Biosafety Manual

1. 1.1 Foreword
2. 1.2 Major Updates
3. 1.3 Introduction

"We wish to suggest a structure for the salt of deoxyribose nucleic acid (D.N.A.). This structure has novel features which are of considerable biological interest….. It has not escaped our notice that the specific pairing we have postulated immediately suggests a possible copying mechanism for the genetic material."

–Watson and Crick (1953), Nature 171, pg 737

The use of recombinant DNA (rDNA) and synthetic nucleic acids (sNA) are regulated by the National Institutes of Health (NIH); the guidelines can be found in the publication NIH Guidelines for Research Involving Recombinant or Synthetic Nucleic Acid Molecules (NIH GUIDELINES) (OBA - NIH Guidelines). These guidelines are the official guide to all rDNA and sNA work done at Stanford. It is important to realize that following these guidelines is the responsibility of all investigators at Stanford University and not solely that of investigators that are funded by NIH.

1. 2.1 Exempt rDNA/sNA
2. 2.2 Non-Exempt rDNA/sNA
3. 2.3 Viral Vectors and Transgenes
4. 2.4 Human Gene Transfer
5. 2.5 Transgenic Plants

The Sciences gain by mutual support. When, as the result of my first communications on the fermentations in 1857-1858, it appeared that the ferments, properly so-called, are living beings, that the germs of microscopic organisms abound in the surface of all objects, in the air and in water; that the theory of spontaneous generation is chimerical; that wines, beer, vinegar, the blood, urine and all the fluids of the body undergo none of their usual changes in pure air, both medicine and surgery received fresh stimulation.

Louis Pasteur, Germ Theory and its Applications to Medicine and Surgery, 1878

Laboratories that work with infectious agents pose risks to people within and near them. Infections have been contracted in connection with laboratory work throughout the history of microbiology (a dubious distinction). Studies have illustrated that laboratory-acquired infections are not confined to any one kind of lab or group of people, and that the incidence of infection among untrained and ancillary workers is high, averaging approximately one-third of all acquired infections.

1. 3.1 Biosafety Level Classification
2. 3.2 Laboratory Facility Requirements
3. 3.3 Biosafety Level Work Practice Requirements
4. 3.4 Tissue Culture, Human and Primate Tissue
5. 3.5 Universal Precautions
6. 3.6 Select Agents and Toxins
7. 3.7 Stanford University Select Toxins Program
8. 3.8 Requirements for Research with Prions and Prion-Like Proteins

Good sense is the most evenly distributed thing in the world, for all people suppose themselves so well provided with it that even those who are the most difficult to satisfy in every other respect never seem to desire more than they have. It is not likely that everyone is mistaken; rather this attitude reveals that the ability to judge and distinguish the true from the false, which is properly what one calls good sense or reason, is in fact naturally equally distributed among all people. Thus the diversity of our opinions does not result from some of us being more reasonable than others, but solely from the fact that we conduct our thoughts along different paths, and consider different things. . . .

René Descartes: Discourse on Method (1637)

The NIH has mandated the presence of an Institutional Biosafety Committee for all organizations that come under NIH regulations. At Stanford University, this committee is called the Administrative Panel on Biosafety (APB); the charge of the panel is as follows:

1. 4.1 Charge To The Administrative Panel On Biosafety
2. 4.2 The APB Review Process

Life happens – be prepared.

Fortune cookie, 2001

1. 5.1 Training

This is the story of Leeuwenhoek, the first of the microbe hunters. It is the tale of the bold and persistent and curious explorers and fighters of death who came after him……some of them who were too bold have died – done to death by the immensely small assassins they were studying – and these have passed to an obscure small glory.

–Paul De Kruif, Microbe Hunters (1926), Harcourt, Brace and Co., pub, pg. 3

1. 6.1 Bloodborne Pathogens
2. 6.2 Aerosol Transmissible Diseases
3. 6.3 Vaccinations
4. 6.4 Medical Surveillance

Biosafety is a two-way path involving the creation of a safe working environment for all personnel and ensuring that the work being done does not impact the environment. It is essential to understand that the most important factor in biosafety is the laboratory worker. Good work practice, facility design, training, and protective clothing all fall to the wayside in the presence of a worker who is ignorant or uncaring of proper work procedures.

There are obvious dangers to working with infectious agents/rDNA/sNA. Pathogens can infect a host through a number of routes, and it is important to be aware that a laboratory-acquired infection may not follow the same route as a naturally occurring one. The following are some of the more common accidents that can result in infection, listed in decreasing order of occurrence:

• Spills and splashes
• Needle and syringe sticks
• Sharp objects (including glass)
• Animal bite or scratch
• Mouth pipetting

1. 7.1 Universal Precautions
2. 7.2 Safety Engineered and Needleless Sharps
3. 7.3 Biological Safety/Biosafety Cabinets
4. 7.4 Liquid waste
5. 7.5 Signs and Hazard Communication
6. 7.6 Exposures
7. 7.7 Spill Response
8. 7.8 Reporting

Beam me up, Scotty.

-Captain James T. Kirk, Star Trek

If only it could work that way…. However, until we have a transporter, transportation of biohazardous goods requires a bit more planning and training.

1. 8.1 Shipping of Biohazardous Goods Within Stanford University
2. 8.2 Shipping of Biohazardous Goods Off Stanford University
3. 8.3 What is a Dangerous (Biohazardous) Good?
4. 8.4 Export Controls Related to Biologicals and Toxins
5. 8.5 Importation of Biohazardous Goods onto Stanford University

1. 9.1 Medical Waste
2. 9.2 Sharps Waste
3. 9.3 Mixed Waste
4. 9.4 Animal Carcasses
5. 9.5 Autoclave Waste
6. 9.6 Decontamination
7. 9.7 Autoclaves

….those who pass their lives shut up in houses and offices are not often strong. Their muscles are not thick and hard, and their blood is not rich. But, worse than that, they make their brains and their nerves work too hard; they fatigue their heads and become irritable, or nervous, as it is called, being excited to gayety or anger without sufficient cause. Sometimes, indeed, their brains become altogether deranged, and are no longer able to act properly; the persons are then insane, or lunatic. It is by no means true, however, that the professions and sedentary occupations furnish all of the cases of insanity….

-Paul Bert, First Steps in Scientific Knowledge (1886), J.B. Lippincott, Pub, Part VI, Pg 65.

Laboratories which utilize biological materials must notify the Biosafety program prior to terminating work to ensure that the laboratory has been decontaminated and that the biological material has been secured or properly disposed. If the principal investigator intends to cease work, he or she must notify Biosafety at least 60 days prior to the set departure/closing date. This will allow Biosafety to consult with the principal investigator and perform a walkthrough of the lab to provide recommendations on the most expeditious way to prepare for the move and the final termination of the biohazardous work in the lab. A final lab deactivation inspection will be scheduled accordingly.

1. 10.1 Lab Close Out Procedures
2. 10.2 Disposal of Used Lab Equipment

Note: the following definition is used for the term “exotic plant pathogen” per NIH Guidelines:

In accordance with accepted scientific and regulatory practices of the discipline of plant pathology, an exotic plant pathogen (e.g., virus, bacteria, or fungus) is one that is unknown to occur within the U.S. (see Section V-G, Footnotes and References of Sections I-IV). Determination of whether a pathogen has a potential for serious detrimental impact on managed (agricultural, forest, grassland) or natural ecosystems should be made by the Principal Investigator and the Institutional Biosafety Committee, in consultation with scientists knowledgeable of plant diseases, crops, and ecosystems in the geographic area of the research (Section V-M).

1. 11.1 Biosafety Level 1- Plants (BL1-P)
2. 11.2 Biosafety Level 2 - Plants (BL2-P)
3. 11.3 Biosafety Level 3 - Plants (BL3-P)
4. 11.4 Biosafety Level 4 - Plants (BL4-P)
5. 11.5 Containment Practices for Biosafety Level 1- 3 Plants