Monoclonal antibodies (MAbs) are identical antibodies produced by a single parent cell. MAbs can be created that specifically bind to any substance and then serve to detect or purify that substance. The production of MAbs involves immunizing an animal, fusing the animal's immune B cells with myeloma cells to form a hybridoma, selecting the hybridomas that produce the desired MAb, and propagating the hybridomas to produce large amounts of MAbs. MAbs have many applications including use as diagnostic, therapeutic, and research tools due to their homogeneity, specificity, and ability to be produced in unlimited quantities.
Immunity
It can be defined as the resistance to disease, specifically to infectious disease or pathogens. The term “immune” is derived from the Latin word “immunis” that is exempt from charges. In medical term, it refers to the being protected from infectious pathogens.
Immune system
It is adaptive defense system which is able to generate a variety of cell and molecules capable of specifically recognizing and eliminating a variety of limitless foreign invaders into the system.
In 1975 Georges Kohler and Milstein succeeded in making fusions of myeloma cell lines with B cells to create hybridomas that could produce antibodies.
antibody
Also known as immunoglobulin is a large, Y shaped glycoprotein produced mainly by plasma cells that is used by the immune system to neutralize pathogens.
monoclonal antibodies
Antibodies that are made by identical immune cells that are clones of a unique parent cell.
polyclonal antibodies
A polyclonal antibodies represents a collection of antibodies from different B cells that recognize multiple epitopes on the same antigen.
Immunity
It can be defined as the resistance to disease, specifically to infectious disease or pathogens. The term “immune” is derived from the Latin word “immunis” that is exempt from charges. In medical term, it refers to the being protected from infectious pathogens.
Immune system
It is adaptive defense system which is able to generate a variety of cell and molecules capable of specifically recognizing and eliminating a variety of limitless foreign invaders into the system.
In 1975 Georges Kohler and Milstein succeeded in making fusions of myeloma cell lines with B cells to create hybridomas that could produce antibodies.
antibody
Also known as immunoglobulin is a large, Y shaped glycoprotein produced mainly by plasma cells that is used by the immune system to neutralize pathogens.
monoclonal antibodies
Antibodies that are made by identical immune cells that are clones of a unique parent cell.
polyclonal antibodies
A polyclonal antibodies represents a collection of antibodies from different B cells that recognize multiple epitopes on the same antigen.
Monoclonal antibodies (mAb or moAb) are antibodies that are made by identical immune cells that are all clones of a unique parent cell. Monoclonal antibodies can have monovalent affinity, in that they bind to the same epitope (the part of an antigen that is recognized by the antibody). In contrast, polyclonal antibodies bind to multiple epitopes and are usually made by several different plasma cell (antibody secreting immune cell) lineages. Bispecific monoclonal antibodies can also be engineered, by increasing the therapeutic targets of one single monoclonal antibody to two epitopes. Given almost any substance, it is possible to produce monoclonal antibodies that specifically bind to that substance; they can then serve to detect or purify that substance. This has become an important tool in biochemistry, molecular biology, and medicine. When used as medications, non-proprietary drug names end in -mab and many immunotherapy specialists use the word mab anacronymically.
Hybridoma technology is a method for producing large numbers of identical antibodies (also called monoclonal antibodies). This process starts by injecting a mouse (or other mammals) with an antigen that provokes an immune response.
Hybridoma technology is a method for producing large number of identical antibodies called monoclonal antibodies.
It was discovered by G.kohler and C.milstein in 1975. they were awarded nobel prize for physiology and medicine in 1975.
The hybrid cells are produced by fusing B- lumphocyte with myeloma cells or tumour cells.
The B-lymphocyte have the ability to produce large number of antibodies and tumour cells have indefinite growth.
This is why two cells are used for the production of hybrid cell
Production and applications of monoclonal antibodiesKaayathri Devi
production and applications of monoclonal antibodies, monoclonal antibodies ,applications of monoclonal antibodies, production of monoclonal antibodies,
INTRODUCTION: Monoclonal antibodies can be produced through a technique known as hybridoma technology.
HISTORY: The production of monoclonal antibodies was invented by Niels K.J. Georges, J.F. Kohler and Cesar Milstein in 1975.
PRINCIPLE FOR CREATION OF HYBRIDOMA CELLS: HAT (hypoxanthine aminopterin and thymidine) medium – Only hybridoma cells can proliferate in HAT medium.
PRODUCTION OF MONOCLONAL ANTIBODIES (HYBRIDOMA TECHNOLOGY): The establishment of hybridomas and production of monoclonal antibodies involves the following steps-
Immunization (ii) Cell fusion (iii) Selection of hybridomas (iv) Screening the products (v) Cloning and propagation (vi) Characterization and storage.
ADVANTAGES AND DISADVANTAGES OF MONOCLONAL ANTIBODIES:
Advantages- Monoclonal antibodies is specific to a given antigenic determinant.
Disadvantages- There is no guarantee that monoclonal antibodies produced is totally virus-free, despite the purification.
APPLICATIONS OF MONOCLONAL ANTIBODIES: Diagnostic applications, therapeutic applications, protein purification and miscellaneous applications.
REFERENCES:
• Satyanarayana, U. 2016. Biotechnology. Books and Allied (P) Ltd, Kolkata. pp. 213-226.
• Gupta, P.K. 2016. Biotechnology and Genomics. Rastogi Publications, Meerut. pp. 299-311.
• Owen, J.A., Punt J., Stranford, S.A. and Patricia, P.J. 2013. Kuby Immunology. 7th Ed. W.H. Freeman and Company, New York. pp.645-655.
• Singh, B.D. 2017. Biotechnology Expanding Horizons. Kalyani Publishers, New Delhi. pp. 172-174.
• Dubey, R.C. and Maheshwari, D.K. 2018. A Textbook of Microbiology. S Chand and Company Limited, New Delhi. pp. 662-663.
Monoclonal Antibody-Preparation & Application - MPH201T.pptxRAHUL PAL
Monoclonal antibodies (mAbs) are proteins produced by a single type of B cell. They are identical to each other and recognize a specific antigen. Antigens are molecules that the body's immune system recognizes as foreign. When an antigen binds to a monoclonal antibody, it triggers a series of reactions that can lead to the destruction of the antigen.
Monoclonal antibodies can be used to treat a variety of diseases, including cancer, autoimmune diseases, and infections. They are also used in research and diagnostics.
Monoclonal antibodies (mAb or moAb) are antibodies that are made by identical immune cells that are all clones of a unique parent cell. Monoclonal antibodies can have monovalent affinity, in that they bind to the same epitope (the part of an antigen that is recognized by the antibody). In contrast, polyclonal antibodies bind to multiple epitopes and are usually made by several different plasma cell (antibody secreting immune cell) lineages. Bispecific monoclonal antibodies can also be engineered, by increasing the therapeutic targets of one single monoclonal antibody to two epitopes. Given almost any substance, it is possible to produce monoclonal antibodies that specifically bind to that substance; they can then serve to detect or purify that substance. This has become an important tool in biochemistry, molecular biology, and medicine. When used as medications, non-proprietary drug names end in -mab and many immunotherapy specialists use the word mab anacronymically.
Hybridoma technology is a method for producing large numbers of identical antibodies (also called monoclonal antibodies). This process starts by injecting a mouse (or other mammals) with an antigen that provokes an immune response.
Hybridoma technology is a method for producing large number of identical antibodies called monoclonal antibodies.
It was discovered by G.kohler and C.milstein in 1975. they were awarded nobel prize for physiology and medicine in 1975.
The hybrid cells are produced by fusing B- lumphocyte with myeloma cells or tumour cells.
The B-lymphocyte have the ability to produce large number of antibodies and tumour cells have indefinite growth.
This is why two cells are used for the production of hybrid cell
Production and applications of monoclonal antibodiesKaayathri Devi
production and applications of monoclonal antibodies, monoclonal antibodies ,applications of monoclonal antibodies, production of monoclonal antibodies,
INTRODUCTION: Monoclonal antibodies can be produced through a technique known as hybridoma technology.
HISTORY: The production of monoclonal antibodies was invented by Niels K.J. Georges, J.F. Kohler and Cesar Milstein in 1975.
PRINCIPLE FOR CREATION OF HYBRIDOMA CELLS: HAT (hypoxanthine aminopterin and thymidine) medium – Only hybridoma cells can proliferate in HAT medium.
PRODUCTION OF MONOCLONAL ANTIBODIES (HYBRIDOMA TECHNOLOGY): The establishment of hybridomas and production of monoclonal antibodies involves the following steps-
Immunization (ii) Cell fusion (iii) Selection of hybridomas (iv) Screening the products (v) Cloning and propagation (vi) Characterization and storage.
ADVANTAGES AND DISADVANTAGES OF MONOCLONAL ANTIBODIES:
Advantages- Monoclonal antibodies is specific to a given antigenic determinant.
Disadvantages- There is no guarantee that monoclonal antibodies produced is totally virus-free, despite the purification.
APPLICATIONS OF MONOCLONAL ANTIBODIES: Diagnostic applications, therapeutic applications, protein purification and miscellaneous applications.
REFERENCES:
• Satyanarayana, U. 2016. Biotechnology. Books and Allied (P) Ltd, Kolkata. pp. 213-226.
• Gupta, P.K. 2016. Biotechnology and Genomics. Rastogi Publications, Meerut. pp. 299-311.
• Owen, J.A., Punt J., Stranford, S.A. and Patricia, P.J. 2013. Kuby Immunology. 7th Ed. W.H. Freeman and Company, New York. pp.645-655.
• Singh, B.D. 2017. Biotechnology Expanding Horizons. Kalyani Publishers, New Delhi. pp. 172-174.
• Dubey, R.C. and Maheshwari, D.K. 2018. A Textbook of Microbiology. S Chand and Company Limited, New Delhi. pp. 662-663.
Monoclonal Antibody-Preparation & Application - MPH201T.pptxRAHUL PAL
Monoclonal antibodies (mAbs) are proteins produced by a single type of B cell. They are identical to each other and recognize a specific antigen. Antigens are molecules that the body's immune system recognizes as foreign. When an antigen binds to a monoclonal antibody, it triggers a series of reactions that can lead to the destruction of the antigen.
Monoclonal antibodies can be used to treat a variety of diseases, including cancer, autoimmune diseases, and infections. They are also used in research and diagnostics.
Topics included :- Introduction to monoclonal antibody; Principle for creation of hybridoma cells and steps involved in it; Second generation monoclonal antibodies; Advantages, disadvantages and applications (Diagnostic and therapeutic) of MAbs.
Waste water treatment is a process used to convert wastewater into an effluent that can be returned to the water cycle with minimum impact on the environment, or directly reused. The latter is called water reclamation because treated wastewater can then be used for other purposes.
Industrial microbiology is a branch of applied microbiology in which microorganisms are used in industrial processes; for example, in the production of high-value products such as drugs, chemicals, fuels and electricity.
An ecosystem is a community of living organisms in conjunction with the nonliving components of their environment, interacting as a system. These biotic and abiotic components are linked together through nutrient cycles and energy flows.
Biochemistry, sometimes called biological chemistry, is the study of chemical processes within and relating to living organisms.[1] Biochemical processes give rise to the complexity of life.
Overpopulation is an undesirable condition where the number of existing human population exceeds the carrying capacity of Earth. Overpopulation is caused by number of factors. Reduced mortality rate, better medical facilities, depletion of precious resources are few of the causes which results in overpopulation.
Hypersensitivity (also called hypersensitivity reaction or intolerance) refers to undesirable reactions produced by the normal immune system, including allergies and autoimmunity.
Lipids include fats, waxes, phospholipids, sterols, such as cholesterol, and fat-soluble vitamins. Broadly speaking, there are three possible sites where lipids are synthesized: the smooth endoplasmic reticulum (SER), the cytosol and, in plants specifically, the chloroplast.The ER and Golgi apparatus together constitute the endomembrane compartment in the cytoplasm of eukaryotic cells. The endomembrane compartment is a major site of lipid synthesis, and the ER is where not only lipids are synthesized, but membrane-bound proteins and secretory proteins are also made.
There are different bacterial characteristics . These include cultural characteristics, cellular characteristics, and biochemical characteristics. The simplest of these to observe are the cultural characteristics. Learning the cultural characteristics of the organisms we use in the lab will help you greatly, especially when working on your morphological unknown.
the branch of science concerned with the chemical and physico-chemical processes and substances that occur within living organisms.
the processes and substances with which the science of biochemistry is concerned.
The digestion of certain fats begins in the mouth, where short-chain lipids break down into diglycerides because of lingual lipase. The fat present in the small intestine stimulates the release of lipase from the pancreas, and bile from the liver enables the breakdown of fats into fatty acids.
Protein is an important component of every cell in the body. Hair and nails are mostly made of protein. Your body uses protein to build and repair tissues. You also use protein to make enzymes, hormones, and other body chemicals. Protein is an important building block of bones, muscles, cartilage, skin, and blood.
Immune System Organs. The key primary lymphoid organs of the immune system include the thymus and bone marrow, as well as secondary lymphatic tissues including spleen, tonsils, lymph vessels, lymph nodes, adenoids, skin, and liver.
In cellular biology, membrane transport refers to the collection of mechanisms that regulate the passage of solutes such as ions and small molecules through biological membranes, which are lipid bilayers that contain proteins embedded in them.
Antigen-antibody interaction, or antigen-antibody reaction, is a specific chemical interaction between antibodies produced by B cells of the white blood cells and antigens during immune reaction. ... The specificity of the binding is due to specific chemical constitution of each antibody.
Earliest Galaxies in the JADES Origins Field: Luminosity Function and Cosmic ...Sérgio Sacani
We characterize the earliest galaxy population in the JADES Origins Field (JOF), the deepest
imaging field observed with JWST. We make use of the ancillary Hubble optical images (5 filters
spanning 0.4−0.9µm) and novel JWST images with 14 filters spanning 0.8−5µm, including 7 mediumband filters, and reaching total exposure times of up to 46 hours per filter. We combine all our data
at > 2.3µm to construct an ultradeep image, reaching as deep as ≈ 31.4 AB mag in the stack and
30.3-31.0 AB mag (5σ, r = 0.1” circular aperture) in individual filters. We measure photometric
redshifts and use robust selection criteria to identify a sample of eight galaxy candidates at redshifts
z = 11.5 − 15. These objects show compact half-light radii of R1/2 ∼ 50 − 200pc, stellar masses of
M⋆ ∼ 107−108M⊙, and star-formation rates of SFR ∼ 0.1−1 M⊙ yr−1
. Our search finds no candidates
at 15 < z < 20, placing upper limits at these redshifts. We develop a forward modeling approach to
infer the properties of the evolving luminosity function without binning in redshift or luminosity that
marginalizes over the photometric redshift uncertainty of our candidate galaxies and incorporates the
impact of non-detections. We find a z = 12 luminosity function in good agreement with prior results,
and that the luminosity function normalization and UV luminosity density decline by a factor of ∼ 2.5
from z = 12 to z = 14. We discuss the possible implications of our results in the context of theoretical
models for evolution of the dark matter halo mass function.
Multi-source connectivity as the driver of solar wind variability in the heli...Sérgio Sacani
The ambient solar wind that flls the heliosphere originates from multiple
sources in the solar corona and is highly structured. It is often described
as high-speed, relatively homogeneous, plasma streams from coronal
holes and slow-speed, highly variable, streams whose source regions are
under debate. A key goal of ESA/NASA’s Solar Orbiter mission is to identify
solar wind sources and understand what drives the complexity seen in the
heliosphere. By combining magnetic feld modelling and spectroscopic
techniques with high-resolution observations and measurements, we show
that the solar wind variability detected in situ by Solar Orbiter in March
2022 is driven by spatio-temporal changes in the magnetic connectivity to
multiple sources in the solar atmosphere. The magnetic feld footpoints
connected to the spacecraft moved from the boundaries of a coronal hole
to one active region (12961) and then across to another region (12957). This
is refected in the in situ measurements, which show the transition from fast
to highly Alfvénic then to slow solar wind that is disrupted by the arrival of
a coronal mass ejection. Our results describe solar wind variability at 0.5 au
but are applicable to near-Earth observatories.
Slide 1: Title Slide
Extrachromosomal Inheritance
Slide 2: Introduction to Extrachromosomal Inheritance
Definition: Extrachromosomal inheritance refers to the transmission of genetic material that is not found within the nucleus.
Key Components: Involves genes located in mitochondria, chloroplasts, and plasmids.
Slide 3: Mitochondrial Inheritance
Mitochondria: Organelles responsible for energy production.
Mitochondrial DNA (mtDNA): Circular DNA molecule found in mitochondria.
Inheritance Pattern: Maternally inherited, meaning it is passed from mothers to all their offspring.
Diseases: Examples include Leber’s hereditary optic neuropathy (LHON) and mitochondrial myopathy.
Slide 4: Chloroplast Inheritance
Chloroplasts: Organelles responsible for photosynthesis in plants.
Chloroplast DNA (cpDNA): Circular DNA molecule found in chloroplasts.
Inheritance Pattern: Often maternally inherited in most plants, but can vary in some species.
Examples: Variegation in plants, where leaf color patterns are determined by chloroplast DNA.
Slide 5: Plasmid Inheritance
Plasmids: Small, circular DNA molecules found in bacteria and some eukaryotes.
Features: Can carry antibiotic resistance genes and can be transferred between cells through processes like conjugation.
Significance: Important in biotechnology for gene cloning and genetic engineering.
Slide 6: Mechanisms of Extrachromosomal Inheritance
Non-Mendelian Patterns: Do not follow Mendel’s laws of inheritance.
Cytoplasmic Segregation: During cell division, organelles like mitochondria and chloroplasts are randomly distributed to daughter cells.
Heteroplasmy: Presence of more than one type of organellar genome within a cell, leading to variation in expression.
Slide 7: Examples of Extrachromosomal Inheritance
Four O’clock Plant (Mirabilis jalapa): Shows variegated leaves due to different cpDNA in leaf cells.
Petite Mutants in Yeast: Result from mutations in mitochondrial DNA affecting respiration.
Slide 8: Importance of Extrachromosomal Inheritance
Evolution: Provides insight into the evolution of eukaryotic cells.
Medicine: Understanding mitochondrial inheritance helps in diagnosing and treating mitochondrial diseases.
Agriculture: Chloroplast inheritance can be used in plant breeding and genetic modification.
Slide 9: Recent Research and Advances
Gene Editing: Techniques like CRISPR-Cas9 are being used to edit mitochondrial and chloroplast DNA.
Therapies: Development of mitochondrial replacement therapy (MRT) for preventing mitochondrial diseases.
Slide 10: Conclusion
Summary: Extrachromosomal inheritance involves the transmission of genetic material outside the nucleus and plays a crucial role in genetics, medicine, and biotechnology.
Future Directions: Continued research and technological advancements hold promise for new treatments and applications.
Slide 11: Questions and Discussion
Invite Audience: Open the floor for any questions or further discussion on the topic.
Comparing Evolved Extractive Text Summary Scores of Bidirectional Encoder Rep...University of Maribor
Slides from:
11th International Conference on Electrical, Electronics and Computer Engineering (IcETRAN), Niš, 3-6 June 2024
Track: Artificial Intelligence
https://www.etran.rs/2024/en/home-english/
Introduction:
RNA interference (RNAi) or Post-Transcriptional Gene Silencing (PTGS) is an important biological process for modulating eukaryotic gene expression.
It is highly conserved process of posttranscriptional gene silencing by which double stranded RNA (dsRNA) causes sequence-specific degradation of mRNA sequences.
dsRNA-induced gene silencing (RNAi) is reported in a wide range of eukaryotes ranging from worms, insects, mammals and plants.
This process mediates resistance to both endogenous parasitic and exogenous pathogenic nucleic acids, and regulates the expression of protein-coding genes.
What are small ncRNAs?
micro RNA (miRNA)
short interfering RNA (siRNA)
Properties of small non-coding RNA:
Involved in silencing mRNA transcripts.
Called “small” because they are usually only about 21-24 nucleotides long.
Synthesized by first cutting up longer precursor sequences (like the 61nt one that Lee discovered).
Silence an mRNA by base pairing with some sequence on the mRNA.
Discovery of siRNA?
The first small RNA:
In 1993 Rosalind Lee (Victor Ambros lab) was studying a non- coding gene in C. elegans, lin-4, that was involved in silencing of another gene, lin-14, at the appropriate time in the
development of the worm C. elegans.
Two small transcripts of lin-4 (22nt and 61nt) were found to be complementary to a sequence in the 3' UTR of lin-14.
Because lin-4 encoded no protein, she deduced that it must be these transcripts that are causing the silencing by RNA-RNA interactions.
Types of RNAi ( non coding RNA)
MiRNA
Length (23-25 nt)
Trans acting
Binds with target MRNA in mismatch
Translation inhibition
Si RNA
Length 21 nt.
Cis acting
Bind with target Mrna in perfect complementary sequence
Piwi-RNA
Length ; 25 to 36 nt.
Expressed in Germ Cells
Regulates trnasposomes activity
MECHANISM OF RNAI:
First the double-stranded RNA teams up with a protein complex named Dicer, which cuts the long RNA into short pieces.
Then another protein complex called RISC (RNA-induced silencing complex) discards one of the two RNA strands.
The RISC-docked, single-stranded RNA then pairs with the homologous mRNA and destroys it.
THE RISC COMPLEX:
RISC is large(>500kD) RNA multi- protein Binding complex which triggers MRNA degradation in response to MRNA
Unwinding of double stranded Si RNA by ATP independent Helicase
Active component of RISC is Ago proteins( ENDONUCLEASE) which cleave target MRNA.
DICER: endonuclease (RNase Family III)
Argonaute: Central Component of the RNA-Induced Silencing Complex (RISC)
One strand of the dsRNA produced by Dicer is retained in the RISC complex in association with Argonaute
ARGONAUTE PROTEIN :
1.PAZ(PIWI/Argonaute/ Zwille)- Recognition of target MRNA
2.PIWI (p-element induced wimpy Testis)- breaks Phosphodiester bond of mRNA.)RNAse H activity.
MiRNA:
The Double-stranded RNAs are naturally produced in eukaryotic cells during development, and they have a key role in regulating gene expression .
Richard's aventures in two entangled wonderlandsRichard Gill
Since the loophole-free Bell experiments of 2020 and the Nobel prizes in physics of 2022, critics of Bell's work have retreated to the fortress of super-determinism. Now, super-determinism is a derogatory word - it just means "determinism". Palmer, Hance and Hossenfelder argue that quantum mechanics and determinism are not incompatible, using a sophisticated mathematical construction based on a subtle thinning of allowed states and measurements in quantum mechanics, such that what is left appears to make Bell's argument fail, without altering the empirical predictions of quantum mechanics. I think however that it is a smoke screen, and the slogan "lost in math" comes to my mind. I will discuss some other recent disproofs of Bell's theorem using the language of causality based on causal graphs. Causal thinking is also central to law and justice. I will mention surprising connections to my work on serial killer nurse cases, in particular the Dutch case of Lucia de Berk and the current UK case of Lucy Letby.
This pdf is about the Schizophrenia.
For more details visit on YouTube; @SELF-EXPLANATORY;
https://www.youtube.com/channel/UCAiarMZDNhe1A3Rnpr_WkzA/videos
Thanks...!
Deep Behavioral Phenotyping in Systems Neuroscience for Functional Atlasing a...Ana Luísa Pinho
Functional Magnetic Resonance Imaging (fMRI) provides means to characterize brain activations in response to behavior. However, cognitive neuroscience has been limited to group-level effects referring to the performance of specific tasks. To obtain the functional profile of elementary cognitive mechanisms, the combination of brain responses to many tasks is required. Yet, to date, both structural atlases and parcellation-based activations do not fully account for cognitive function and still present several limitations. Further, they do not adapt overall to individual characteristics. In this talk, I will give an account of deep-behavioral phenotyping strategies, namely data-driven methods in large task-fMRI datasets, to optimize functional brain-data collection and improve inference of effects-of-interest related to mental processes. Key to this approach is the employment of fast multi-functional paradigms rich on features that can be well parametrized and, consequently, facilitate the creation of psycho-physiological constructs to be modelled with imaging data. Particular emphasis will be given to music stimuli when studying high-order cognitive mechanisms, due to their ecological nature and quality to enable complex behavior compounded by discrete entities. I will also discuss how deep-behavioral phenotyping and individualized models applied to neuroimaging data can better account for the subject-specific organization of domain-general cognitive systems in the human brain. Finally, the accumulation of functional brain signatures brings the possibility to clarify relationships among tasks and create a univocal link between brain systems and mental functions through: (1) the development of ontologies proposing an organization of cognitive processes; and (2) brain-network taxonomies describing functional specialization. To this end, tools to improve commensurability in cognitive science are necessary, such as public repositories, ontology-based platforms and automated meta-analysis tools. I will thus discuss some brain-atlasing resources currently under development, and their applicability in cognitive as well as clinical neuroscience.
Lateral Ventricles.pdf very easy good diagrams comprehensive
Hybridoma technology
1. MONOCLONAL antibody (hybridoma technology)
Archana Soni
Assistant Professor
Shri Shankaracharya
Mahavidyalaya, Junwani,
Bhilai (C. G.) INDIA
2. Monoclonal Antibodies
Contents
1. Definition.
2. Introduction.
3. Pharmacology.
4. Effects of MAbs.
5. Principle for creation of MAbs.
6. Production of MAbs.
7. Human monoclonal antibodies.
8. Advantages of MAbs.
9. Limitations of MAbs.
10. Applications of MAbs.
11. Antibody fingerprinting.
12. Conclusion.
13. Reference.
3. Definition
MAbs are antibodies that are identical
because they were produced by one of
the immune cell (B cell), all clones of a
single parent cell.
Given any substance, it is possible to
create monoclonal antibodies that
specifically bind to that substance; they
can then serve to detect or purify that
substance. This has become an
important tool in biochemistry,
4. Introduction
• Antibodies or immunoglobulins are protein molecules produced by a specialized group of cells called
B-lymphocytes (plasma cells) in mammals.
• Monoclonal antibody (MAb) is single type of antibody that is directed against a specific
antigenic determinant (epitope). In the early years, animals were immunized against a specific antigen,
B-lymphocytes were isolated and cultured in vitro for producing MAbs. This approach was not
successful since culturing normal B-lymphocytes is difficult, and the synthesis of MAb was short-lived
and very limited.
• It is interesting that immortal monoclonal antibody producing cells do exists in
nature. They are found in the patients suffering from a disease called multiple myeloma (a cancer of B-
lymphocytes). It was in 1975. George Kohler and Cesar Milstein (Noble Prize, 1984) achieved large
scale production of MAbs. They could successfully hybridize antibody—production B-lymphocytes
with myeloma cells in vitro and create a hybridoma.
• The production of monoclonal antibodies by the
hybrid cells is referred to as hybridoma technology.
5. Pharmacology
• A) Mechanismof actionof MAbs:
• Blocking or steric hindrance of the function of target antigen i.e., T-lymphocytes, B-
lymphocytes, tumor necrosis factor-a (TNFa) and interleukin (IL) which are capable
of transducing intracellular signals.
• Cytotoxicity to the cell expressing target AG by ADCC or CDC.
• Inhibition of growth .
• B) Pharmacokinetics:
• MAbs are used by intravascular route and remain essentially intravascular.
Intravenous injection is not always be appropriate for long-term treatment for a
variety of reasons. Hour-long infusion require a hospital environment and are often
associated with mind to very severe side effects. Continous and sustained delivery
of antibodies can lead to induction of neutralizing anti-idiotypic immune
responses, which sometimes develop when massive doses of purified
immunoglobulins are repeatedly injected into patients. Additionally, the
bioavailability of therapeutic antibodies is often detrimental to the treatment
efficacy. They have small volume of distribution and limited tissue penetration.
They remain in circulation for 2 days to 2 weeks. Another limitation is the high cost
of recombinant proteins certified for human use.
7. PRINCIPLE FOR CREATION OF HYBRIDOMA
CELLS
• The myeloma cells used in hybridoma technology must not be capable of
synthesizing their own antibodies. The selection of hybridoma cells is based on
inhibiting the nucleotide synthesizing machinery. The mammalian cells can
synthesize nucleotides by two pathway—de novo synthesis and salvage pathway.
• The de novo synthesis of nucleotides
require tetrahydrofolate which is formed from dihydrofolate. The formation of
tetrahydrofolate can be blocked by the inhibitor aminopterin.
• The salvage pathway involves
the direct conversion of purines & pyrimidines into the corresponding nucleotides.
Hypoxanthine guanine phosphoribosyl transferase (HGPRT) is a key enzyme in the
salvage pathway of purines. Thymidine kinase (TK), involved in the salvage
pathway of pyrimidines converts thymidine monophosphate (TMP). Any mutation
in either one of the enzymes blocks the salvage pathway.
• When cells deficient in HGPRT are
grown in a medium containing hypoxanthine aminopterin & thymidine (HAT
medium), they cannot survive due to inhibition of de novo synthesis of purine
nucleotiodes. (salvage pathway is not operative due to lack of HGPRT). Thus, cells
lacking HGPRT, grown in HAT medium die.
• The hybridoma cells, possess the ability of myeloma cells
to grow in vitro with a functional HGPRT gene obtained from lymphocytes. Thus,
only the hybridoma cells can proliferate in HAT medium, & this procedure is
successfully used for their selection.
8.
9. Production of Mab’s
• The establishment of hybridomas
and production of MAbs involves
the following steps.
• 1.) Immunization.
• 2.) Cell fusion
• 3.) Selection of hybridomas.
• 4.) Screening the products.
• 5.) Cloning & propagation.
• 6.) Characterization & storage.
10. Human monoclonal
antibodies
• From SCID mouse by injecting human B and T
cells and grafting mouse by transplanting
spleens and lymph nodes.
• The transplanted mouse is immunized with
target antigens to produce human antibodies.
• Phage derived combinatorial antibody library,
phage display technique-mimics in vitro
affinity maturation.
11. ADVANTAGES OF MAB’S
• 1.) Homogeneity: Monoclonal antibody represents a single antibody molecule that
binds to antigen with the same affinity and promote the same effectors functions.
• 2.) Specificity: The product of a single hybridoma reacts with the same epitope on
antigens.
• 3.) Immunizing Antigen: Need not be characterized and is ultimately not needed in
large quantities to produce large quantities of antibody.
• 4.) Selection: It is possible to select for specific epitope specificities and generate
antibodies against a wider range of antigenic determinants.
• 5.) Antibody Production: Unlimited quantities of a single well-defined monospecific
reagent.
12. Limitations of MAb’s
• As they are specific to a particular antigen, they cannot distinguish
molecule as a whole.
• Some times they cannot distinguish groups of different molecules. Ex:
Presence of retro viruses as a part of mammalian chromosomes is not
distinguished.
• The presence of some of these viruses is detected in hybridomas. This poses
a great danger since there is no guarantee for MAb produced is totally
virus free.
• For this reason US food and drug administration insists that MAb for
human use should be totally free all pathogenic organism including
viruses.
13. APPLICATIONOF MAB’S
• 1.) Diagnostic applications.
• a) MAbs in biochemical analysis
• b) MAbs in diagnostic imaging
• 2.) Therapeutic uses
• a) MAbs as direct therapeutic agents
• b) MAbs as targeting agents in therapy
• 3.) Protien purification
• 4.) Miscellaneous applications
(ABZYMES).
14. ANTIBODYFINGERPRINTING
• The occurrence of antibodies and their
involvement in certain diseases is well known
(e.g., rheumatic arthritis). A new category
of individual specific (IS) autoantibodies
have been discovered in recent years.
These IS--autoantibodies are produced
after birth and reach maximum in number
by 2 years, and the remain constant for
the later part of life. Monoclonal
antibodies produced against IS--
autoantibodies can be used for their
detection, and identification of individuals.
This technique referred to as autoantibody
15. CONCLUSION
• Since for human health care, the subject of production and use
of antibodies has become a very important area of research, not
only for academic purposes but also for its relevance to
industrial growthfor diagnosis and therapeutics, monoclonal
antibodies can be utilized for it. In near future more such
monoclonal antibodies shouldbe produced for biochemical
research that will be of great commercial and medical value.