Introduction to Microscopy

Types of MicroscopesTypes of Microscopes

Dissection or StereoscopeDissection or Stereoscope

SEM ( Scanning Electron Microscope)SEM ( Scanning Electron Microscope)

TEM ( Transmission Electron TEM ( Transmission Electron Microscope)Microscope)

Compound ( Light) Microscope***Compound ( Light) Microscope***

Difference between Difference between Magnification and Magnification and

ResolutionResolution MagnificationMagnification: is just power of an instrument to enlarge : is just power of an instrument to enlarge

things...things...Just as you would increase the size of a picture in your PC by Just as you would increase the size of a picture in your PC by zoom function. It would enlarge it...zoom function. It would enlarge it...

ResolutionResolution : is power to show two closely situated particles : is power to show two closely situated particles as different... that obviously improves quality of the picture..as different... that obviously improves quality of the picture..

The magnification of human eye is 1 and the resolution is 1 The magnification of human eye is 1 and the resolution is 1 micrometer.micrometer.i.e. The eye does not magnify a thing and two particles i.e. The eye does not magnify a thing and two particles spaced at a distance of 1 micrometer will be able to be seen spaced at a distance of 1 micrometer will be able to be seen different.different.No wonder objects closer than this will be seen as one. No wonder objects closer than this will be seen as one.

Human hair: 50 micrometersHuman hair: 50 micrometers

Dissection or Dissection or StereoscopeStereoscope

Used to get a better look at a larger Used to get a better look at a larger specimenspecimen

Low magnification- 10X- 40XLow magnification- 10X- 40X

3-D image3-D image

Light illuminatedLight illuminated

Dissection Microscope

Spider- Dissection Microscope

Scanning Electron Scanning Electron MicroscopeMicroscope

SEMSEM Used to examine very small, multicellular, Used to examine very small, multicellular,

anatomical structuresanatomical structures 3-D structures3-D structures Specimen is coated with metalSpecimen is coated with metal Electron beam is swept across specimen Electron beam is swept across specimen

causing electrons from specimen to get causing electrons from specimen to get excited causing secondary electrons to be excited causing secondary electrons to be emitted and collectedemitted and collected

Electrons are detected and an image is Electrons are detected and an image is createdcreated

High Magnification/High Resolution High Magnification/High Resolution

Scanning Electron Microscope

Human Egg Sitting on the point of a pin-

Follicle cell to nurture cell

Scanning Electron Microscope

Magnification: X 500

Black Widow Spider Claw

Scanning Electron Microscope

Mascara brush with mascara and flakes of skin-50X

Scanning Electron Microscope

Scanning Electron Microscope

Velcro- hook and loop-35X

Transmission Electron Transmission Electron MicroscopeMicroscope

Works similar to a slide projector: Works similar to a slide projector: beam of electrons ( not light) passes beam of electrons ( not light) passes through specimen and projected on through specimen and projected on screen below.screen below.

2-D view of internal organelles2-D view of internal organelles Specimen sliced thinSpecimen sliced thin Very high resolution and high Very high resolution and high

magnification. Much higher than a magnification. Much higher than a compound microscopecompound microscope

Transmission Electron Microscope

Normal Collagen Degenerating Collagen

A pair of images of collagen fibrils in the eye of a patient with glaucoma revealed.

Transmission Electron Microscope

Cell undergoing Apoptosis- committing suicide

Transmission Electron Microscope

Compound Light Compound Light MicroscopeMicroscope

Light IlluminatedLight Illuminated Image is 2-DImage is 2-D High Magnification but low resolutionHigh Magnification but low resolution View cells that are living- example View cells that are living- example

algaealgae Tissue samples may be sectioned, Tissue samples may be sectioned,

stained and put onto a slidestained and put onto a slide There are several different There are several different

magnification lensesmagnification lenses

Frog’s blood- 400X Magnified

Compound Light Microscope

Frog’s Blood- 1000X Magnified

Compound Light Microscope

•What is visible with a Compound Light Microscope?

Animal Cell Plant Cell Bacterium

Cell Membrane Cell Wall It's SO SMALL,

all you will see is

The Cell Envelope

(Composed of,from the

inside/outa cell

membrane,a cell wall,

and in somespecies, a capsule.)

Cytoplasm Cell Membrane

Nuclear Membrane

Cytoplasm

NucleusNuclear

Membrane

Chromatin(Its in there,even if you

can'tsee the detail!)

Nucleus

Chromatin

Chloroplast

high=40X; low=10X; scanning=4Xhigh=40X; low=10X; scanning=4X

changes objective lenseschanges objective lenses

directs light into microscopedirects light into microscope

focuses lightfocuses light

amount of light entering the microscopeamount of light entering the microscope

brings object into slow focusbrings object into slow focus

look through; lens is 10Xlook through; lens is 10X

carry and support carry and support

supports slide; positions slidesupports slide; positions slide

brings object into rapid focusbrings object into rapid focus

Type of Light MicroscopyType of Light Microscopy

IMMUNOFLUORESCENCEIMMUNOFLUORESCENCE Specific molecules are located within the Specific molecules are located within the

cell by an antibody produced against the cell by an antibody produced against the molecule of interest.molecule of interest.

The antibody- fluor attaches to the The antibody- fluor attaches to the molecule and under a light microscope molecule and under a light microscope the molecule can be observed.the molecule can be observed.

This can be used in order to see whether This can be used in order to see whether certain molecules are present or not certain molecules are present or not present and at what concentration.present and at what concentration.

Telomeres on Metaphase Telomeres on Metaphase ChromosomesChromosomes