Numerical aperture and limits of resolution of microscope

1. Microscope ;
An instrument that is used to see
very small objects that can not be seen with naked
eye. The science of investigating small objects by using
an instrument is caleed miccroscopy.
Types of microscope;
1) Light Microscope.
2) Compound Microscope.

2. Working Principal
of light
microscope.

3. Important properties of microscope;
 Magnification ; ability to make
small objects larger or zooming
capability.
 Resolution ; ability to distinguish
two objects from each other or the
measure of clarity of an image.

4. Resolution limit
 The smallest distance between two objects at
which two small objects can still be seen as
separate objects or the lowest distinguishable
distance between 2 objects.
 It is the measuremet of the ability of the objective
lens to separate in the image adjacent details present
in the object.

5. Mathmatically ;

6. Resolving power
It is the reciprocal of limit of reolution.
Formula;

8. Hence d can be manipluated to by using the equation
and it is used to increase the resolution of microscope.
Factors that can increase the
resolving power of microscope by
decreasing d ;
 Wavelength of light.
 Numerical aperture of lens.

9. Effect of wavelength of light on
increasing the resolving power ;
• Resolving power is inversely propptional to
resolving power of microscope.
• By using the light of lower wavelength the
resolving power of a microscope can be
increased.
• Blue light and UV light have lower wavelength.
• Blue light of visible spectrum is mostly used in
microscope for having good resolution.

11. Numerical Aperture
 The numerical aperture of
microscope of an objective lens is
a measure of its ability to gather
light and resolve fine specimen
detail.
 Image forming light pass through
the specimen and enter the
objective lens in an inverted cone
shape.

12. Formula of NA ;
n×sintheta

13. Effect of NA on resolving power ;
NA is directly
related to resolving power of microscope.
Thus the greatest resolution is obtained by using a lens
with largest possible NA .
Resolution of microscope is increased by increasing
the two components of NA that are refractive index of
medium and the half angle of light entering cone.

14. When an air is a medium between lens and
specimen the maximum angle is 90 which is equal to
1 so the NA is one but when we use an immersion oil
more light is gathered on specimen and lens as a
result NA is increased as well as resolution is also
increased.
Immersion oil is used as a medium for having
high resolution ofmost microscope.
 Refractive index of air = 1.
 Refractive index of water=1.33
 Refractive index of immersion
oil=1.6_1.7

15. Working distance
The distance between the surface of lens and surface
of cover glass or specimen.

16. Working distance and numerical aperture ;
When the working distance is smaller the numerical
aperture increases as a result resolution also increases.

17. Thank you