Radiation Environment and Medicine 2022 Vol.11, No.1 34–39

ReportCytogenetic Biodosimetry in Radiation Emergency Medicine:
2. Biosafety and Chemical Safety in Biodosimetry Laboratory
Kosuke Kasai1, Yu Abe2, Valerie Goh Swee Ting3, Mai Tran Thanh4, Yohei Fujishima5,
Ryo Nakayama1, 5, Kai Takebayashi1, 5, Akifumi Nakata6, Kentaro Ariyoshi7, Hiroyuki Hanada8,
Mitsuaki A. Yoshida5, 9, Katsuhiro Ito8* and Tomisato Miura5**
Department of Bioscience and Laboratory Medicine, Hirosaki University Graduate School of Health Sciences,66-1 Hon-cho, Hirosaki, Aomori 036-8564, Japan2
Department of Radiation Biology and Protection, Atomic Bomb Disease Institute, Nagasaki University,
1-12-4 Sakamoto, Nagasaki, Nagasaki 852-8523, Japan3
Department of Radiobiology, Singapore Nuclear Research and Safety Initiative, National University of Singapoore,
1 Create Way, Singapore 138602, Singapore
4Biodosimetry Group, Centre of Radiation Technology and Biotechnology, Dalat Nuclear Research Institute,
1 Nguyen Tu Luc, Ward 8, Dalat City, Lamdong Province, Vietnam
5Department of Risk Analysis and Biodosimetry, Institute of Radiation Emergency Medicine, Hirosaki University,
66-1 Hon-cho, Hirosaki, Aomori 036-8564, Japan
6
Faculty of Pharmaceutical Sciences, Hokkaido University of Science, 15-4-1, Maeda 7-jo, Teine-ku, Sapporo, Hokkaido 006-8585, Japan
7
Center for Integrated Science and Humanities, Fukushima Medical University, 1 Hikariga-oka, Fukushima City, Fukushima, 960-1295, Japan
8
Advanced Emergency and Critical Care Center, Hirosaki University Hospital, Hirosaki University,
53 Hon-cho, Hirosaki, Aomori 036-8563, Japan
9
Institute of Chromosome Life Science, 11-5-409, Fukuokachuo 2-Chome, Fujimino-shi, Saitama 356-0031, Japan
1

Received 25 October 2021; revised 19 November 2021; accepted 6 December 2021

In a biodosimetry laboratory, blood collected from exposed patients is cultured and the exposure
dose is estimated based on the frequency of chromosome aberrations. Blood is defined as aninfectious specimen because it may contain hepatitis virus and human immunodeficiency virus(HIV) and must be handled in a biosafety level (BSL) 2 facility. Due to the recent coronavirus
pandemic with SARS-CoV-2, further strengthening of infection control measures is required.
This article outlines the requirements for setting up a BSL2 laboratory, personal protective
equipment for infection control, treatment of infectious biological waste and emergency response
measures. Furthermore, it is essential to safely manage hazardous chemicals used in biodosimetry.
Biodosimetry laboratories should conduct risk assessments of blood handling and chemical use
and consider risk mitigation measures. In addition, laboratory personnel must educate workers on
infection control and chemical safety.
Key words: cytogenetic biodosimetry, laboratory management, biosafety, chemical safety,
emergency response*Katsuhiro Ito: Advanced Emergency and Critical Care Center, Hirosaki University Hospital, Hirosaki University, 53 Hon-cho, Hirosaki, Aomori 036-8563, Japan
E-mail: itohkcck@hirosaki-u.ac.jp**Tomisato Miura: Department of Risk Analysis and Biodosimetry, Institute of Radiation Emergency Medicine, Hirosaki University, 66-1 Hon-cho, Hirosaki, Aomori036-8564, Japan
E-mail: tomisato@hirosaki-u.ac.jp
Copyright © 2022 by Hirosaki University. All rights reserved.

Kosuke Kasai et al./ Radiation Environment and Medicine 2022 Vol.11, No.1 34–39
A35

BFig. 1. Comparison of a clean bench and a safety cabinet. A. clean bench, B. safety cabinet.

1. Introduction
Laboratory facilities are designated to biosafety levels(BSL) 1 to 4 depending on the type of pathogens handled.
BSL1 has the most basic level of containment while
BSL4 has the maximum level of containment. The
risk levels of pathogens are categorized based on thepathogenicity of infectious microorganisms, transmission
route and host range of microorganisms, availability ofeffective preventive measures and availability of effective
therapeutic methods1).
In biological dose assessment (biodosimetry),
blood collected from exposed patients is cultured andchromosome aberrations are analyzed. Human blood is aninfectious specimen that is classified as a United Nations(UN) standard category B biological substance2). The
World Health Organization (WHO) laboratory biosafety
guidelines recommend a BSL2 environment for blood
handling. While handling blood specimens, it is necessary
to take thorough infection control measures and properly
dispose of infectious waste1). In addition, methanol, acetic
acid and other hazardous and flammable substances areused when preparing chromosome specimens. Chemicals
used should also be properly handled with care.
Biodosimetry laboratories are required to manage
their laboratories in accordance with national laws andguidelines, as well as international guidelines. Biosafety
guidelines and chemical handling guidelines must be
established for each facility. Efforts must be made toensure rigorous safety management and laboratory users
must be educated in workplace health and safety. This
paper introduces the establishment of a biodosimetrylaboratory that complies with the laws and guidelines in
infection control, chemical safety, infectious biologicalwaste treatment and liquid chemical waste disposal.
2. Infection control and BSL2 laboratory maintenance
Peripheral blood used for biodosimetry may containpathogens such as hepatitis B virus (HBV), hepatitis
C virus (HCV), human immunodeficiency virus (HIV)and the novel coronavirus (SARS-CoV-2). Therefore,
human blood must be treated as an infectious specimen.
In particular, as the SARS-CoV-2 virus is transmitted
by aerosols, infection protection against aerosols andother biohazard control measures should be taken whenhandling blood collected from patients with suspectedradiation exposure and positively diagnosed with
COVID-19 3）.
Blood is usually handled in a BSL2 laboratory as
microbial infectious agents may cause moderate levels of
disease in the event of exposure. Laboratories responsible
for biodosimetry in radiation emergency medicine should
set up BSL2 laboratories in accordance with WHOʼslaboratory biosafety guidelines1) to prevent infections andcontamination. In addition, BSL2 laboratory certifications
and approval are required and regulated by the laboratoryʼs
respective institutes.
The requirements of the BSL2 laboratory and theirdetails are shown below. These requirements are notexhaustive. For more information, please refer to yourinstitutesʼ BSL2 certification criteria.
2.1. BSL2 laboratory requirementsa) The laboratory has the structure and equipment of anormal biological laboratory.
b) The laboratory is equipped with a biological safetycabinet (only when performing operations that areprone to aerosols).
Note 1. Aerosols may be generated when the
blood collection tube or cell culture tube isopened.
Note 2. The safety cabinet needs to be inspected
at least once a year.
Note 3. A clean bench is a device that protects
the cell culture from contamination byproviding an aseptic working environment.
Aerosols generated from an infectious

36Kosuke Kasai et al./ Radiation Environment and Medicine 2022 Vol.11, No.1 34–39

Fig. 2. Representative image of a Biohazard sign for a BSL2 laboratory.
Contact personnel listed in the sign must be contactable at all times.

Fig. 3. Personal protective equipment for biodosimetry.

Fig. 4. Centrifuge bucket with a lid for biohazardous aerosol protection.

sample can be released outside of thecabinet. It is not designed for personalprotection (Fig. 1A). Clean benches arethus not recommended for biodosimetry.
On the other hand, infectious aerosols are
not released outside the Class II safetycabinet and can be used for personal
protection (Fig. 1B).
c) Install the autoclave in the building where thelaboratory is located if an autoclave is used forsterilization.
2.2. BSL2 laboratory display and access restrictionsa) Internationally recognized biohazard symbols must be
displayed on the doors outside of rooms which handle
infectious specimens such as blood (Fig. 2).
b) Only certified personnel are allowed access to thelaboratory work area.
c) Laboratory doors must be closed.

d) Food and drink cannot be stored in the laboratorywork area.
e) Eating, drinking, smoking, makeup application andcontact lens handling in the laboratory work area areprohibited.
2.3. Personal protective equipment (PPE)a) Wear the following personal protective equipment(Fig. 3) when handling blood, bodily fluids or otherpotentially infectious samples.
• Isolation gown
• Surgical mask
• Goggles or face shield• Disposable gloves• Head cover• Closed-toed footwearb) The gloves used must be removed aseptically. Wash
your hands after removing your gloves.
c) It is prohibited to wear lab coats outside theKosuke Kasai et al./ Radiation Environment and Medicine 2022 Vol.11, No.1 34–39

laboratory, such as in the office, library and staffroom.
d) Do not wear exposed footwear in the laboratory. Feetand toes should be fully covered.
e) Lab coats should not be stored in the same lockers or
cupboards as everyday clothing.
2.4. Other biohazard safety measures required for biodosimetrya) Centrifuge
Centrifuges are used for lymphocyte isolation,
cell wash and cell harvest and fixation. To prevent
biohazardous aerosol transmission and contamination
during centrifugation, it is highly recommended toattach the lid to the bucket of the centrifuge (Fig. 4) or
to install the centrifuge in the biological safety cabinet.
Pathogen contamination in the working environment canbe prevented by removing the centrifuge tube from theclosed bucket in the safety cabinet.
b) CO2 incubator
In biodosimetry, blood or isolated peripheral bloodmononuclear cells are directly cultured in a CO 2
incubator without pathogen inactivation. Disinfection andsterilization inside the CO2 incubator should be regularly
performed with its in-built heat-sterilization function.
Moreover, CO2 is often supplied from a compressedgas cylinder connected to the incubator. In accordancewith the safety regulations outlined by your country on
the use of compressed gases, efforts must be made toprevent accidents caused by pressurized gas, such as fall
prevention measures. Compressed gas cylinders should
also be used before the inspection expiration date, or be
replaced or re-inspected if expired. Valves should also
be maintained and inspected regularly. This reduces therisk of chemical and physical hazards associated withcompressed gas cylinders.
3. Infectious liquid and solid biohazardous waste
handling
After using infectious specimens that may containpathogens, pathogen spread must be inhibited. Leftover
blood, culture supernatant after blood culture, reagentsused for blood cell washing and other equipment whereblood and its components were in contact with must be
disinfected or sterilized by an appropriate method anddiscarded.
3.1. Biological liquid waste disinfection
Any liquid reagents in contact with blood should beequally treated as an infectious specimen like blood.
After collecting these liquids in an appropriate storage
container, sodium hypochlorite solution (effective chlorine
concentration 0.1% as a regular disinfectant, effective
chlorine concentration 0.5% for large amounts of organic37matter) can be added as a first step to decontamination1).
Proper waste disposal can be finally performed after
high pressure thermal sterilization (121 °C , 20 minutes).
If there is a waste disposal facility capable of handling
biological liquid waste disposal according to nationalguidelines, biological liquid waste in the lab should befirst treated with suitable disinfectants before contacting
the facility for disposal.
If chemical disinfectants other than a hypochlorous
acid-based solution are used, ensure that the concentrationand type of the disinfectant used is suitable for effectivesterilization of the target pathogen and liquid waste
volume. In addition, if other chemicals are found in the
biological liquid waste such as acids or ammoniumcontaining materials, hypochlorous acid treatment will
generate hazardous gases. In that case, another suitabledisinfectant should be used instead.
3.2. Disposal of solid biohazardous waste in contact withinfectious specimens
Pipette tips, cell culture tubes, blood collection tubes
and other biological disposable equipment which werein contact with infectious specimens must first be
sterilized by high pressure thermal sterilization (121 °C,20 minutes). They can then be discarded or separately
disposed of as medical waste. If the laboratory isunable to perform autoclaving within its premises, solid
biohazardous waste must be sent to a waste disposal
facility capable of handling solid biohazardous waste
disposal according to national guidelines.
4. Chemical safety in biodosimetry
Several hazardous chemicals and organic solvents areused in biodosimetry and its related studies. The person
responsible for the biodosimetry laboratories must be
educated and trained in the hazardous chemical handlingin accordance with relevant laws and guidelines. Safety
data sheets (SDS) for each hazardous chemical used in
the laboratory must be prepared and managed so thatthey can be viewed promptly. All hazardous chemicals
used should also be labelled with their appropriate
Globally Harmonized System of Classification andLabelling of Chemicals (GHS) hazard pictograms4).
4.1. Hazardous chemicals
Proper management of hazardous chemicals used in thelab is necessary for workplace and health safety. Any
hazardous chemicals used must be made known to theinstitute, locked and stored in their respective chemicalcabinets, and the amount of the chemical stored andused must be recorded. Laboratories that use hazardous
chemicals must be well-equipped with precautions
regarding health hazards and have an emergency

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Kosuke Kasai et al./ Radiation Environment and Medicine 2022 Vol.11, No.1 34–39

response plan in place. In addition, laboratory personnel
must educate users on chemical safety and potential
health hazards associated with hazardous chemicals usedin the lab.
4.2. Flammable chemicals
Flammable chemicals such as methanol are used inbiodosimetry research. Flammable chemicals should be
stored in a flammable safety cabinet for fire protection.
When using these chemicals, it is also necessary toevaluate the risk of fire and explosion to workers andconsider measures to reduce the associated risks.
4.3. Local exhaust ventilation
When using hazardous chemical substances or organic
solvents, exhaust equipment specific for each chemicalclassification is required. Local exhaust devices such as
fume hoods must be used to reduce the risk of harmful
health effects in researchers by preventing prolonged
exposure to noxious chemical fumes.
4.4. Liquid chemical waste
The chemicals used must be classified according tofacility regulations and disposed of in an appropriatemanner.
5. Measures related to emergency response
In laboratories that handle infectious samples and toxic
substances, emergency response must be establishedin case of potential exposure accidents, spills, fires andnatural disasters. In the event of an exposure accident(e.g. needle stick injuries, ingestion, aerosol exposure)while working in the laboratory, the worker must
promptly contact the laboratory manager and treat theaffected area immediately. In the event of a biological orchemical spill, laboratory staff should be trained to handle
and clean up spills with an appropriately prepared spillkit. All accidents and spills should be promptly reported
to the respective organizationʼs Occupational Safety and
Health department.
In addition, emergency contact net works andevacuation routes must be established in case of natural
disasters such as earthquakes and fires. In the event of anatural disaster or fire, it is important to take appropriate
actions in response to the disaster, giving top priority toensuring personal safety. The laboratory manager must
educate these emergency responses during the laboratory
safety briefing.
6. Conclusion
As blood used in biodosimetry in radiation emergencymedicine is an infectious biological sample, efforts must

be made to protect researchers against infection inaccordance with various laws and guidelines set by eachcountry and international guideline. In addition, there
are various regulations regarding the use of chemicals
and the handling of liquid waste. The person in chargeof the biodosimetry lab must understand these laws andguidelines to equip the laboratory and educate its users.

BSL2 laboratory• Install a Class II biological safety cabinet.
• Install an autoclave in the building where thelaboratory is located.
• Display the biohazardous signs and emergency
contact information on the BSL2 laboratory
entrance door.
• Onl y aut horized personnel can enter t helaboratory.
• Wear personal protective equipment.
Infectious biohazardous waste• Pathogen spread should be inhibited.
• Any solution in contact with blood must be
sterilized and treated appropriately with adisinfectant.
• Any solid waste in contact with blood must be
disposed of properly.
Chemical safety and management• Safety data sheet (SDS) of each chemical must beprepared.
• Strictly manage the use of hazardous chemicals inaccordance with national regulations.
• Install a local exhaust system such as a fume
hood. Chemicals emitting hazardous fumes oraerosols should be used inside the fume hood.
• It is necessary to carry out a risk assessment of
all hazardous chemicals and consider measures toreduce the risks.
• Liquid chemical waste must be sorted according
to regulations and disposed of properly.
Measures related to emergency response• Ensuring personal safety is a top priority.
• In the event of a spill or an exposure accident
while working in the laboratory, the worker must
promptly contact the laboratory manager.
• Promptly report to the Occupational Safety andHealth department set up in each organization.
• Emergency contact networks and evacuationroutes must be established.

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39

2.

The authors declare that they have no conflict ofinterests.

WHO. Guidance on regulations for the transport of infections
substances 2021-2022: applicable as from 1 January 2021. Geneva:
World Health Organization; 2021.

3.

WHO. Laboratory biosafety guidance related to coronavirus
desease (COVID-19): Interim guidance, 28 January 2021. Geneva:
World Health Organization; 2021.

References

4.

United Nations. Globally Harmonized System of Classificationand Labelling of Chemicals, Ninth revised ed. New York: United
Nations Publication; 2021.

Conflict of Interest

1.

WHO. Laboratory biosafety manual. 3rd ed. Genera: World HealthOrganization; 2004.