TECHNIQUES

USED IN
CELL BIOLOGY-II
 INTRODUCTION
 What is the purpose of using different

techniques in cell biology.

 Why do we used differential staining?

 What is the purpose of microdissection

 Is tissue culture can be used for both plants

and animals?
CONTENTS

 Chromatography

 Electrophoresis

 Spectrophotometry

 Resolution and magnification in microscopy

 Micrometry
CHROMATOGRAPHY
Used to separate different chemical compounds from a

mixture

Paper chromatography is a simple and most widely used

technique

Involves two phases

Stationary phase

Mobile phase
CHROMATOGRAPHY

Mobile phase consists of a solvent in which

mixture sample is dissolved

Stationary phase consists of a filter paper

Mobile phase travels through the stationary phase

CHROMATOGRAPHY
Molecules of mixture sample begin to separate

as dots at different places on stationary phase

Paper is called Chromatogram

Apparatus is called Chromatography

Chamber
ELECTROPHORESIS
Used to separate fragments of charge bearing

polymer according to size, shape, molecular weight

and surface charge e.g. DNA, RNA, Protein

Utilizes a gel medium for separation of fragments

under the influence of electric field

ELECTROPHORESIS

Gel is sandwiched between glass to form a viscous slab

Two ends of the slab are suspended in two salt solutions

that are connected via electrodes to a power source

When voltage is applied, molecules present in the gel

migrate through the electric field

ELECTROPHORESIS
Negatively charges molecules will move towards the positive

pole

Molecules having positive charges will move towards the

negative pole

Velocity of movement of fragments is inversely proportional

to size

Smaller fragments move faster than larger

SPECTROPHOTOMETRY
Technique used to determine the absorption of

different wavelength of light by particular chemical

compound of a photosynthetic pigment

Spectrophotometer is an instrument that measures

amount of light that passes through the sample and

calculate how much light was absorbed

SPECTROPHOTOMETRY

Amount of light absorbed at each wavelength

is plotted in a graph i.e. absorption spectrum

Can be used to determine wavelengths of light

that take part in photosynthesis

RESOLUTION AND MAGNIFICATION IN MICROSCOPY

Our naked eye is capable to distinguish two points

which have atleast 0.1mm apart

Minimum capacity of a lens to differentiate between

two adjacent points is called resolution power of lens

Resolution of naked eye is 0.1 mm

RESOLUTION AND MAGNIFICATION IN
MICROSCOPY

Magnification is the capacity of an optical

instrument to increase the size of an object than

its original size

Objects which can’t be seen by naked eye can

be observed by increasing magnification

RESOLUTION AND MAGNIFICATION IN
MICROSCOPY

 Microscopy is a technique used to view objects

that cant be seen by the naked eye
 Range can be anything between mm and nm
 Most plants in animal cells are between 10μm and
30μm
MICROMETRY
 Objects can be measured under the microscope by

an eye piece

 This is a transparent scale mounted in the focal

plane of the eye piece

 It can be seen in the field of view at the same time as

an object is being examined under the microscope

MICROMETRY
 This can be done by placing a stage micrometre under the

microscope

 This is a glass slide on which a series of vertical lines by the

distances of 1.0mm, 0.1mm and 0.01mm by superimposing

the images of the eye piece and stage micrometre scales it is

possible to calibrate so that the size of a given object can be

estimated
RECAP
 Chromatography

 Electrophoresis

 Spectrophotometry

 Resolution and magnification in microscopy

 Micrometry
 HOME WORK

Explain the purpose of different

techniques used in Cell Biology

 CONCLUSION

Today, we have discussed some of the

techniques of Cell Biology, tomorrow we

will be discussing the Cell Wall.