2. Table of Contents
what is the
cytokines?
04
Direct and Indirect
Detection Methods
of protein ?
03
01
difference between a
primary antibody and a
secondary antibody?
02
Definition and grouping
of immunology
6. Definition
Immunology is a branch of the biology involved with the study of the immune
system, components of the immune system, its biological processes, the
physiological functioning of the immune system, types, its disorder .
The immune system acts as a body’s defence system by protecting our body
cells, tissues and organs from invading infection through various lines of
defence. Overall, the immune system functions by recognising and destroying
foreign antigens including the harmful microorganisms and other disease-
causing microbes.
7. Types of Immune System
humans, have two types of Immune System and are classified
based on its resistance power against the infectious agents
Innate
Immune
System.
Adaptive
Immune
System.
8. Innate Immune System
Innate Immune System is composed of cells and proteins
that are always present and are ready to fight against microbes in
the infection area. Innate Immune System is present from the time
of our birth.
Main elements of the innate immune system are –
Dendritic cells.
Phagocytic leukocytes.
Natural killer (NK) cell.
Physical epithelial barriers.
Circulating plasma proteins.
9.
10. Adaptive Immune System
The Adaptive Immune System is required to fight against pathogens
that cannot be controlled by innate immune defenses. It is also
referred to as the Acquired Immune System because it is acquired
during the course of life.
Two Types of adaptive responses are – humoral immunity
moderated by antibodies which are developed by B lymphocytes and
cell-mediated immunity, moderated by T Lymphocytes.
11.
12.
13. What is an antibody?
1. An antibody is a protein component of immune responses against
invaders, such as viruses or bacteria. After detecting foreign substances
on the invaders, also known as antigens, the immune responses generate
an antibody or an immunoglobulin.
2. What are primary antibodies and secondary antibodies?
Primary antibodies: antibodies that bind specifically to an
antigen.
Secondary antibodies: antibodies that bind to primary
antibodies.
14.
15. What is the main difference between a primary antibody
and a secondary antibody?
The difference between a primary antibody and a secondary
antibody primarily lies on the binding to an antigen.
1. A primary antibody binds directly to a particular antigen, whereas a secondary antibody doesn’t
bind to the target antigen. Instead, it binds to the primary antibody.
2. During antibody production, the target antigen and host species used to produce the primary
antibodies are different from those used to make the secondary antibodies.
16. Direct and Indirect Detection Methods
of protein ?
1. Antibody detection of the target protein is accomplished using a one-step
or two-step protocol. The one-step procedure, direct detection, relies upon
a single antibody that has been covalently joined to an easily detected
label molecule (biotin, an enzyme, or a fluorescent dye).
2. indirect detection, two different antibodies are used in sequence for the
detection step. First, the Western blot is incubated with an unlabeled
primary antibody directed against the target protein. After washing, a
labeled secondary antibody is used to detect the presence of the primary
antibody, and thus the target protein. The labeled secondary antibody is
typically directed against the immunoglobulin class or subclass of the
primary antibody’s species.
18. Comparison of Direct and Indirect Detection Methods
Direct Detection Indirect Detection
Advantages Advantages
1 Faster overall, since there are fewer
steps.
2 Less chance of non-specific signal.
o Often gives a stronger signal because
multiple secondary antibodies bind to
each primary antibody.
o Easy to change label type or detection
methods for a new experiment by
swapping secondaries.
o Saves labeling time and expense,
especially when all primary antibodies
are made in the same species.
o Provides access to a wider range of
labels.
Disadvantages Disadvantages
o Coupling of label to the primary
antibody may affect the antibody’s
ability to bind to the target protein.
o Labeling every primary antibody adds
time and cost.
o More non-specific signal can arise from
the binding of the secondary antibody
to other proteins on the blot.
o Extra incubation and wash steps add
time to the experiment.
19. Cytokines
1. Cytokines are peptides that have a fundamental role in
communication within the immune system and in allowing the
immune system and host tissue cells to exchange information.
2. Cytokines act via binding to a receptor that in turn sends a signal
to the recipient cell, leading to a change in function or phenotype.
Such signal cascades are complex and integrate a variety of
environmental factors.
3. Cytokines exist in broad families that are structurally related but
exhibit diverse function (e.g., the TNF/TNF receptor superfamily,
interleukin [IL]-1 superfamily, and IL-6 superfamily).
21. References
1. Iain B. McInnes, in Kelley and Firestein's Textbook of Rheumatology (Tenth Edition), 2017.
2. Watson, J. D., Baker, T. A., Bell, S. P., Gann, A., Levine, M., Losick, R. (2013). Molecular Biology of the
Gene. United Kingdom: Pearson Education.
3. Brandt, et al. (2019). Evaluation of Patient IgM and IgG Reactivity Against Multiple Antigens for
Improvement of Serodiagnostic Testing for Early Lyme Disease. Frontiers in Public Health, 7, pp.
370.
4. Yu, H.-W., Halonen, M. J., & Pepper, I. L. (2015, January 1). Chapter 12 - Immunological Methods (I. L.
Pepper, C. P. Gerba, & T. J. Gentry, Eds.). ScienceDirect; Academic Press.
https://www.sciencedirect.com/science/article/pii/...
5. Oettgen, H.C. (2017). Fifty Years Later: Emerging Functions Of Ige Antibodies In Host Defense,
Immune Regulation, And Allergic Diseases. The Journal Of Allergy And Clinical Immunology,
137(6), pp. 1631–1645.
