This document discusses techniques for characterizing cell lines, including karyotyping and isozyme analysis. Karyotyping allows visualization of chromosome number and structure to identify genetic abnormalities. It provides information on species identification, genome stability, and numerical chromosome abnormalities. Isozyme analysis examines differences in enzyme structure and mobility between species using electrophoretic banding patterns. This can determine species identity and detect contamination. Common isozymes analyzed include LDH, MD, G6PD and NP. Both techniques are used to accurately characterize cell lines.

1. SHRI SHIVAJI SCIENCE COLLEGE,
AMRAVATI
2022 - 2023

2. Department Of Biotechnology
B.Sc.Part - III (Semester- V)
Topic- Characterization of cell lines -
1) Karyotyping
2) Isozyme analysis.
Presented By - Mayur
Anant Dakhore

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● The term cell line refer to the progogation of culture after
the first subculture.
● Once the primary culture is subculture, it becomes the cell
line.
● A given cell line contain several cell lineages of either
similar or distinct phenotype’s.

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What does Characterization
mean?
● Many traits of the cell line.
● Some of which may be unique.

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● Genetic instability
● Identify:- the species of origin.
● Relationship:- correlation with the tissue of origin.
● Purity – This test is used to detect a range of possible
contaminants, including bacteria, fungi, mycoplasma.
● To detect transformed cell line

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1) Karyotyping:-
✘ Karyotype is an organized profile of an
individual's chromosomes.
✘ Karyotyping is a technique that is use to examin
chromosomes in a sample of cells which can help
identify genetic problems as the cause of disorder
.

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Human
Karyotype:-
Breast cancer
Karyotype:-

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MAIN PURPOSE OF KARYOTYPING :-
✘Main purpose of the karyotyping is to
locate or visualize the changes in the
number of chromosomes and abnormality
in the structure.
✘Also to locate the evolution.

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Basis of karyotyping :-
● Basically karyotyping is based on
three patterns.
● On the basis of size of chromatids.
● On the basis of bending patterns.
● On the basis of centromeric positions.

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Karyotype of patient:- A: normal cell line 46,XY;
B: abnormal karyotype with the 18q deletion:-
A

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✘Species identification.
✘Index of genome stability.
✘Validation of normal diploid karyotype.
✘Numerical chromosomes abnormalities.
Through karyotype analysis, you will obtained
the following information about your cell lines
:-

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✘ Different chemical and
physical
✘ properties:
✘ Electrophoretic mobility.
✘ Kinetic properties.
✘ Amino acid sequence.
✘ Amino acid composition.
✘ Isoenzymes or
isozymes are
mutiple forms of
same enzyme that
catalyse the same
chemical reaction.
2) Isoenzyme Or Isozymes are :-

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Isoenzyme
Table :-
Isoenzyme
Name
Composition Electrophoretic
migration
Present in Elevated in
LDH 1 Heat
Resistance
(H4) Fastest moving Myocardium,
RBC,kidney
Myocardial
infarction
LDH 2Heat
Resistance
(H₂M₁) Myocardium,
RBC,kidney
Kidney
disease,megalob
lastic anemia
LDH 3 (H₂M₂) Brain Leukemia,malig
nancy
LDH 4 Heat
Labile
(H1M3) Lung,spleen Pulmonary
infarction
LDH Heat
labile
Inhibited by
urea
(M4) Slowest moving Skeletal muscle,
Liver
Skeletal muscle
and liver
diseases
● LDH:-Lactate
dehydrogenase.
● M - muscle -
chromosome
11(basic).
● H - heart -
chromosome
12(acidic).

14. Lactate dehydrogenase (LDH):-
The isozymes are also differentially expressed in different
developmental stages.
EXAMPLE:-
Before birth the heart is more anaerobic compared with
adulthood. Indeed, before birth the main isozyme in the heart is
the M4, and with time it switches to HM3 (at birth), to H2M2
and HM3at 1 year after birth, and to H3M and H4 after 2
14

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Isoenzyme Analysis:-
The isoenzyme analysis method utilizes
electrophoretic banding patterns to
examine slight differences from species
to species in the structure and mobility
of individual.

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● Isoenyzyme analysis can determine the species
identity of cell line .
● determine of your cell line if it has been
contaminated by a cell line of a different
species. From the banding patterns of the
enzymes used, a "process of elimination" is
employed to determine the identity of cell line of
species.

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Standard isoenzyme analyses included determination of the
isoenzyme gel electrophoresis banding pattern for at least three
of the following enzymes :-
● Glucose - 6 - Phosphate Dehydrogenase (G6PD)
● Lactate Dehydrogenase (LD)
● Malate Dehydrogenase (MD)
● Nucleoside Phosphorylase (NP)
● Aspartate Aminotransferase (AST)
● Peptidase B (PepB)
● G-Protein Coupled Receptors (GPCR)
● cAMP Phosphodiesterase Assay
● Protein Kinase Assay (PK)
● Other isoenzyme analyses are available

18. References :-
● Biotechnology,
By U Satyanarayan.
● Culture Of Animal Cells,
By R.Ian Freshney.

19. Thanks!
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Any Questions?