1. Identifying a Potential Biotechnology Product
Chapter 6

2. Learning Outcomes
Give examples of biotechnology products derived from plant and animal sources and discuss the challenges of extracting compounds
Identify the steps in a Comprehensive Product Development Plan and use it to determine whether a potential biotechnology product is worth manufacturing
Discuss the types of assays done as potential products move through process development and identify the additional assays required for pharmaceutical development
Describe how an ELISA or a Western blot is conducted and what the results of each assay can reveal
Explain how scientists test the effectiveness of antibiotics and antimicrobials and discuss the significance of antibiotic resistance.
Describe the role of CHO cells in protein product development
Describe the typical recombinant DNA protein product pipeline, additional steps required by the FDA for pharmaceutical proteins, and possible formulations of the final product

3. 6.1 Sources of Potential Products
For thousands of years, people have used plant and animal organs for many purposes.
Recently, scientists have learned how to use plant and animal parts as sources of products:
Pancreas of livestock is used to make insulin.
Foxglove plant is used to regulate irregular heartbeat.

4. Harnessing the Potential of Materials Produced in Nature
Sometimes a product is made in very small quantities in nature so it must be produced
Modeling the Research and Development of a Potential Product
Estimating market size
Identifying product sources
Creating a Comprehensive Product Development Plan (CPDP)
Does the product meet a critical need? Who will use the product?
Is the market large enough to produce sufficient sales? How many customers are there?
Does preliminary data support that the product will work? Will it do what the company claims?
Can patent protection be secured? Can the company prevent other companies from producing it?
Can the company make a profit on the product? How much will it cost to make it? How much can it be sold for?

5. Vocabulary
Amylase – an enzyme that functions to break down the polysaccharide amylose (plant starch) to the disaccharide maltose
Assay – a test

6. 6.1 Review Questions
Why are antibiotics important biotechnology products?
What is the function of the enzyme, amylase?
Why might a company be interested in producing amylase as a product?
Summarize the criteria that a potential product must meet in a CPDP review.

7. 6.2 The Use of Assays
If a substance is chosen as a potential product, researchers must be able to test for its presence, activity, and concentration. The product must be “assayed.”
Other assays are done for potency, toxicity, and stability.
Assays are performed at every step in the development of a product.

8. The 24-well plate reveals samples with different amounts of amylase activity. Amylase breaks down starch to sugar. In the activity assay, as amylase breaks down starch, the dark color of a starch/iodine mixture becomes lighter. The lighter the color, the more active the amylase.

9. Decreasing amounts of protein are indicated by less blue color in the tube. The right-hand tube is a negative control that contains no protein. The tubes to the left have increasing concentrations of protein.

10. Vocabulary
Activity assay – an experiment designed to show a molecule is conducting the reaction that is expected
Concentration assay – a test designed to show the amount of molecule present in a solution
Investigational New Drug (IND) application – a document to the FDA to allow testing of a new drug or product on humans.
Pharmacodynamic (PD) assay – an experiment designed to show how a drug is metabolized (processed) in the body
Potency assay – an experiment designed to show the biochemical effect of a drug on the body
Toxicology assay – an experiment designed to find what quantities of a drug are toxic to cells, tissues, and model organisms
Stability assay – an experiment designed to determine the conditions that affect the shelf life of a drug

11. 6.2 Review Questions
What kind of assay would use Bradford reagent in the test?
For what purpose would a technician use an ELISA?
What does a stability assay measure?
In a large company, which department would have several employees developing and conducting assays?

12. 6.3 Enzyme Linked Immunosorbent Assay (ELISA)
One of the most important and frequently used molecular assays is the ELISA.
An ELISA is very specific and will recognize a single type of protein or other antigenic molecule in a mixture of others.
ELISA utilizes two important phenomenon:
1) antibody antigen specificity to recognize only very specific molecules
2) enzyme activity on colorimetric reagents for visualization purposes.
In an ELISA, the goal is to recognize the antigen and measure its concentration.

14. Vocabulary
Direct ELISA – an ELISA where a primary antibody linked with an enzyme recognizes an antigen and indicates its presence with a colorimetric reaction
Indirect ELISA – an ELISA where a primary antibody binds to an antigen and then a secondary antibody linked with an enzyme recognizes the primary antibody—the antigen presence and concentration is indicated by the degree of a colorimetric reaction

15. 6.3 Review Questions
Explain how antibodies and enzymes are used in ELISAs.
How can a technician know that an antigen is present during an ELISA?
How can a technician know the concentration of an antigen in an ELISA?
What is the difference between a direct and an indirect ELISA?

16. 6.4 Western Blots
In a Western blot, samples are run on a PAGE gel and then the protein bands are transferred to a blotting membrane (PVDF or nitrocellulose).
The visualization of a Western blot can confirm the presence of a particular protein at very low concentrations.

17. Western Blot Gel/Transfer Membrane Setup Diagram
Western Blot Gel/Transfer Membrane Setup Diagram. During a Western blot, electrical current carries protein bands from the PAGE gel to the blot transfer membrane.

18. Vocabulary
PVDF– polyvinylidene fluoride, a high molecular weight fluorocarbon with dielectric properties that make it suitable for Western blots because it is attractive to charged proteins
Nitrocellulose – a modified cellulose molecule used to make paper membrane for blots of nucleic acids and proteins
Antiseptic – antimicrobial solution, such as alcohol or iodine, that is used to clean surfaces

19. 6.4 Review Questions
What kind of molecule is being blot during a Western blot?
What causes molecules to move from a gel to a membrane in a Western blot?
How are blotted molecules visualized during a Western blot?
For what purpose are Western blots used in industry?

20. 6.5 Looking for New Products in Nature
Herbal remedies from the rainforest

21. Vocabulary
Herbal remedies – the products developed from plants that exhibit or are thought to exhibit some medicinal property
Antimicrobial – a substance that kills or slows the growth of one or more microorganisms
Antiseptic – an antimicrobial solution, such as alcohol or iodine, that is used to clean surfaces

22. 6.5 Review Questions
From where do scientists expect that most of the remaining naturally occurring biotechnology products will come?
How can a technician know if a certain type of bacteria is sensitive to an antimicrobial substance?
List a few herbal products that claim to have therapeutic value against depression.
How can molecules be extracted from plant samples for testing purposes?

23. 6.6 Producing Recombinant DNA (rDNA) Protein Products
rDNA was first produced in the mid 1970s.
Recombinant refers to the process of combining DNA from two sources.

24. Vocabulary
Transfection – the genetic engineering, or transformation of mammalian cell lines
Formulation – the form of a product, as in tablet, powder, injectable liquid, etc.

25. 6.6 Review Questions What are CHO cells and what are they used for?
How long does it take to develop, test, and market a typical rDNA protein product?
What does GMO stand for and what does it cover?
Biotechnology products must be formulated before they can be marketed. Name two formulations for a pharmaceutical product other than tablet form.

26. Questions and Comments?