29 - Tool of Laboratory (Microscopy)

Ocular Lens

Eyepiece you look through which provides minimal magnification

Objective Lens

Major step in magnification, difference lenses have difference magnification

Magnification

Ability to enlarge the apparent size of an object/organism 

Resolution

Ability to distinguish 2 seperate objects (dictated by wavelength of light and property of the microscope)

Light Microscopy

Uses visible light, resolution 200nm, cheap and easy and works on live or fixed samples, samples are colorless with low resolution/magnification

Gram Stain

Stain used to classify bacteria based on their cell wall structure (either gram+ or gram-)

Gram + Gram Stain

Large peptidoglycan cell wall on the exterior of the cell, purple

Gram - Gram Stain

Has Peptidoglycan cell wall but has an outer cell membrane, pink

Causative Agent of TB

Mycobacterium Tuberculosis

What bacteria has a waxy/lipid rich that doesn’t easily gram stain?

Mycobacterium Tuberculosis

Fluorescence Microscopy

Uses visible light (laser/halogen), resolution 200nm, labels specific structures and works on live/fixed samples

Fluorescence

Molecule that absorbs light at 1 wavelength and emits light at a sighly larger wavelength 

Specific Fluorescent Markers

Mitotracker Red, Alexa Fluor 488/Phalloidin, Hoechst

Mitotracker Red

Detects mitochondria (Red)

Alexa Fluor 488/Phalloidin

Detects actin (Green)

Hoechst

Detects DNA (Blue)

Confocal Microscopy

Uses visible light (laser/halogen), resolution of 200nm, highlights specific structures, focuses on single planes, and can make 3D images

What does Confocal Microscopy do to get its image? 

Takes a series of images and takes all the planes of the images and makes a much sharper image (improving the resolution)

What is resolution linked to?

Wavelength and Illumination Source

Elections have shorter wavelength, so…

Higher resolution power

Transmission Electron Microscopy

Uses electronic beam, resolution 0.2nm, high magnification/resolution to see fine details, requires fixed/toxic samples, slow and expensive

What does Transmission Electron Microscopy do to get its image?

Uses thin sliced sections of the organism/tissue

Scanning Electron Microscopy

Uses electron beam, resolution 10nm, high magnification to show fine external structures, scans unsliced surfaces, requires fixed/toxic samples