NBC2 Presentation to the NVC March 31, 2009 - Northeast Regional ...

mutebabiesBiotechnology

Dec 6, 2012 (4 years and 11 months ago)

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How to Start a Biotechnology

Program

Mary Jane Kurtz, Ph.D NBC
2

Consultant

mjkurtz@biomanufacturing.org


Why Biotechnology?

A number of good reasons

1.
Academics

2.
Technical skills

3.
Biotechnology as a future career

4.
Biotechnology is exciting



Biology can be seen with a molecular perspective



Can answer questions about ourselves


Can provide new medical aid to the afflicted


Getting Support from Your
Administration

This is important to the future success of your
new program

Allows for obtaining outreach to new students,
space to grow, and expenses for needed
equipment

Supports the teachers who are involved in
program


Academic Advantages


Allows students to do up
-
to
-
date hands
-
on
experiments


Demonstrates the integration of all sciences,
math, and engineering through experiences in
the laboratory


It develops writing skills for clarity, focus and


documentation




Technical Advantages


Curriculum can introduce career tracks for
students in a developing scientific area



Using hands
-
on activities, students have a
opportunity to develop skills needed in the
workplace at all levels of formal training



Potential career paths flowing from the
knowledge gained is broad based


Biomanufacturing Curriculum:

National Standard Concepts Covered


Math, Biology,
Chemistry in Curriculum


Measurement


Solutions


Enzyme reactions


Transformation of cells
with DNA


Forces used in
centrifugation,
electrophoresis etc.


National Academy of
Sciences Standards


Unit of Math & Science


Structure/properties of
matter


Chemical
reactions/conservation
of matter


Cell structure and
functions, heredity


Motions and forces










Biotechnology


A plus for Teachers and Students





Integrated science education: Science Technology
Engineering and Math (STEM)


Career pathways towards work/school focused on
science with thousands of new jobs predicted in the
next few years


Laboratory based activities = increased interest


More exciting ways of introducing concepts


Hands
-
on learning is more inclusive


State
-
of
-
the
-
art laboratories







Biomanufacturing vs Biotechnology


Offers diverse career pathways


Hands
-
on learning of science


Understanding of good laboratory practices


Stability in career pathways


Assistance in obtaining college degrees
through company support


Both highly academic and technically oriented
students

Ten Technician Jobs Anchor

Ten Biomanufacturing Departments


Facilities/Metrology


Validation


Environmental Health and
Safety (EH&S)


QA


Upstream Processing


Downstream Processing


QC Microbiology


QC Biochemistry


Process Development








RESEARCH &

DEVELOPMENT
(pre
-
clinical):

Discovery Research

OPERATIONS:


Process

development,
Manufacturing

& Production

QUALITY:


Quality Control


&

Assurance

CLINICAL RESEARCH:

Clinical

Research:

Regulatory

Affairs

Discovery Research


Senior

Scientist

Scientist III,II,I

Research Associate

Process

Development

Director

Supervisor &

Process Development

Technician

Quality Control


(QC)



Clinical Research


Clinical Research Manager

Bioinformatics

Scientist

Engineer

Analyst

Programmer

Manufacturing

&

Production

Supervisor

Associate

Technician (Operator)

Instrumentation Tech

Calibration Technician

Facilities

Management

Manager

Facilities Technician

Shipper/receiver

Chemistry

Chemistry QC

Analyst

QC technician

Microbiology

Microbiology

QC

analyst

QC Technician

Quality

Assurance


(QA)

Documentation

Specialist

QA Documentation
Coordinator

Clinical Research

Clinical Research Manager

Regulatory Affairs

Manager

Associate

Data

Manager

Business Development

Director of Business
Development

Administration

Human resources

Safety Manager



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Student Success Stories

Students completing courses with certificate or

Associate degrees in Biomanufacturing:


www.biomanufacturing.org

http://www.biomanufacturing.org/







1. How to Start A Biotechnology
Program

(After getting support from the administration)

Gain the support the Industry in your Area:


Find out the location and focus of each of the biotechnology/support
companies within (10) miles of your school


Understand the desired skills individuals (such as your students)
should have (Most important of these are soft skills)


Organize an advisory group that can assist you


in curriculum building towards skills needed



This will be a source of information as well as help with technical
supplies







2. Find or Develop a Curriculum


Resources


High School Level
-
Entry Level Community College:
Protein is Cash:


Northeast Biotechnology Center and Collaborative , NBCC:
www.biomanonline.org, Biomanufacturing.org


Biotechnology: Science for the New Millennium
:


Ellyn Dougherty, EMC publisher,
www.emcschool.com



Basic Laboratory Methods in Biotechnology:Textbook and
Laboratory Reference:


Lisa Seidman et al, Prentice Hall,
ISBN 013
-
795535
-
9


Align Curriculum with

Program Goals


What are your goals?


Two year associate degrees transfer to four year
college


Two year associate degree and work


Certificate and immediate work


Research and Development or
Manufacturing?


Goals can vary but should allow majority


of your students to participate in.


Biology Undergraduate

Other Undergraduate
or Engineer

Biomedical Ph.D student

Junior Postdoc &
Senior Postdoc

Other Ph.D student

Industry

Assistant Professor

Staff Scientist

Independent Investigator

Other Science Position

Four year college career pathway



1

Introduction to Biotechnology

Use of bacteria, enzymes and
fermentation by yeast

2

Metrology

Validation of
Pipets

and Calibration

SOP
‘s &

Documentation
of Testing

3

Basic Microbiology

Unit 1

Basic Aseptic Technique and Testing of
Bioload in Biomanufacturing Areas

Gowning up

4

Basic Microbiology

Identification of Bacterial Types

Unit 2

Identify of
Microbe


Morphology

and Electrophoresis

5

Transformation of Cells

With DNA

PGlO transformation of

Bacteria

6

Spectrophotometry

Protein determination of
samples

By

Bradford and UV analysis

7

Genetic Transformation of Bacteria

E. coli transformation with
pGLO

plasmid

8

The Rain
Forest:How

do we find new
pharmaceuticals ?

Growth of
pGlo

transformed bacteria
and isolation of the
p
-
GLO
product

9

Proteins and their purification

Isolation of casein from milk

Using solubility and filtration
techniques

10

Good Manufacturing Practices

Documentation

GMP activity with popcorn

And use of Documentation

11

Chromatographic separation and how it
works

Use of ion exchange and hydrophobic
columns in

Isolating GFP

12

QC Sampling of Product to

Determine its

Purity

Polyacrylamide

Gel Electrophoresis

Intro to Biotechnology for High School/CC

Biotechnology Curriculum

2.
Associate Degree or Certificate Program


Resources:


Introduction to Biomanufacturing: Global
Biomanufacturing Lab manual


NBC2, at

www.biomanufacturing.org



Introduction to Biomanufacturing: Textbook,


NBC2, at
www.biomanufacturing.org


Introduction to Biotechnology Lab Manual


ATE central: Linnea Fletcher, et al

ISBN BITC1311006



Techniques in Biotechnology

for Community College

Week

Lecture

and Topic

Laboratory

1

Acids, Bases and
Buffers

Protein &
Spectrophotometry


Preparation

of

Buffers

Determination of
Protein in solution

2

Enzyme

Kinetics

Kinetics of LDH

Protein

structure

Km of enzymes

3

Purification

by pH

Column
Chromatography

Precipitation of pro

tein

and Affinity
chromatography
4

Enzyme Purification
by Ion Exchange

Purification

of LDH
using a IEX column

5

Identification of

Proteins

Concentration and

Electrophoretic

Techniques

6

High

Pressure
Chromatography

Quality control

Case study of Two Different
Programs

College Degree


Requirements will be more


demanding in math, science


Comprehension, graphing
&computer work is good
can work independently


Has appropriate soft skills
and can work well as a team
member


Is good at trouble shooting
problems in labs




High School or Certificate


Entry level math is pre
-
algebra or algebra 1


Science background is
sketchy (lacks chemistry)


Desire to be placed into


a position after graduation


Likes science but needs
reassurance


Needs to have SOP to carry
out work

High School Curriculum

A Workshop to Instruct Teachers and Students


in

Biomanufacturing and the Bioeconomy

Career Tracks: Protein is Cash
Introduces


Upstream Processing


Production of pGLO into protein by transformed cell


Downstream Processing


Separation of cellular debris and cell supernatant


Purification of pGLO protein by Chromatography


Quality Control


Identification of protein product by electrophoresis


Discovery Research


How new drugs are discovered


Biofuels




1.
Metrology

2.
Transformation of Bacteria

3.
Upstream Processing

4.
Downstream Processing

5.
Quantitative Analysis

6.
Discovery Research


$
Protein is Cash
$

Local Teachers
Workshops

Biomanonline.org

Day 1 : Metrology/Instrumentation

Activities



Calibration of top balance



Verification of


pipet performance



Pipetman Challenge


Outcomes



Introduction of Good Manu
-
facturing Practices



SOP and documentation



Confidence in
measurements by instru
-


mentation

Metrology

Day 2 Transformation of Bacteria

Activities


Addition of pGLO plasmid to
bacteria in Ca++ solution


Heat /shock the mixture


Plate cells onto selective
Luria broth agar + ampicillin


Incubate overnight at 37oC


Note: arabinose acts to turn
on the gene which starts
production of pGLO protein


Outcomes


Selection of cells by growing
on ampicillin plates


Only transformed with cells
will survive due to amp
-
r
gene


Selected colonies will
multiply in Luria broth at
37oC overnight


Aseptic technique and
proper disposal of bacteria



Day 2 E.coli Transformation

with pGLO plasmid

Ampicillin resistance gene (Amp
r
)

and target gene on bacterial plasmid

Bacterial clones

Cell division

Transformation mixture is plated

on to agar plate containing

Ampicillin

Only E. coli containing plasmid

survive on Ampicillin plates

Results of Inserting Foreign DNA
into an Organism








Cells will multiply and produce desired
gene product






pGlO gene expression vector: Green
Fluorescent Protein


Day 2: Upstream Processing:


Cell Growth and Lysis

Activities


Transformed cells grown
overnight in selected media
are separated from media
by centrifugation


Media is removed and
packed cells are lysed


Homogenate is centrifuged


Supernatant with pGLO
protein is retained for


downstream processing


Outcomes


Multiplication of cells


Initial separation of


fluorescent protein from


cell homogenate





Day 3: Downstream Processing:
Purification of Green Fluorescent Protein by
Chromatography

Activities


Separation of product by
Different types of
Chromatography



Size exclusion


Hydrophobic Interaction


Cationic Interaction


Anionic Interaction


Outcomes


Fractions with green
fluorescent protein will
glow and be selected


for analysis



Understand concept of
chromatography as
selective interactions of
compounds with matrix


Isolation and Purification of Green
Fluorescent Protein

Transformed
cells

#1 #2 #3

Fraction

number

Test
tubes

Courtesy of Bio
-
Rad

Separation of Proteins

by Column Chromatograph


Separation by size


Size exclusion chromatography


Separation by hydrophobic characteristics


Water loving vs water hating environments


Separation by ionic charge


Protein has a positive charge & attaches to
negative matrix on column: Cation Exchange


Protein has a negative charge & attaches to
positive matrix on column: Anion Exchange

Size Exclusion Chromatography


Red molecule =10^6 daltons
-

Blue molecule = 600 daltons

How Size Exclusion Works


Molecular size of molecule will separate two
or more molecules


Large molecules can not go into a bead of a
certain size and flows quickly through a
column


Small molecules enter into a bead and flows
slowing through a column.


Size of two different molecules are separated

Hydrophobic Interaction
Chromatography




Low salt




H+

H+

H+

H+

High salt:

2 M
(NH
4
)
2
SO
4

Wash buffer:

1.3 M

(NH
4
)
2
SO
4

Elution

buffer:

10 mM
Tris

Hi Salt

Ion Exchange Chromatography


Proteins bind to
opposite charges on
the matrix


An example of ion
Exchange



Addition of
increasing


Salt/pH should
release proteins



Cation Exchange vs Anion exchange


How proteins are attracted to ion exchangers

Day 4: Quality Control

Activities


Chromatography fractions
prepared for
electrophoresis


Electrophoresis Box is
assembled with PAGE gel


Samples of chromatography
fractions are added to PAGE
gel and ran for 30 minutes


Gels stained and viewed


Outcomes


Analysis of protein samples
by observation on light box
determines protein purity


Standard molecular weight
markers indicate size of
protein


Verification of mol wt by
comparison with standard
proteins and number of
proteins in a single sample


Quality Control

Analysis of Column Fractions



Isolated fractions using Ion Exchange
Chromatography are then analyzed


Electrophoresis by SDS PAGE of fractions
collected











Day 5. Discovery Research and

FDA approval process

Questions


Discovery Science in Drug
development


Good Manufacturing
Practices and the FDA


Initial Biomanufacturing


Process Development


Scale up to full production


of biologic /clinical trials



Answers


Visit local biomanufacturing
plants


Discuss importance of
documentation in the
workplace ie. SOP and Batch
Records


Invite industry
representatives to speak to
the regulated workplace

Bio
-
Rad Partnership

Northeast Biomanufacturing Center and Collaborative
(NBC
2
) is partnered with Bio
-
Rad:



Provides teachers with engaging hands
-
on


biomanufacturing education that is easily


accessible



Introduction to:



metrology,



production (upstream and downstream processing)



quality control biochemistry and clinical trials.

These modules can be brought directly into the classroom



HTTP

Virtual Chromatography

http://www.Atelearning.com/BioChrom

Useful Information for
Biomanufacturing


http://www.Biomanufacturing.org


http://www.Biomanonline.org


http://www.Bio
-
link.org

Background Resources:


“Development of Biotechnology Curriculum for the Biomanufacturing
Industry”, Robert McKown, and George L. Coffman, May/June 2002,
Pharmaceutical Engineering pages 1
-
6.


“ Introduction to Biomanufacturing a Global Biomanufacturing Curriculum”


Northeast Biomanufacturing Center & Collaborative (NBC2) 2011,


publisher :Lulu.com