Dark
Field
Microsco
Presented by:
Ananya Rajappa B R
Fig: A colony of g r e e n planktonic
alg ae Volvox under the dark fi eld
py (23SM1017) Aniket Sharma
(23SM1019)
Ann Theress (23SM1020)
Anna Thomas
microscope. Image Credit: D. (23SM1021) Anoushka
Kucharski K.
Singh (23SM1022)
History
In 1830, J.J. Lister (the father of Joseph Lister)
invented the dark fi eld microscope, in which the
standard bright field (Abbe) condenser is
replaced with a single or double- refl ecting dark
fi eld condenser.
In 1906 in Vienna, Karl Landsteiner and Viktor
Mucha were the first to use dark fi eld microscope to
visualise T pallidum from syphilis lesions.
What is Dark Field
Microscopy?
Dark Field Microscopy is used to examine live
micro- organisms that are either invisible in the
ordinary light microscope, cannot b e stained by
standard methods, or are so distorted by staining
that their characteristics then cannot b e identifi ed.
In dark fi eld microscopy, the object appears bright
against a dark background.
Principle
• Bright image on a dark background
• This is similar to the ordinary light microscope; that has
a simple
modifi cation
•The
Light * Specimen
light path consists of
source stage
• Condenser
Op aque * Objective
* Ocular lens
An dise
opaque dise is placedlensb e t w e e n the light source and the
condenser lens.
• This disc blocks most of the light from the illuminator.
Hence it
produces a hollow c o n e of light.
• The only light that reaches the objective lens is light that
has b e e n refracted, refl ected or scattered b y the
specimen.
• Other lights that don't hit the sample, are not passed through
objective lens. They are passed around the objective lens. As
Applications
▶ Enhanced contrast for transparent structures
▶ Visualisation of thin bacteria
▶ Rapid detection of Treponema pallidum
▶ Observation of Bacterial and Protozoan motility
▶ Study of marine organisms and materials like
crystals and polymers
▶ Examination of cells and tissues
▶ Surface and e d g e detail examination
Advantages
▶ Simple and cost-eff ective setup
▶ Ideal for Unstained, Transparent
Specimens
▶ Higher resolution and contrast
▶ No sample preparation needed
▶ Suitable for observing live organisms
▶ Detailed view of external features
▶ Eff ective for water-borne organisms
Disadvantages
▶ Images formed are prone to degradation,
distortion and inaccuracies
▶ Since the sample has to b e adequately
illuminated,
sometimes strong illumination may damage the
specimen
▶ Not only the specimen but dust and other
particles
scatter the light and are easily observed
▶ Low Level of light and limited colours
Advancements
▶ Integration with Fluorescence Techniques
▶ Digital imaging and Real-time analysis
▶ Label-free and non-invasive imaging
▶ Portability and Accessibility
▶ Enhanced contrast for organisms without
staining,
preserving their natural state
▶ Super Resolution Techniques extending
visibility to nanoscale structures
Thank
you
