tissue culture technology,242013

7/28/2019 Tissue Culture Technology,242013

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ANIMALTISSUE CULTURE


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Topics

Introduction1

Tissue Culture2

Hybridoma Tech3

Future Medicine4


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Biotechnology: Present and Future


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IMMUNITY


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Figure 1-7Anatomi

sistemimun


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CONTROL OF VIRUS INFECTION


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T Helper (Th) Cells in Immune Responses

Th cell

Cellular

Immunity

Humoral

Immunity

Th cells-Play a central role in immune protection.

-Orchestrate immune responses through

their production of cytokines and

chemokines.


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Th cell subsets

Th1Th17

TregTh2 Th cell


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Diseases with Imbalance of

Th1/Th2 Response

Th1 > > Th2 Th1 < < Th2

VS

Autoimmune diseases

Multiple sclerosis

DiabetesRheumatis arthritis

Viruses

HIV, MAIDS

HSV-1

BacteriaLister ia

monocytogenesOrgan transplantation

Allergic diseases

Allergy

AsthmaAllergic rhinitis

ProtozoansLeishmania major

Tryanosoma cruzi

BacteriaMycobacter ium

tuberculosisFungiCandida


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Figure A-35Cel l St imu lat ing Agen ts


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Future Medicine


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Epidemiology

Surveillance &

Disease Investigation

Biostatistics

Environmental

Science

Community Health

Promotion

Infectious Disease

Competencies


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Morens, D., Folkers, G., & Fauci, A.(2004) The challenge of emerging and re-emerging infectiousdiseases. Nature, 430: 242-249.


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Global distribution of Relative Risk for an EID Event

caused by Zoonotic Pathogens from Wildlife*

Jones KE, Patel NG, Levy MA et al. Nature 2008; 451:990-993.


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Inhibition of IL-12 Production as a

Therapeutic Strategy of Autoimmune diseases

To screen active compounds with inhibitory

activity on IL-12 production1. Retinoids, oxLDL

2. Chemicals from some medicinal plants

3. Combinatorially synthesized chemicals

Major IL-12 producers

Macrophage Dendritic cell


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Tissue Culture: Teory, Techniques,and Application

D i d


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Design dan Layout

Laboratorium

Service Bench

Preparation room

Wash up (Tempat pencucian)

Storage

Construction and layoutEssential Equipment


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Sterile Handling Area

Laminar flow

Incubation

Heaters

Air circulation

Thermostats


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PENANGANAN KONTAMINASI

Sumber infeksi

1. Sel

2. Media kultur

3. Glassware dan Plastic ware

4. Airborne modes

Laminar flow TransferRoom


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Biologi cell culture

Kuliah ke 3


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Petunjuk praktis kultur sel

Ruang kultur dan lingkungannya

Sterilitas dan peralatan

Kualitas bahan untuk kulturPemilihan Media

Pemilihan Serum

Material untuk kultur sel


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Biologi Sel Kultur

Lingkungan kultur: In vivo x in vitro

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Transformation

Cell death

Log cell


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Istilah

Dedifferentiation:

Undefferentiated cell

Absence of appropriate inducer

Penting untuk di pahami: dedifferentiation,deadaptation, differentiation and selection

CLONE SELECTION THEORY


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CLONE SELECTION THEORY


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Sumber infeksi

1. Sel

2. Media kultur

3. Glassware dan Plastic ware

4. Airborne modes

Laminar flow TransferRoom


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Teknik Aseptik

Antibiotika

Prosedur di Ruang Transfer

P d di R T f


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Prosedur di Ruang Transfer

Personel Ruang kultur

Bahan yang dipakai harus di tes

sebelumnya (media, serum, trypsin,

HBSS dan komponen lainnya dari

media)

KulturSkills


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TC APPLICATION



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Th cell

Cellular

Immunity

Humoral

Immunity

Th cells-Play a central role in immune protection.

-Orchestrate immune responses through

their production of cytokines and

chemokines.


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Teknik Hibridoma

o Isolasi Limfosit

o Isolasi Makrofag

o Kultivasi Myeloma

o Fusi Sel

o Seleksi Hibrid


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HAT-SELECTION

B-cells Non fused B-cells

Non fused myeloma

Fusion

False hybrid

Myeloma cells True Hybrid


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Hybridoma clones and normal feeder cells

THE STEP THAT YOU SHOULD FREEZE


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THE STEP THAT YOU SHOULD FREEZE

STOCK IN HYBRIDOMA PRODUCTION

Growing myeloma cells from freeze stock

fusionScreen

Split pos clones into 24 well plates

Screen to weed out false pos

split pos clones into T-25 flasks

Freeze at least Single cell clone2 vials/ clone

Screen

Split pos clones into plates

split in to culture flasks

Freeze stock

Expanded for Ab production

Ad t f Ab d Ab


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Advantage of mAb and pAb

mAb pAb

Homogeneous of product Heterogeneous of productonly one specificity Multiple specificity

constant Reactivity Variable reactivity

Unlimited Limited

No diff. batch-to batch Batch-to batch variationImmunization with crude antigen Immunization with purified antigen

Small doses for therapy High doses for therapy

Di d t f Ab d Ab


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Disadvantage of mAb and pAb

mAb pAb

High production cost Low production cost

Long of production: 4-6 months 3-6 monthsDefinite affinity & Ig class Variable

Definite biol. Character & funct. Variability

No agglutination reaction Agglutination

reaction


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Figure A-15Antibody engineering


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Figure A-15 part 2 of 2


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Diferensiasi sel

Dedifferentiation:

Undefferentiated cell

Absence of appropriate inducer

Penting untuk di pahami: dedifferentiation,

deadaptation, differentiation and selection

To oplasma gond i i


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Toxoplasma gond i i


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Aplikasi TC


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Virus propagation

Trypanosoma


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Trypanosoma

INVASI T gond ii


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INVASI T. gond ii

Apoptosis


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Apoptosis


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Genetic disorders:Kl inefel ter sy ndrom e


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Life long learner & building net work,

TERIMAKASIH

tissue culture technology,242013

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