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Title: ????? ???????????????? Electron Microscopy technique


1
????? ???????????????? Electron
Microscopy technique
  • ????? / ????? ??????
  • ??????? ???????- ????? ?????? ????????? ??????

2
??? ?? ????? ??? ??? ??? ????? ??? ??????? ???
??? ?????? ???? ??? ?????? ??????????
  • ?????? ?????? ?????? ????? ??? ????? ???? High
    vacuum ??? ??? ?? ??????? ?????? ??? ??? ???? ??
    ????? ??? ?? ???? ???? ???? ?????????? .
  • ????? ???? ?????? ??? ?????? ??????? ??? ??????
    ???????????? ???? ???? ??????? .

3
1924
???? ??? ??? ??????? ???????? ??????????? ???
???? ????? ???? ???????? ??????? ??????? ???????.
???? ??? ?????? ???????
????? ????? ????? ???? ???? ?? ???? ?? ???? ???
????? ?? ????? ???? ????? ???? ?? ???? ????????
?? ????? ???? ?????? ?????? ??? ????? ??????
?????? ????? ?? ( - ) ???? ?????? ??? ??????
????? ????? ?????? ????? ?????? ??? ????? ??????
???? ?????? ????? ( / - ) ?? ?????? ????? ?????
????? ????? ???? ??? ????? ??????? ??? ???? ??
??? ???????? ?????? ??? ????? ??? ???? ?????
??????? ??????? ?????? ?? ???????? ?????? ???
???? ???????? ??? ????? ??????? ?? ????? ???? ??
???? ????
???? ?? ??? ?????? (???? ?? ??????) ????
???????????? ???? ??????? ???????
4
1955
  • - ??? ???? ??? ?????? ????? ????? ???? ?????
    ??????? ?? ????? ??? ?????? ?? ????? ????????
    ??????? ???? ?????? ????? ?? ????? ?????? ???????
  • ??????? ?????? ???????? ????????? ??????????
    ?????? ?????? ????? ?????? ?????? ?????? ????????
    ?????? ???????????? ?????? ?? ??? ?????? ????????
    ??? ???? ???? ?? ??????? ?????? ??? ???????
    ??????? ??????????? ???????? E.M. ????? ???? ??
    ???????? ????????????? ??????? ??????? ? ???????
    ???? ?? ?? ??? ???
  • ???? ???? ???????? ?? ????? ??? ????????? ??????
    ?????? ?????? ????? ???????? ??? ????? ????????
    ??? ?????? ?????????? ??????
  • ???? ?? ??? ????????? ????? ??????? ??????

5
1980
??? ???? ?????? ?????????? ??????
1986
???? ???? ???????? ???? (SEM) ?????? ??? (35JSM)
6
????? ??????? ? ???????
L.M ?????
E.M ?????????
7
L.M?????
E.M?????????
1-Transmission Electron Microscopy
Compound L.M
Simple L.M
(TEM)
1- Inverted L.M
Single lenses
2- Scanning Elect Microseopy
(SEM)
2- Dark field L.M
3- Phase Contrast Microscop
3- Enery Dispersiv X-ray Spectroscop
4- U.V.M.
5- Immuno flourecent I.F.L.M.
4- Field Emission Scanning Electron
Microscopy
(F E.SEM)
6- Descetting L.M attaching
5- Electron Probe Micro- Analyzer.
7- Camera L.M
(EPMA) (SEMEDS)
8- Cooliscop L.M
8
?????? ??????
  • ??? ????? ??? ????? ????? ?????? ?????????? ???
    ?????? ????
  • ???? ?????? ?????? ?? ??????? ?????????? ???
    ????? ???????
  • ???????? ????????.
  • ??????? ???? ??????? ?????? ?? ?? ??? ???????
    ?????????
  • ????? ??????.

9
??????????? ??? ??? ??????? ???????? ??? ????????
  • ??? ??????? ??? ?????? ?????? ?????? ???
  • 1500x ?????? ?????? ???? ?????? , 25x ??????
    ?????? ?? ?????? ????????
  • ??? Magnification power (M.P.)
  • ?????? ???? ??? ??????? ???????
  • ???? ???? ???? ??? ??????? ??? ?????? ??? ??? ???
    ?? ???? ?????? ????? ????
  • ?????? ????????? ???? ?? ???????? ??????? ?????
    ??? ???????? ? ?????
  • ?????? ??? ??????? ??? ??? ??????? ? ???????
  • ???? ???? ?? ???? ???? ??????? ?? ??? ???????
    Resolution power (R.P.)

10
??? ?? ??? ??????? ??? ?????? ?????????? ??????
???? ?? ??? ????? ??? 600,000 ??? ?????? ?? ????
??????? ??? ?? ????? ??? ??? ?? ??? ?? ????? ???
??? ????? ???? ??????? ??????? ???? ???? ?? ????
???? ??????? ??? ????? ?? ??? ??????? ?? ??????
?????? ?????? ??????? ??? ??? ??? ?? ?? ????
?? ??? ??????? ?? ???? ??????? ???????- ? ???
??????? ?? ??? ??????? (Resolution power) ??
(Resolving power) ?? ???? ????? ??? ???? ?????
???????? ???? ?? ?????? ????? ??? ??????? ??????
?? ?????? ?? ??? ?? ?????
11
??? ??? ??? ?????? ??????????....?
  • ??? ??? ?????? ?????? ?????????? ????? ????? ??
    ??? ?????? ??? ?????
  • ??????? ??????????? ??? ??????? ?? ??? ???????
    ???????? ???????? ????? ???
  • ??????? ?????? ???? ????? ??? (R.P.) ??? ?????
    ?????? ?????????? ????
  • ????? ?????? ???? ( ?????? ) ?????? ??? ????
    ????? ???? ????? ??? ??
  • ????? ??? ????? ?? ??? ??????? ???? ???????
    ??????????

12
?????? ?? ???????? ?????????
  • 1m 1000mm
  • 1mm 1000um
    0,1mm 100um
  • 1nm 10 um 1um 1000nm
    0,1um 100 nm
  • 1nm 1000A

13
???????????? ???????? ??? ???????? ?? ?????
???????
  • ??? ??????? ????? ??????? ????? ??? ??? ??????? ?
    ???????

???? ??? ??????? ?? ??????? ??????? ????? ???
??????? ???????(????? ?????? ??????)
- ????? ??? ??????? ?? ??????? ??????????? ?????
??? ????? ????????? ( ??? ?????? ) ??? ??????
?? ?????? ????? ???? ?? ???? ??????? ????? ???
?????? ?????????
14
??? ??????? ?? ?????? ??? ?????? (?? ?????? ??
??????????) ????? ?????? (????? ??????
wave length)( ? ???? )
?????? ??????? ????? ?????? ????? ????? ? ????
????? ( 500 ) ??????? ???????
(0,5 um) ????? ?????? ?? ??????
???? ??? ??????? ??? ?????? (0,2 um) ???? ???
??????? ?????? ??? ????????? ?? ??? ???????
U.V.M. ????? ????? ?????? W.L.0,3um ?
???? R.P.0,1um ????? ?? ???????? ??? ???? ??
????? ?????? ???? ???? ???? ??? ??????? ???????
????
?
R.P
??????? ????????? ???? ????? ?????? ????
???? ?? ??????? ? ??????? ??????
?????? ?????????? ????? ?????? ?????? ??????????
? ???? ???? ?????????? ??? ???? ????
?????? ?????????? ?????? ??????
15
?????? ????
?? ????? ?????? ?????? ?????????? ????? ???
??????? ???????? ???? ????? ????? ??? ??????
????? ??????? ????? ?
??? ????? ??? ?? ???????? ?????????? ?????????
?????? ???? ???????? ?? ???? ????
????? ????? ?????? ?????? ?????????? ??????
?????????? ????????? ??????? ??? ? ?????
?????? ( ???? ) V ???????? ???????
??????
16
???? ??????? 60,000 ???? (60K.v.) ?????? ????????
??? ?????...........?
????
???? ??? ??? ????? ????? (0,5nm) ?? (0,05um)
???? ?????? ??? ?????? ?? ???? ??????? ?? ??????
?????? ??? ????? ?????? ??? ????????
17
?? ??????? ??? ???????? ?????? ?????? ???? ??
???? ??????? ??? ?????? ?????? ?? ?????? ?????
?????? ??? ????? ?? ??? ????? ?????? ?????. ???
???? ?? ?????? ?????? ?? ???? ??? ?? ????? ?????
??? ??????? ??? ??? ??? ?? (250nm) ??????
???? ?? ???? ??? ??? ?????? ???? ????????
???????? ??? ????? ?? (250nm) ?? ?? ??????
??????? ?????? ?????? ??? ?????? ??? ?????
??????? ???? ??? ??????? ??? ???????? ?? 250
???????
18
???? ??? ?? ?????? ?????? ???? ??????? ?? ??????
?????? ?? ????? ?????? ?????
????
???? ????? ?????? ???? ??????? ?????? ?????? ???
?? ???? ?? ???? ????? ?????? ?????? ?????? ?????
??? ????
19
??? ??????? ???? ????? ?????? ??????????
????????? ????? ?????? ?????? ?????????? ???
?????? ??????? ????? ??????? ????? ?? ?????
?????? ??? ?? ??????????? ??? ????? ?????? ?????
????? ??? ??? ???????? ????????? ?? ?????
??????????? ???? ??????? ??????????? ????? ????
?? ?????? ???? ?????? ??? 60.000 ???? ??
(60K.V) ??? (180K.V) ????? ??? ??????? ??????
???? ???? ????? ???? (1000K.V)
20
????? ??? ??????? ?? ??? (600,000X) ???? ???? ???
(1,600,000X) ?????? ????????? ???? ???????
??????????? ??? ?? ??? 0,2-0,1nm ?? ????? ???
??? ????? ??????? ????? ??????? ???????. ??????
?????? ?????????? ?? ??? ??????? ???? ????? ????
?????? ?????? ??? ?? ???????? ?? ???? ????? ??
?? ????? ???????? ??? ?????? ???? ??? ???
??????. ??? ???????? ???????? ??????? ????????
???? ????? ?? ?? ??? ???????????? ??? ??? ????
????? ???? ?????? ???? ??????. ????? ????? ???
????? ??? ??? ????? ??? ??? ????? ????
??????????? ??? ????? ??????? ??????? ??? ????
????? ??? ?????? ??????????.
21
?????? ?????? ???? ??? ?????? ??? ??????
?????????? ??????? ??????
E.M L.M





22
???? ???? ?????? ??????????
  • ????? ????? ?????? ??? ??? ???? ????? ???????
    ??????????? ???? ?????? ????? ???? ??????
    ???????????.
  • ??? ?? ?????? ?????? ??? ????? ?????? ????
    ??????? ????????? ????? . ?? ?? ??????? ???? ???
    ????? ??? ?? ????????? Photons ???????? ?? ???
    ????? ?????? Different wavelengths ?? ??????
    ?????? ??????

23
???? ???? ?????? ??????????
24
Basic structure
  • Electron microscope divided into the following
  • Basic components -
  • 1- Illumination system.
  • 2- Imaging system.
  • 3- Specimen stage.
  • 4- Vacuum pump system.
  • 5- Image recording system.

25
1- Illumination system.
  • 1- Cathode housing.

Electron gun
Cathode (filament)
Anode
2- Electromag Alignment system
26
???? ??????? ?? ???? ?????? ?????????? ??????? ??
????? ??????
  • ????
  • ????? ?? ??? ?????? Cathode???? ???? ???? ??????
    (??????) ?????? ???-
  • 1- ????? ??????????? Electron gun???? ????? ???
    ?????? Filament
  • ??? ?????? Cathode ??? ??? ??? ??? V .
  • ?????? / ?????? ???? ???? ?? ??? ????? ?????
    ?????? ????? ??????????? ??? ??????
  • ( ???? ???? ?????? ??? ????? ????? ???? ?????
    )
  • ???? ??? ????? ???????? ??? ????? ???? ?????
  • ??? ??? ???? ?? ( 3000 ???? ????? )
  • ????? ????? ????? ??????? ?? ??? ??????
    ????? ??? ??? ?????????.
  • ?????? ?? ?????? ????????? ????? ????????
    ?????????
  • ?? ??? 40 K.V ???? 12000 40000) K.V????(
  • ????? ????? ????? ??? ???? ?????? (???
    ?????? ) ??? ????? ????.

27
???? ??????? ?? ???? ?????? ?????????? ??????? ??
????? ??????
  • 2- ??????(wehneltyl cylinder) Anode ??? ????
    ?????? ??????? ????? ??? ??? ????
  • ?? ?????? ???? (1mm Dia.)?? ???? ??? ?????
    ??? ?????? ??????????
  • ??? ???? ???? ??? ??? ???? ?? ?????? ????
    ??????? ?????? ????? ??????????? ???
  • ?????? ??????? ????? ??? ?????? ?? ??????
    ???????

28
??????- Electromagnetic Alignment system ??
?????? ?????????????? ?????? ?? ????? ????????
????? ??? ???? ?????? ???? ???? ???? ????????
???? ?????? ?????????? ?????? ?????? ?????? ??
???????? ?? ???? ????? The electron beam can be
focused by passing trough a magnetic
field
29
2- Imaging system.
???? ????? ??????-
Lenses
??????? -
????? ?????? ?????????? ?????? ?? ???? ?????
????????? ???? ??? ????? ?????? ???????
???????? ??
1- ?????? ??????? Objective lens ( O.L. )

2- ?????? ??????? Projector lens ( P.L. )

3- ?????? ???????? Intermediate lens ( I.L.
)
4-?????? ??????? Condenser lens (C.L. )

30
???? ?? ???? ?? ???? ???? ??????? ?????? ??- 1-
??? ??????? ??? ????? ?? ??? ????? ????
?????? ???? ??????? ???? ??? ???? ???????
??? ???? ??? ???? ????? ? ?????? ??????????
?????? ?? ???? ?????? ????????? ???? ?????? ?????
?????? ????? ???? ???? ?????? ?????? ??
????? ??? ????? ??????????? ??? ?? ?????? ????
?????? ??????? ???? 2- ?????? ??????? ???? ????
??????? ??????? ?????? ?? ???? ??????
?????? ????? ?????? ?????? ??????????? ??? ???
?????? ????????? ???? ??? ???? ?????
????? ?? ???? ????? ?????? ?????? ??? ??????
??????? ??? ????? ?????? ??????? ???????????
??? ??? ????????? (3-5mm)
31
????? ????? ??????? ????????
  • 1- ???? ??? ?????? ??????? ?? ?????? ????????????
  • ?????? ??????? ??????? projector lens
  • ??? ???? ????? ?????? ??????? ?? ??????
    ??????.
  • 2- ????? ?????? ???????? Intermediate lens
  • 3- ?? ?????? ??????? Projector lens
  • ??? ?????? ???????? ?????? ???????? ????
    ?????? ??????? ??????? ??????
  • ????? ???? ????????/ ???????
  • 4- ??????? ?????? ??????? Objective lens
  • ??? ???? ???? ?? ?????? ?? ???? ??? ???
    ?????? Cathode
  • ??? ???? ??? ?????? ???? ????? ???????????
    ??? ?????? ?? ?????? (??????)
  • ???? ?????? ?????????? ?? ???? ????? ?????
    ????.

32
  • ???? ?????? ??????? ?? ???????? ????? ??? ??????
    ??????? ??????? ????????
  • ????? ?????? ?? (Aperture)
  • ????? ??? ??? ?????? ???? ????? ??????? ???
    ???????-
  • Apertune 125um, Apertune 250um Apertune
    3100um
  • ?????? ???? ?????? ??????? (??????) ??????
    ?????? ??????????? ??????? ????? ??? ??
  • ???? ?????? ???????? ????? ???? ??????? ?? ??????
    ????????
  • ???? ???? ???? ??? ??????? ???? ??????? (contrast
    diaphragm)

33
  • ??????? ???? ?????? ?????? (??????) ?????? ??????
    ??????????? ????????
  • ???? ?????? ??????? ????? ??????? ??? ??????
    ??????? ?????? ??????
  • ????? ???? ???? ?????? ??????? (selective
    diaphragm)
  • ??? ??? ?? ????? ?????? ???????? ?????? ???
    ?????? ???????? I.L.
  • ??????? ??????? ????? ?????? ???? ???? ?????
    ?????? ????
  • ????? ???? ??????? ??? ??????? ?????????

34
????? ???? ??????? Magnification instrument
???? ???????Magnification instrument ????? ??
????? ????? ??? ?????? ??? ????? ?????? ???????
??????? ???? ??????? ??? ??????? ???? ??????
?????? ?????? ???????
35
?????? ?????????? ??????
36
?????? ?????????? ??????
  • ?????? ???? ????? ??????? ?????? ??????
  • ???? ?????? ??? ??????? ?? ?????? ??????
  • ????? ??? ?????? ??????? ????? ??????????? ???
    ?????????? ???? ?????? ?????? ?????
  • ????? ??? ?????? ?? ??????? ??? ?? ?????? ??????
    .
  • ???? ???? ??????? ??????? ???? ???? ??? ?????
    ???? ???? ?? ???????????
  • ??? ????? ??????? ????? ????? ????? 5nm
  • ??????? ?? ??????? ??????? ?? ????? ????? ?? ???
    ?????? ???? ??????
  • ???? ?????? ?????? ???? ????? ?? ???? ??????
    ???????? ?? ????? ?????? ?????? ????? ??? ?????
    ?????? ?? ?????? .

37
???? ??????? Resolving
????? ?????? ?????? ?????? ?? ???? ??????? ??????
???? ?? ??????? . ??????? ??????? ( Condenser
lens ) ???? ??? ????? ????? ??? ???? ???????
????? ????? ???? ??????? ???? ?? ????? ???????
??? ???????? ??????? ???????? ( Objective
eye Piece lens ) ??? ?? ???? . ???? ???????????
???? ????? ?????? ??? ??? ?????? ???? ?????
??????? , ??????? ????? ??????? ????? ???? ?????
x25 ?x1500 ????? ?????? ??? ??? ?????? ??
???????? ??????? ???? ????? ??????? ?? ???? ????
????? ??????? , ????? ????? ???? ????? ??? ?????
???????? ???? ?? ?????? ??????? ?? ?????? ????? .
38
???? ??????? Resolving
  • ????? ??????? ??? ??????? ????? ??????? ???? ????
    ????? ?? ??????? . ???? ???? ???? ????? ???????
    ???? ?? ??? ???? ????? ???????? ?????? ??????? ,
    ????? ?????? ??????? ???? ?????? ??????? .
  • ??????? ???? ????? ?????? ?? ??????? ??? ?? ???
    ???? ???? .

39
?????? ?????????? ??????
?? ??? ????? ?? ??????? ??? ?????? ??? ???? ????
?? ??????????? ????? ?????? ?? ?? ???????????
????? ?????? ?????? ????? ??? ??????
???? ?????? ???? ?? ??? ??????????? ???? ?????
????????? ?????? ????? ????????? ?????? ??????
???? ??? ?????? ?????? ?????? ??? ???? ????
?????? 1965? ???? ??? ????? ???? ?? ??? ???? ??
??? ??????? ??????? ??????????? ????????? .
?????? ?????? ?????? ???? ???? ??????
40
?????? ???? ??????? ?? ?????? ?????? Resolving
Power
??? ???? ???? ??????????? ???? ???? ??????
???? ?????. ???? ???? ?????? ???????
????? ?????? ???????? ???????? ???? ?????? ????
???? ???????????
???? ?????? ??? ???? ??????? ?? ?????? ??? ???
10nm
41
Tissue Processing
????? ????? ?????? ???????? ???????? ????? ??????
?????? ??????????
42
Tissue Processing
  • ???????
    Fixation
  • ???? ???????
    Washing
  • ??? ?????
    Dehydration
  • ??????? ??????? ?????? Critical Point
    drying
  • ?????? ?? ??? ???????
  • Processing Specimens after drying
  • ??????? ??? ??????? ??????? Specimen stubs

43
??????? Fixation
  • ???? ??? ???????
  • ?????? ????? Primary
    Fixation
  • ?????? ??????????? 2-3 Bufferd glutraaldhyd
  • ??? ???? ????? ?????? ??? 2-3 ????? ?? ????
    ????? ?? ???????
  • ??? ??? ?? ????? ???? glycogen protein ??
    ?????? ????????

44
??????? Fixation
  • ?????? ??????? Secondary
    Fixation
  • ???????? ??? ????????? 1 Bufferd Osmic acid
  • Osmium tetroxid OsO4
  • ??? ???? ????? ??????
  • ??? ??? ??
  • 1- ????? ?????? ??????? Lipid ?? ?????? ????????
  • 2- ????? ????? ?????? ?????? ?????????? ??????
    ???????? ????????
  • 3- ???? ?? ????? ??????? ????????? ??? ??????
    ????? ????? ??????????? ???? ???? ??? ?????
    ??????? ???? ????? ??????? ?????????

45
????? ??????? ??????
  • ?????? ??? ?????? ?? ??????? ??????? .
  • ?????? ??? ?????? ?? ?????? ?????? ???? ?????????
    ?????????? .
  • ????? ?????? ?? ?? ??????? ??????? ????? ?????
    ?????? ?????? ??? ??? .
  • ????? ?????? ?? ???? ???????? .
  • ???? ????? ??? ???????? ??????? .
  • ???? ?????? ??? ?????? ?????? ?? ????? .

46
????? ???? ???? ??? ???????
  • ????? ?????? ?????? ??? ??? ??????
  • ??? ?????? ( ???? ????? )
  • ???? ??? ?????? ?? ??????
  • ??? ??????? ??? ?? ?????? ?? ??? ??????
  • ???? ???????
  • ???? ?????? ( ???? ???? ?????? ?? ?? ????
    ??? ???? ???? )
  • ??? ??????? ??????
  • ???? ???????
  • ?????????

47
???? ??????? Washing
  • ???? ?????? ??? ???????? ?????? ??? ??? ???? ???
    ?????? ?????? O.A , ??? ????? ?? ????? ?????
    ????? ???? ??? ??????
  • ????? ?? ?????? ?????? ?????? ?? ?????? ????
    ????? ????? ?? ??? ??????? ?????? ?? ??????
    ?????? ??????? .
  • ???? ????? ??? ?????? ???? ????? ???????

48
??????? dehydration
  • ????? ??? ????? ?? ??????
  • ?? ????? ?????? ?? ?????? ????? H2O ?? ??????
    ???????? ????????? ????????? ???? ?????? ???
    ????? ???????? ??? ????????? ??????????
  • ?? ?????? ???? ???????? ???? ?????? ???????? ??
    ????? ??????
  • ???? ??? ?? ??? ???? ?????? ?????? ??????? ??????
    ??? ??????? ??????? ????????
  • ???? ?????? ?????? ???? ??? ??? ???? ?????? ?????
    ??? ????? ???? ???? ???????
  • ????? ??? ??? ??? ????? ???? ???? ????? ????????

49
???? ?????? ??? ?????? ????? ?????? ????????
10 MIN 2X 30 ETHANOL
10 MIN 2X 50 ETHANOL
10 MIN 2X 70 ETHANOL
10 MIN 2X 90 ETHANOL
10 MIN 2X 95 ETHANOL
10 MIN 3X 100 ETHANOL
50
????? ???? ?????? ??? ?????
  • ?????? ?????? ????? ?????? ???????? ???????? ??
    30 ?????? ????????? ?? ?? ??? ????? ????????
    ???? ?? 5 ?? 10 ???? 100 ??? ?? ??????
    ??????????? ??? ???????? ???????? ?? ????? ??????
    ????? ??? ?????? ?? ???? ??? ????????? ???????
    ??????
  • ???????- ???????? ????????
  • ????????? ??? ??? ???????? ???? ??? ?????? ??
    ????? ??????
  • ????? ??????? (????? ??? ??? ??????)
  • ???????? ??? ???
  • ??????? (?????-????-?????) ???? ???.

51
???? ??? ?????????? ?????? ???? ??? ????? ???????
??????? ??????????
????? ?????? ???? ?????? ?? ???????? ?????????
??????
??? ?????? ?? ?? ???? ????? ??? ???? ?? ???????
????? ??? ???????
??????? ???? ?? ??? ??????? ?????? ??? ???????
52
?????? ?? ??????? ?????? ???? ??????? ??????
????? ??????? ?????????? ??????
53
(No Transcript)
54
(No Transcript)
55
(No Transcript)
56
(No Transcript)
57
??????
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