Laboratory Manual for Biotechnology and Laboratory Science

Professor Lisa Seidman earned her PhD from the University of Wisconsin and has taught for more than thirty years in the Biotechnology Laboratory Technician Program at Madison Area Technical College. She is presently serving as Emeritus Faculty at the college. Dr Mary Ellen Kraus has been a faculty member in the Biotechnology Laboratory Technician Program at Madison Area Technical College for more than twenty years. She earned her BS in Biochemistry from the Pennsylvania State University and her PhD in Biochemistry from Cornell University. Dr Diana Lietzke Brandner earned her MS in Biotechnology Education from the University of Wisconsin-Madison. She has been a Lead Laboratory Coordinator in the Biotechnology Laboratory Technician Program at Madison Area Technical College for more than thirty years. Professor Jeanette Mowery earned her PhD in Biomedical Science from the University of Texas Health Science Center-Houston. She has taught for more than 20 years in the Biotechnology Laboratory Technician Program at Madison Area Technical College and is currently serving as Emeritus Faculty at the college.

UNIT I SAFETY IN THE LABORATORY

Unit Introduction

SAFETY PART 1: CREATING A SAFE WORKPLACE

Fundamental Principles

Classroom Activity 1: Performing a Risk Assessment

Classroom Activity 2: Exploring Safety-Related Government Web Sites

Classroom Activity 3: Responding to Emergencies

SAFETY PART 2: WORKING SAFELY WITH CHEMICALS

Fundamental Principles

Classroom Activity 4: Understanding the Chemicals with Which You Work

Classroom Activity 5: Personal Protection

Laboratory Exercise 1: Tracking the Spread of Chemical Contamination

Classroom Activity 6: Analyzing Safety Issues in a Laboratory Procedure

SAFETY PART 3: WORKING SAFELY WITH BIOLOGICAL HAZARDS

Fundamental Principles

Laboratory Exercise 2: Production of Bioaerosols and Factors Affecting Aerosol

Production

Unit Discussion

Safety Contract, student copy

Safety Contract, to turn in

UNIT II DOCUMENTATION IN THE LABORATORY

Unit Introduction

Classroom Activity 7: Being an Auditor

Laboratory Exercise 3: Keeping a Laboratory Notebook

Classroom Activity 8: Writing and Following an SOP

Unit Discussion

UNIT III METROLOGY IN THE LABORATORY

Unit Introduction

Laboratory Exercise 4: Recording Measurements with the Correct Number of Significant
Figures
Classroom Activity 9: Constructing a Simple Balance

Laboratory Exercise 5: Weight Measurements 1: Good Weighing Practices

Laboratory Exercise 6: Weight Measurements 2: Performance Verification

Laboratory Exercise 7: Volume Measurements 1: Proper Use of Volume
Measuring Devices

Laboratory Exercise 8: Volume Measurements 2: Performance Verification of a M Micropipette

Laboratory Exercise 9: Measuring pH with Accuracy and Precision

Unit Discussion

UNIT IV SPECTROPHOTOMETRY AND THE MEASUREMENT OF LIGHT

Unit Introduction

Laboratory Exercise 10: Color and the Absorbance of Light

Laboratory Exercise 11: Concentration, Absorbance, and Transmittance

Laboratory Exercise 12: Preparing a Standard Curve With Food Coloring and Using it for Quantitation

Classroom Activity 10: Beer’s Law and Calculating an Absorptivity Constant

Laboratory Exercise 13: Determination of the Absorptivity Constant for ONP

Unit Discussion

UNIT V BIOLOGICAL SOLUTIONS

Unit Introduction

Classroom Activity 11: Getting Ready to Prepare Solutions with One Solute: Calculations

Classroom Activity 12: Getting Ready to Prepare Solutions with One Solute: Ordering
Chemicals

Laboratory Exercise 14: Preparing Solutions with One Solute

Laboratory Exercise 15: Preparing Solutions to the Correct Concentration

Laboratory Exercise 16: Working with Buffers

Laboratory Exercise 17: Preparing Breaking Buffer

Laboratory Exercise 18: Preparing TE Buffer

Laboratory Exercise 19: More Practice Making a Buffer

Laboratory Exercise 20: Making a Quality Product in a Simulated Company

Unit Discussion

UNIT VI ASSAYS

Unit Introduction

Laboratory Exercise 21: Two Qualitative Assays

Laboratory Exercise 22: UV Spectrophotometric Assay of DNA: Quantitative Application

Laboratory Exercise 23: UV Spectrophotometric Assay of DNA and Proteins: Qualitative Applications

Laboratory Exercise 24: The Bradford Protein Assay: Learning the Assay
Laboratory Exercise 25: The Bradford Protein Assay: Exploring Assay Verification
Laboratory Exercise 26: The Beta-Galactosidase Enzyme Assay
Laboratory Exercise 27: Comparing the Specific Activity of Two Preparations of
Beta-Galactosidase
Laboratory Exercise 28: Using Spectrophotometry for Quality Control: Niacin
Unit Discussion
UNIT VII BIOLOGICAL SEPARATION METHODS

Unit Introduction

Classroom Activity 13: Planning for Separating Materials Using a Centrifuge
Laboratory Exercise 29: Separation of Two Substances Based on Their Differential
Affinities for Two Phases
Laboratory Exercise 30: Separation and Identification of Dyes Using Paper
Chromatography
Laboratory Exercise 31: Separating Molecules by Agarose Gel Electrophoresis
Laboratory Exercise 32: Using Agarose Gel Electrophoresis to Perform an Assay
Laboratory Exercise 33: Optimizing Agarose Gel Electrophoresis

Laboratory Exercise 34: Quantification of DNA by Agarose Gel Electrophoresis
Laboratory Exercise 35: Introduction to Ion Exchange Column Chromatography

Unit Discussion

UNIT VIII GROWING CELLS

Unit Introduction

GROWING CELLS PART I: BACTERIAL CELLS

Laboratory Exercise 36: Using a Compound Light Microscope

Laboratory Exercise 37: Aseptic Technique on an Open Lab Bench

Laboratory Exercise 38: Working with Bacteria on an Agar Substrate: Isolating
Individual Colonies

Laboratory Exercise 39: Gram Staining

Laboratory Exercise 40: Preparing Phosphate-Buffered Saline

Laboratory Exercise 41: The Aerobic Spread-Plate Method of Enumerating Colony-

Forming Units

Laboratory Exercise 42: Preparing a Growth Curve for E. coli

GROWING CELLS PART 2: MAMMALIAN CELLS

Laboratory Exercise 43: Aseptic Technique in a Biological Safety Cabinet

Laboratory Exercise 44: Making Ham’s F12 Medium from Dehydrated Powder

Laboratory Exercise 45: Examining, Photographing, and Feeding CHO Cells

Laboratory Exercise 46: Counting Cells Using a Hemocytometer

Laboratory Exercise 47: Subculturing CHO Cells

Laboratory Exercise 48: Preparing a Growth Curve for CHO Cells

Unit Discussion

APPENDICES

1. Abbreviations and Acronyms Used in this Laboratory Manual

2. Glossary

3. Selected Bibliography

4. Brief Metric Review

5. Calculating Standard Deviation

6. Equipment, Supplies, and Reagents Required for Each Unit

7. Recipes and Preparation Notes