Frequently Asked Questions

The main documents are as follows:

1. NIH Guidelines for Research Involving Recombinant or Synthetic Nucleic Acid Molecules (2019): https://osp.od.nih.gov/policies/biosafety-and-biosecurity-policy#tab2/
2. Biosafety in Microbiological and Biomedical Laboratories (6^th edition): https://www.cdc.gov/labs/BMBL.html
3. The Kansas State University Laboratory Safety Manual (2018): https://www.k-state.edu/safety/docs/LSM.pdf
4. The Kansas State University Chemical Hygiene Plan (2017): https://www.k-state.edu/safety/docs/CHP-FINAL.PDF
5. Federal Select Agent Regulations: https://www.selectagents.gov/regulations/index.htm
6. Arthropod Containment Guidelines: https://www.liebertpub.com/doi/10.1089/vbz.2018.2431

Depending on the work you are planning on conducting, you will need to complete an IBC registration document and possibly have it reviewed by the IBC before beginning the work. You will need to submit an IBC registration document if your work involves:

1. Research with microbial agents (bacterium, viruses, prions, or parasites) (including microbial work that is generally performed under BSL-1 containment).
2. Research with toxins from biological origins.
3. Research involving recombinant or synthetic nucleic acids.
4. Research with material of human origin (including tissue, blood, cell lines, etc.).
5. Research involving transgenic animals.
6. Research involving transgenic arthropods.
7. Research involving transgenic plants.

The following research/teaching activities will require a full IBC review and approval:

1. Activities that involve the use of RG2 or greater agents.
2. Activities that involve the use of agents requiring a BSL-2 or greater containment.
3. Activities that involve recombinant or synthetic nucleic acids.
4. Activities that involve the use of human materials (blood, tissue, cell lines, etc.).
5. Activities that involve the use of tumorigenic materials.

Yes, CRISPR-Cas9, ZFNs, and TALENs typically involve some type of gene insertion and therefore are subject to the NIH Guidelines for Research Involving Recombinant or Synthetic Nucleic Acid Molecules (NIH Guidelines) and thus require IBC approval. Additionally, if the gene editing system is being expressed from a plasmid or any other type of recombinant vector then the research would be covered under the NIH Guidelines and would also require IBC approval.

Basically, there are two parts to this answer:

1. The K-State IBC has had a standing practice of considering any materials from a human source to be RG2-BSL-2, even when companies distributing the materials (such as ATCC) list the materials as only requiring BSL-1 containment.
2. This determination is based on a passage from Appendix H in the BMBL (page 383):

Recommended Practices
Each institution should conduct a risk assessment based on the origin of the cells or tissues (species and tissue type), as well as the source (recently isolated or well-characterized). Human and other primate cells should be handled using BSL-2 practices and containment. All work should be performed in a BSC, and all material decontaminated by autoclaving or disinfection before discarding.

2a. As cell lines can contain latent viruses, OSHA Bloodborne Pathogens Standard is also considered (https://www.osha.gov/OshDoc/data_BloodborneFacts/bbfact01.pdf):

Bloodborne pathogens are infectious microorganisms present in blood that can cause disease in humans. These pathogens include, but are not limited to, hepatitis Bvirus (HBV), hepatitis C virus (HCV), and human immunodeficiency virus (HIV), the virus that causes AIDS. Workers exposed to bloodborne pathogens are at risk for serious or life-threatening illnesses.

Tumorigenic materials have the following properties (from table 3.2 The Biology of Cancer, Garland Science, 2014, which was adapted in part from S.J. Flint, L.W. Enquist, R.M. Krug et al., Principles of Virology, Washington, DC: ASM Press, 2000):

1. Altered morphology (rounded shape, refractile in phase-contrast microscope).
2. Loss of contact inhibition (ability to grow over one another).
3. Ability to grow without attachment to solid substrate (anchorage independence).
4. Ability to proliferate indefinitely (immortalization).
5. Reduced requirement for mitogenic growth factors.
6. High saturation density (ability to accumulate large numbers of cells in culture dish).
7. Increased transport of glucose.
8. Tumorigenicity (the ability to form tumors in immunocompromised animals).

The following activities do not require IBC review, however, you still may need to submit an IBC registration document (see question 2):

1. Activities using only commercially available deregulated transgenic crops.
2. Activities that involve only the in vitro use of nucleic acids (i.e. PCR, synthetic double stranded RNA) and does not involve the cloning and propagation of recombinant DNA in cells.

ORICS verifies any exemption, so an IBC registration document must still be submitted. If the work is exempt, it won’t need to go to the IBC for a full review. If the work involves the following activities, however, it will require an IBC review and approval prior to initiation of the work:

1. Activities that involve the use of RG2 or greater agents.
2. Activities that involve the use of agents requiring BSL-2 or greater containment.
3. Activities that involve the use of human materials (blood, tissue, cell lines, etc.).
4. Activities that involve the use of tumorigenic materials.

Once an IBC application form is received by ORICS, it will be pre-reviewed by a member of ORICS staff. Initial comment/edits may be sent back to the PI, and an updated copy incorporating the changes may be requested. Depending on the nature of the work, it will either be exempted and required no further action from the PI (generally true of work requiring BSL-1 containment, however, some BSL-1 work involving recombinant DNA work may need to go for a full IBC review) or need to go to the IBC for a full review. A schedule of deadlines and IBC meetings is found here: https://www.k-state.edu/comply/ibc/committeemeetings/untitled.html

After the IBC review, the application will enter into one of three statuses:

1. Approved without stipulations: no further action by PI is needed.
2. Approved with stipulations: the PI will need to address the concerns of the IBC before approval is granted. This is usually accomplished by incorporating the comments into a new draft and submitting for review to ORICS.
3. Withheld: the concerns of the IBC will need to be addressed and incorporated into a new draft and submitted back to the IBC for another review.

In short, whatever happens in the laboratory is the full responsibility of the PI and all those who are in the space must be aware of the risks that are present. How you can ensure compliance can vary depending on the level access that they will need.

1. Laboratory staff member: will need to complete all K-State safety training, CITI training, and any laboratory or project specific training as required by the PI. If you think it best to have one of your staff supervise all the time or at first and then clear them, etc., that is entirely up to your judgement.
2. Outside researcher who will access your laboratory periodically (i.e. use a specific piece of equipment): They will need to complete all K-State safety training, CITI training, and any laboratory or project specific training as required by the PI. If it helps, we are happy to check their records in our system and give you a report back. We can assign any missing training as well.
3. Visitor to the laboratory: Depending on the level of access needed, it would be appropriate to give them a short safety talk about your laboratory before they enter. Describe the hazards present and ask them to stay with their escort and not to touch anything.

To prevent contracting Hepatitis B from human blood or other potentially infectious human materials, all research team members must be offered the Hepatitis B vaccination. Individuals who are already vaccinated or do not wish to receive the vaccine will be asked to sign a declination form.

A sample declination form can be found here: Hepatitis B Vaccine Declination Form (DOC)

Information regarding Hepatitis B and the vaccine is found here: https://www.cdc.gov/hepatitis/hbv/index.htm

Diluted bleach loses its effectiveness after 24 hours, so solutions will need to be made fresh daily.

References:

https://www.osha.gov/sites/default/files/publications/general_decontamination.pdf

https://www.cdc.gov/fungal/candida-auris/c-auris-lab-safety.html

https://www.cdc.gov/foodsafety/communication/salmonella-food.html

https://www.cdc.gov/vhf/ebola/clinicians/non-us-healthcare-settings/chlorine-use.html

Please review this chart and email comply@ksu.edu with any questions.

Use of LVVs at K-State requires the following:

1. Staff members working with LVVs need to be informed of the hazards associated with their use. There is a potential for exposure to LVVs to cause integration of foreign genes into the user’s genome that may result in oncogenic potential in the long term.

2. Staff members working with LVVs need to be aware of the treatments available after a potential exposure. The current recommendation is that Post Exposure Prophylaxis treatment needs to be started 2 to 72 hours after exposure. If an expose occurs, please notify Via Christi Occupational Health 785-776-2813 M-F 8:00AM to 4:30PM. After hours should go to the ER or Urgent care, and the individual exposed should bring the LVV Fact Sheet (here) for the hospital staff to review. Depending on the post exposure risk assessment, prescription drugs may need to be administered very soon after exposure (the current recommendation is 2-72 hours post exposure) so it is important to contact Via Christi Occupational Health ASAP.

3. LVV Fact Sheet will be included in the PI’s Laboratory Biosafety Manual.