Hybridoma cell Technology.pptx

Hybridoma cell technology is the most paramount for monoclonal antibody production that used to develop diagnostic assay and therapeutic against cancer and other disease

1. PMP 4102: Medical Bio-products Development Presentation on: Hybridoma Cells Production and Its Advantage BY Biniam M. May, 2022 Nairobi, KENYA PAN AFRICAN UNIVERSITY INSTITUTE FOR BASIC SCIENCES, TECHNOLOGY AND INNOVATION JOMO KENYATTA UNIVERSITY OF AGRICULTURE AND TECHNOLOGY

2. Introduction Principle of hybridoma c ell production and selection General procedure for Hybridoma cell development Equipment and chemicals for hybridoma cell production Advantage/importance of hybridoma cell Conclusion Reference OUTLINE

3. INTRODUCTION D iscovery of the hybridoma technology Antibody production of by plasma cells (effector B cells) has first described by Astrid Fagraeus in 1948 . However , these cell lines were not effective in-vitro antibody production because of mortality (short lifespan ). In 1975, Kohler and Milstein were developed a technique known as hybridoma technology for developing hybridoma cell(immortal cell line ). They produce murine antibody They were awarded the 1984 Noble prize for physiology and medicine.

4. Principle of hybridoma cells generation and selection Hybridoma cell can be generated through fusing short-lived plasma cells with myelomas cell line (cancer cells ) . After cell fusion, three types of cells are generated:  Unfused  myeloma cells -immortal but lack HGPRTase an enzyme that allows cells to grow in HAT medium. U nfused  plasma spleen cells -short lifespans but have HGPRT, Fused  hybridoma  cells - which contain the genetic machinery of both myelomas and plasma cells, are able to survive and replicate indefinitely.   HAT is the most known medium for selection of hybridoma cell from other unfused cells Notes: HGPRT - H ypoxanthine-guanine phosphoribosyl transferase HAT - Hypoxanthine- Aminopterin - Thymidin )

5. The selection of hybridoma from this mixture depends on the modulation of the nucleotide synthesis process in HAT medium .   Principle of hybridoma cells generation and selection Cell fusion between myelomas and plasma cells is recurrently induced by electric pulses ( electrofusion ) or polyethylene glycol (PEG). Alternatively, the immortalization of plasma cells can be triggered by infection with the Epstein-Barr virus (EBV);   However, this process is less efficient than PEG-mediated cell fusion which remains the gold standard reagent for hybridoma generation . H ypoxanthine-guanine phosphoribosyltransferase and thymidine kinase (TK) 

6. General procedure for hybridoma cells development Hybridoma are immortal antibody-secreting cell lines for diagnostic and therapeutic use. The generation of antibodies through the hybridoma technology is performed in five steps: Development  and optimization of the antigen (immunogen) Immunization  of the host with the target antigen to initiate B cells’ maturation Isolation  of mature B cells from the spleen and fusion with myelomas Screening  and selection of hybridomas Propagation  of hybridomas with highest affinity to the target and purification of mAbs

7. General procedure for hybridoma cells and its antibody generation

8. Equipment and chemicals r equired for fusion and growth of hybridomas Incubator Biohazard or laminar flow hood Liquid nitrogen facility Inverted microscope

9. Generation of highly specific mAbs  Natural affinity maturation  Less labor -intensive  Unlimited quantities of cost-effective, consistent, homogenized antibody production . Perfect for: R esearch , Diagnostic assay development Therapeutic uses. Advantages/Importance of hybridoma cell technology

10. Conclusion Hybridoma cell = plasma cell (B-cell) + myeloma cell Hybridoma cell : Can grow in HAT medium Used for monoclonal antibody production The produced monoclonal antibody can be:

11. Goodman & Gilman’s The Pharmacological Basis of Therapeutics 12 th edn Katzung Basic & clinical Pharmacology 11 th edn Jancie , M Recheit , etal . Naturebiotechnology , 2005, Sep,Vol . 23,No.9Stamatis-Nick C. J Allergy Clin.Immunol , Oct. 2005 www.wikipedia.com www.medscape.com REFERENCES