MICROSPHERES/
MICROCAPSULES
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P H A R M A C Y
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CONTENTS
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▪ INTRODUCTION
▪ POLYMERS USED
▪ ADVANTAGES
▪ METHODS OF PREPARATION
▪ FACTORS AFFECTING THE RELEASE OF THE
DRUG
▪ APPLICATIONS
▪ MONOCLONAL ANTIBODY PRODUCTION
▪ APPLICATIONS
▪ REFERENCES
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INTRODUCTION
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▪ Microencapsulation of pharmaceuticals was first
investigated in the year 1931 by preparing spheres of
gelatin using coacervation technique.
▪ The micro particulate delivery systems are considered
and accepted as a reliable means to deliver the drug to
the target site with specificity, if modified, and to
maintain the desired concentration at the site of
interest without untoward effect.
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Microspheres
▪ Small spherical particles.
▪ Solid matrix particle.
▪ It is a micrometric
reservoir system.
▪ Diameter ranges from 1µ
to 1000µ.
Fig 1:Microspheres
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Microcapsules
▪ Microcapsules can be the
small entities that
contain an active agent
or core material
surrounded by a shell or
embedded into a matrix
structure.
▪ It is a micrometric
matrix system.
Fig 2 : Microcapsules
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ADVANTAGES
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▪ Sustained or prolonged release of the drug. Eg
Glibenclamide.
▪ To masking the organoleptic properties.Eg. Paracetamol,
Nitrofurantoine.
▪ Liquid drugs can be converted in a free flowing powder.
▪ The drugs sensitive to moisture, light and oxygen can be
protected by this technique. Eg.Nifedipine-Photo
instability.
▪ Prevent the incompatibility between drugs.
Eg.Hydroquinone
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▪ Drug volatilation can be prevented by this technique.
Eg. Aspirin.
▪ Reduction in toxicity and GI irritation including with
KCL and ferrous sulphate can be achieved by
microencapsulation.
▪ Enhanced stability, it prevent from oxidation.
Eg.Vitamin A palmitate.
▪ Used to prepare intrauterine contraceptive device.
▪ Change the site of application has been useful for
those drugs which have the toxicity at lower pH.
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DISADVANTAGES
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▪ The costs of the materials higher than those of standard
formulations.
▪ The fate of polymer matrix and its effect on the
environment.
▪ The fate of polymer additives such as plasticizers,
stabilizers, antioxidants and fillers.
▪ Reproducibility is less.
▪ Process conditions like change in temperature,pH,
solvent addition, and evaporation/agitation may
influence the stability of core particles to be
encapsulated.
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Mechanism and kinetics of drug release
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Diffusion
Diffusion is the most commonly involved mechanism
wherein the dissolution fluid penetrates the shell, dissolves
the core and leak out through the interstitial channels.
Thus, the overall release depends on,
▪ The rate at which dissolution fluid penetrates the wall
of microcapsules.
▪ The rate at which drug dissolves in the dissolution
fluid.
▪ The rate at which the dissolved drug leak out and
disperse from the surface.
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Dissolution
▪ Dissolution rate of polymer coat determines the
release rate of drug from the microcapsule.
Osmosis
▪ The polymer coat of microcapsule acts as semi
permeable membrane and allows the creation of an
osmotic pressure.
Erosion
▪ Erosion of coat due to pH and/or enzymatic
hydrolysis causes drug release.
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Prerequisites For Ideal Microparticulate Carriers
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▪ Longer duration of action
▪ Increase of therapeutic efficiency
▪ Control of content release
▪ Protection of drugs
▪ Reduction of toxicity
▪ Biocompatibility
▪ Relative stability
▪ Water solubility or dispersability
▪ Bioresorbability
▪ Targetability
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POLYMERS USED
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SYNTHETIC POLYMERS
Non-biodegradable
▪ Acrolein
▪ Epoxy polymers
Biodegradable
▪ Polyanhydrides
▪ Polyalkyl cyano acrylates
▪ Lactides and glycolides and their copolymers
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Cntd..
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NATURAL MATERIALS
Proteins
▪ Albumins,Gelatin,Collagen
Carbohydrates
▪ Starch,Agarose,Chitosan
Chemically modified Carbohydrates
▪ Poly(acryl)dextran
▪ Poly(acryl)starch
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Examples
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▪ PLGA (poly lactic-co-glycolic acid) microsphere
▪ Gelatin microspheres
▪ Dextran microspheres
▪ Polyanhydride microspheres
▪ Poly phosphazene microspheres
▪ Chitosan microspheres
▪ Poly saccharides or lipid cross linked chitosan
microspheres
▪ Poly alkyl cyanoacrylate microsphere
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▪ Drug loaded albumin microsphere
▪ Poly acrolein microsphere
▪ Hybrido microsphere
▪ Polymer grafted starch microsphere
▪ Starch microsphere
▪ Carrageenan microsphere
▪ Alginate microsphere
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Materials used for preparation
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Core material
▪ The core material, defined as the specific material to
be coated, can be liquid or solid in nature.
▪ The solid core be active constituents, stabilizers,
diluents, excipients, and release-rate retardants or
accelerators.
▪ Liquid Core Material- Solvents, catalyst,sugars,salts.
▪ Solid Core Material- Dextrins, minerals, bases,
pharmaceuticals.
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Coating material
▪ Innovative coating polymers have also been
developed for some special applications particularly
among the bio adhesives and mucoadhesives. Eg.
Ethyl cellulose, carboxylate and amino derivatives.
Water Soluble resins
▪ Hydroxyethylcellulose, Polyvinylpyrrolidine,starch.
Water insoluble resins
▪ EthylCellulose,Polyethylene,Polymethacrylate.
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METHODS OF PREPARATION:
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1) Single Emulsion Technique
2) Double Emulsion Technique
3) Polymerization Technique
4) Phase Separation Coacervation
5) Spray drying
6) Solvent Extraction
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Factors to be considered
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▪ Particle size requirement
▪ The drug and protein should not be adversely affected
by the process.
▪ Reproducibility of release profile and method
▪ No stability problem
▪ No toxic product with the final product.
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1) Single Emulsion Technique
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Aqueous solution /suspension of polymer
Stirring / sonication
Dispersion in organic phase oil/chloroform
Cross linking
Heat denaturation Chemical cross linking
By adding dispersion to heated oil ( Glutaraldehyde,HCHO,
CHCl3 )
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Microspheres in organic phase
Centrifugation,washing,separation
MICROSPHERES
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2) Double Emulsion Technique
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Aqueous solution of polymer
Dispersion in oil/organic phase, vigorous
homogenization(sonication )
Primary emulsion(w/o)
Addition of aqueous solution of PVA
W/O/W multiple emulsion
Addition to large Aqueous phase
denaturation /hardening
Microspheres in solution
Separation, washing, Drying
Microspheres
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3)Polymerization technique
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Types
▪ Bulk polymerization
▪ Suspension precipitation polymerization
▪ Emulsion polymerization
▪ Miceller polymerization
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Bulk polymerization:
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Monomer Bioactive material Initiator
Heated to initiate polymerization
Initiator accelerate rate of
Reaction
Polymer(Block)
Moulded/fragmented
MICROSPHERES
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Suspension polymerization
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Monomer Bioactive material Initiator
Dispersion in water and stabilizer
Droplets
Polymerization Vigorous agitation
Heat/irradiation
Separation and Drying
MICROSPHERES
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Emulsion Polymerization
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Monomer/ Aq.Solution of NaOH,
Bioactive material Initiator, Surfactant , Stabilizer
Dispersion with vigorous stirring
Micellar sol. of polymer in aqueous medium
Polymerization
Microspheres formation
MICROSPHERES
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4)Phase Separation Coacervation
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Aq./organic solution of polymer
Drug
Drug dispersed or dissolved in the polymer solution
Phase separation by different means
Polymer rich in globules
Hardening
Microspheres in aq./organic phase
Separation Drying
MICROSPHERES
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5)Spray Drying
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Polymer dissolve in volatile organic solvent
(Acetone , Dichloromethane)
C
Drug dispersed in polymer solution under high speed
homogenization
Atomized in a stream of hot air
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Due to solvent evaporation small droplet or fine mist
form
Leads to formation of Microspheres
Microspheres separated from hot air by cyclone
separator
Trace of solvent are removed by vacuum drying
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6)Solvent extraction
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Drug is dispersed in organic solvent
Organic phase is removed by extraction with water
(This process decreasing hardening time for
microspheres)
Hardened microspheres
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FACTORS AFFECTING RELEASE OF THE
DRUG
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Drug
▪ Position in microspheres
▪ Molecular weight
▪ Physiochemical properties
▪ Concentration
▪ Interaction with matrix
Environment
▪ pH
▪ Polarity
▪ Presence of enzyme
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Microspheres
▪ Type and amount of matrix polymer
▪ Size and density of the microspheres
▪ Extent of cross linking ,denaturation or polymerization
▪ Adjuvants
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Characterization of Microspheres
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▪ Characterization is an important phenomenon to
determine the micro structure of the microspheres
▪ Used to determine the Release and stability of the
carrier.
Particle size and shape
Determination of shape and structure of micro particles.
▪ Light microscopy
▪ Laser light scattering microscopy
▪ Scanning electron microscopy
▪ Confocal laser scanning microscopy
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Electron microscopy for chemical analysis
▪ Used to determine the atomic composition of the
surface .
▪ ESCA can be used to determine the surfacial
degradation of the biodegradable microspheres.
Attenuated Total Reflectance Fourier Transform Infrared
Microscopy
▪ FTIR – Degradation of polymer matrix of the system
▪ ATR -Surface of the microsphere
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Angle of contact
▪ Measure the wetting property of microparticulate
carrier.
▪ It is measured at the solid/air/water interface.
Density determination
▪ Density measured by using a multivolume
pychnometer.
IR spectoscopy
▪ To measure deviation in composition.
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Isoelectric point
▪ Micro electrophoresis apparatus is used to measure
electrophoretic mobility of microspheres from which
isoelectric point can be determine.
▪ It can be correlated to surface charge or ion
adsorption of microspheres.
Surface carboxylic acid residue
▪ Measured by using radioactive glycine.
▪ Scintillation counter.
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Surface amino acid residue
▪ Determined by using 14 C- acetic acid conjugate.
▪ Liquid Scintillation counter.
Capture Efficiency
▪ The percent entrapment can be determined by allowing
washed microspheres to lyse.
Actual content
% Entrapment = × 100
Theoretical content
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Release study
▪ Usually carried out in phosphate saline buffer pH 7.4.
▪ Two method
1) Rotating paddle dissolution Apparatus
2) Dialysis method
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DIALYSIS METHOD
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▪ Used to study the release of drug or proteins from
the microsphere.
▪ Microsphere are kept in the dialysis bag or tube with
membrane while dialysis media continuously stirred
and sample of dialysate are taken.
▪ The withdrawn sample is estimated for drug content
and each time volume is replaced using fresh buffer
solution.
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Fig 3 : Dialysis assembly
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Rotating Paddle Dissolution Apparatus
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▪ Sample taken at regular time interval.
▪ Sink condition maintained.
▪ Analyzed as per monograph.
▪ Release profile from graph of Amount Release vs
Time.
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Phosphate saline buffer
Ph= 7.4
Drug loaded microsphere
Paddle rotated at 100 rpm
Fig 4 : Rotating paddle
dissolution apparatus
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Application of microspheres in
pharmaceutical industry
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▪ For taste and odour masking
▪ To delay the volatilization
▪ For separation of incompatible substances
▪ For improvement of flow properties of powders
▪ To increase the stability of the drug against the
external conditions
▪ For safe handling of toxic substances
▪ To improve the solubility of water insoluble
▪ Substances by incorporating dispersion of such in
aqueous media .
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▪ For targeting delivery of Anti-neoplastic drug.
▪ In vaccine delivery.
▪ In cosmetic industry.
▪ For delivery of several analgesic drug like Naproxen
sodium.
▪ For delivery of Rifampicin in TB.
▪ Increase retention time of drug in ocular system like
Pilocarpine nitrate
▪ For delivery of certain gastric acid labile antibiotics
like Penicillin.
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Targeting using Micro particulate carriers
▪ Targeting means “The therapeutic efficacy of the drug
relies on its access and specific interaction with its
candidate receptors.”
Ocular
▪ The eye and the cornea are easily accessible targets.
▪ The retention of micro particulate system can be
attained by using gel form.
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Nasal
▪ The intranasal route is exploited for the delivery of
the peptides and proteins.
▪ Bio adhesive microspheres have greater control over
the surface character and the release pattern.
Oral
▪ The most preferred convenient route.
▪ It also preferred for delivery of soluble antigen.
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Magnetic microspheres
▪ It is a biophysical approach.
▪ Magnetic microspheres are prepared by mixing
water soluble drugs and 10 nm magnetite in aqueous
solvent of matrix material.
▪ Magnetic targeting is based on the force exerted by
external magnetic field over the magnetically
susceptible microspheres.
▪ Eg. Amphotericin B ,Interleukin 2.
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Microspheres in vaccine drug delivery
▪ Biodegradable delivery system of vaccine can be
given by parenteral route.
▪ Polymers used- Poly lactic acid, poly glycolic acid,
poly lactides coglycosides. Eg. Diphtheria toxoid.
Hepatitis B surface antigen.
Specific advantages:
▪ Improved antigenicity by adjuvant action
▪ Modulation of antigen release
▪ Stabilization of antigen
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Microspheres in immune system
▪ Microspheres interact with macrophages release the
antigens which are phagocytosized by antigen
presenting cells are responsible for activation of B
and T cells.
▪ Eg. Bovine serum albumin, Tetanus toxoid.
Microsponges
▪ It consist of non collapsible structures with porous
surface through which active ingredients are released
in a controlled manner.
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Imaging
▪ Cells, cell lines , tissues, organs can be imaged using
radiolabelled microspheres.
▪ Eg. The scintiographic imaging of the tumor masses
in lungs using labelled human serum albumin
microsphere.
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Recent advances in microencapsulation
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▪ An injectable slow-release partial opioid agonist or opioid
antagonist in a poly (D, L-lactide) microspheres with a
small amount of residual ethyl acetate.
▪ Enteric polymeric micro particles containing a
proteinaceous antigen in a single or double emulsification
process in which the enteric polymer acts as a stabilizer
for the micro particles which are formed in the process.
▪ Sustained release microsphere containing a LHRH
derivative or its salt in a large amount without containing
gelatin by using a lactic acid-glycolic acid polymer or
salts.
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▪ Double wall microspheres using two biodegradable
polymers by the o/w emulsification solvent extraction
process.
▪ Method of encapsulating DNA retaining its ability to
induce expression of its coding sequence in a micro
particle for oral administration prepared using the w/o/w
emulsion and using biodegradable polymers.
▪ Starch to encapsulate vaccines using emulsification
method
▪ Encapsulation of nucleotides and growth hormone using
simple or double emulsification methods.
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MONOCLONAL
ANTIBODY
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Monoclonal Antibodies
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▪ An antibody is a protein used by the immune system
to identify and neutralize foreign objects like bacteria
and viruses. Each antibody recognizes a specific
antigen unique to its target.
▪ Monoclonal antibodies (mAb) are antibodies that
are identical because they were produced by one type
of immune cell, all clones of a single parent cell.
▪ Polyclonal antibodies are antibodies that are derived
from different cell lines. They differ in amino acid
sequence.
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Advantages
▪ When drugs are delivered as antibody conjugates the
conjugates can specifically reach the target cells
without causing any damage to the normal tissue
Disadvantages
▪ Monoclonal antibody production, a time consuming
process.
▪ Average affinity of Monoclonal antibodies are
generally lower.
▪ Any physical/chemical treatment will affect all
Monoclonal antibodies in that production.
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PRODUCTION OF MONOCLONAL
ANTIBODY
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HYBRIDOMA TECHNOLOGY
Step 1: – Immunization Of Mice & Selection Of
Mouse Donor For Generation Of Hybridoma cells
ANTIGEN ( Intact cell/
Whole cell membrane/
micro-organisms ) +
ADJUVANT
(emulsification) Ab titre reached in Serum
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Step 2: – Screening Of Mice For Antibody Production
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After several
weeks of
immunization
Serum Antibody Titre Determined
(Technique: – ELISA / Flow cytometery)
Titre too low Titre High
BOOST BOOST
(Pure antigen) (Pure antigen)
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Step 3: – Preparation of Myeloma Cells
+ HAT(Hypoxanthine
Myeloma Cells Aminopetrin
Immortal Tumor Of Lymphocytes Thymidine )Medium
Myeloma Cells
HGPRT-
High Viability & Rapid Growth
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Step 4: – Fusion of Myeloma Cells with Immune Spleen Cells
Selection of Hybridoma Cells
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PEG
FUSION
SPLEEN CELLS MYELOMA CELLS
Feeder Cells
Growth Medium
HYBRIDOMA CELLS
ELISA PLATE
HAT Medium
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Step 5: – Cloning of Hybridoma Cell Lines by “ Limiting
Dilution” or Expansion
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A. Clone each +ve Culture
B. Test each supernatant for antibodies
C. Expand +ve Clones
Propagate
Invitro Invivo
Tissue Mouse Ascites method
Culture
Method
HARVEST MONOCLONAL ANTIBODY
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Purification techniques
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▪ Cells, cell debris, lipids, and clotted material are first
removed, typically by filtration with a 0.45 um filter.
▪ Chromatography
(I) Ion-exchange chromatography
(II) Antigen affinity chromatography
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Types of mAbs
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▪ Murin source mAbs: Rodent mAbs with excellent affinities
and specificities, generated using conventional hybridoma
technology. Clinical efficacy compromised by HAMA
(human anti murine antibody) response, which lead to
allergic or immune complex hypersensitivities.
▪ Chimeric mAbs: Chimers combine the human constant
regions with the intact rodent variable regions. Affinity and
specificity unchanged. Also cause human antichimeric
antibody response (30% murine resource)
▪ Humanized mAbs: Contained only the CDRs of the rodent
variable region grafted onto human Framework Regions.
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Applications of Monoclonal Antibodies
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Immunodiagnostic Applications
▪ Detects protein of interest either by blotting or
immunofluorescence
▪ Enzyme linked immunosorbant assay
Therapeutic Applications
▪ Transplant rejection
▪ Cancer
▪ Autoimmune disorders
▪ Inflammatory disease
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Immunodiagnosis of pregnancy
▪ Onset of pregnancy can be determined monoclonal
anti-ß hCG antibody was labeled with the enzyme,
horseradish peroxidase.
Immunodiagnosis of viral, bacterial and parasitic infections
▪ Enzyme immunoassay have been developed for
bacterial, viral, parasitic infections.
Blood group typing
▪ Monoclonal antibodies used as a antisera in blood
grouping.
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Analysis of antigenic determinants
▪ Antigenic fingerprinting with monoclonal antibody used
in the development of immunodiagnostic tests and in
the field of vaccine development.
Monoclonal antibodies in cancer therapy
▪ MCA have been used in patients suffering from
leukemia’s, lymphomas, melanomas, colorectal cancer.
▪ Eg. Arcitumomab – Anti-carcinoembryonic antigen
(CEA) antibody labelled with technetium 99 (99Tc)
▪ Used for imaging patients with colorectal carcinoma.
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▪ Nofetumuma – Mice mab coupled with 99Tc for
diagnosis to determine extent and stage of disease of
small cell lung cancer.
Anti-inflammatory and immunosuppressant mabs
▪ Muromonab (OKT-3)-Blockage suppresses activity of T
cells.
▪ Adalimumab, Etanercept and infliximab- Suppression of
release of inflammatory cytokines IL-1,IL-6.
▪ Omalizumab – Asthma.
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Investigation of receptor ligand interaction
Used to study the receptors structure and function.
It includes
▪ Affinity purification of receptors
▪ Biochemical characterization
▪ Function
▪ Location
▪ Identification of receptor –ligand internationalization
and recycling pathways.
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Vehicle
▪ Monoclonal antibodies as a vehicle for delivery of
drugs.
▪ An antibody can be used for homing of attached drug
or toxin at cancer cells.
By this higher concentration can buildup locally,
minimizing the systemic toxicity.
1. Danuamycin loaded MCA – lymphomas
2. Methotrexate
3. Adiramycin
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References
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1. Gupta AK., Dey BK., Microencapsulation For
Controlled Drug Delivery ,Sunsari Technical
College Journal, Volume 1, Issue 1, October 2012 .
2. Jain N.K.,controlled and Novel Drug Delivery, CBS
Publishers and Distributors ,New Delhi,First edition
1997 Reprint in (2001).Page no 219-250.
3. Vyas S.P. and Khar R.K., Targeted and controlled
drug delivery system, Vallabhprakashan, New Delhi,
First edition. Page no 418-454.
4. R Gupta et al., Monoclonal antibodies and their
production and applications.
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THANK YOU
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