Safety Instruction Manual

July 11, 2000**** Laboratory
XXX Department, Faculty of YYY, ZZZ University

Research Project ID: 2001B0000 -np
Research Project: Crystallographic structure analysis of vaccinia virus particles
Beam line No.: BL01XU
Samples
1. Vaccinia virus(1) Infectious for humans:(2) Disease symptoms:(3) Infectious for animals:(4) Remarks:

Yes.
Pathogenicity has been considerably reduced.
Inoculation causes a benign skin lesion, although
this is typically not contagious. However, a defective
immune system might trigger serious symptoms.
Infectious for vertebrata such as cows, rabbits, andmice.
Crystallized samples lack infectiousness for humans
and animals.

1) Structure of the experiment team and reporting structure- Structure of the experiment teamAffiliation: **** Laboratory, XXX Department, Faculty of YYY, ZZZ University
Principal investigator of biology experiment: Professor xxx, Years of experience inbiology experiments: 20 years
Associate principal investigator of biology experiment: Assistant Professor yyy,
Years of experience in biology experiments: 8 years
Experiment conducted by: AA BBB (student), Years of experience in biology
experiments: 3 years
CC DDD (student), Years of experience in biology experiments:
3 years
・ Reporting structure
The reporting structure is shown in Appendix 1.
2) Entering and leaving the Lab where classified samples are handled
An area of restricted entry should be provided in front of the Lab hatch of BL01XU
where virus samples are handled. Entering and leaving the restricted area should berecorded in a log book. The log book should record the date, name, time, purpose ofentry, and time of leaving.
Special lab gown and footwear should be worn whenever virus samples are brought
in. Personnel should not leave the restricted area while wearing such clothing andfootwear. Hands and fingers should be sterilized using 75% ethanol after handling thesamples.
3) Preventing contamination of the body and clothes when handling specific biologicalsamples, and medications for preventing infections
Goggles, masks, and rubber gloves should be worn during an experiment in order toprevent contamination of the body and clothes. Glutaraldehyde and 75% ethanol
should be immediately available during an experiment in order to avoid contamination.
Vaccinations should be provided as necessary for a particular experimental operationin order to prevent viral infections.
4) Operation of experimental facilities or local confinement facilities that may becontaminated by a specific biological sample
Install a safety cabinet inside the hatch of BL01XU to handle virus samples. The
hatch door should be kept closed and a shutter operation key carried when handling avirus inside the lab hatch. Hatch exhaust equipment should not be operated without afilter.
The inside of the safety cabinet should always be clean and tidy. The floor around thesafety cabinet should be kept free of objects as much as possible in order to prevent thespread of contamination. The floor should be covered with a sheet as necessary.
Appropriate measures should be taken to prevent small animals from entering theexperimental hatch.
Strictly adhere to other precautions that are prescribed by the principal investigator.
5) Storage of specific biological samples (Storage facilities, locking method, andpackaging while in storage)
Virus samples should be kept in the locked refrigerator or freezer dedicated to virus
sample storage in the restricted area in front of the lab hatch. The refrigerator andfreezer should be marked with signage as stipulated in Clause 31 of the Regulations on
Biosafety at SPring-8, and these should not be used for any other purpose.
The keys to the refrigerator and freezer should be kept by the principal investigator.
Virus samples should be placed in crystal form in a glass capillary 0.2 to 1.5 mm indiameter and 0.01 mm in wall thickness. Both ends of the capillary should be sealedwith dental wax or vacuum compound, and each capillary stored in a separate sample
storage bottle. The capillary should be fixed to the lid of the sample storage bottle
using compound. Several sample storage bottles should be packed in a tight box, andthe tight boxes should be placed in a sealed stainless steel tank for transportation.
6) Disinfection and sterilization equipment to be available at all times - types, purposes,
number, and storage locations
Types
Autoclave
75% alcohol
Glutaraldehyde

PurposeHigh-pressure sterilization
Sterilization
Sterilization

Number122

Location
Lab hatch＊
Safety cabinet
Safety cabinet andLab hatch entrance(See Appendix 2)7) Precautions for experiments involving specific biological samples
Verify that the glass capillary is not damaged when you remove a sample bottle fromthe tight box.
Virus crystals should remain in a glass capillary to be mounted on a goniometer.
Sample position and diffraction intensity data collection should be adjusted remotelyusing a computer inside the work control area. Except while data is collected, specificbiological samples should be kept in a refrigerator or a freezer with signage statingthat the specified biological samples are in storage.
Glass capillaries should not be removed from the transportation container outside
the safety cabinet.
8) Handling log for specific biological samples
Whenever a biological sample is handled, the name of the person handling it and thedate, time, location, and purpose should be recorded.
9) Disinfection, sterilization and disposal of specific biological sample experimentapparatuses, culture media, chemicals, animals, etc.
Items that may come in contact with virus samples during an experiment, including
the following, should be brought back to the University after sterilization withhigh-pressure steam.
Sample holders used during sample transportation
Tweezers to handle samples
Culture media and chemicals also should be sterilized using high-pressure steam andbrought back to the University.
Personal protective equipment, including goggles, masks and arm covers, should beplaced in a plastic bag and brought back to the University.
10) Safety education and training for experiment personnel and records thereof.
The principal investigator of a biological experiment is responsible for conducting
education and training regarding experimental procedures and safety measures,
including the properties and handling of a vaccine virus and the handling of glass
capillaries. The education and training should cover Regulations on Biosafety atSPring-8, Detailed Regulations on Biosafety at SPring-8, and Criteria for Laboratory
Facilities Where Samples are Handled and for Safety Work.
Whenever education and training is conducted, the date, time, content, name ofinstructor, and names of students should be recorded.
11) Medical examination of experiment personnel
Personnel engaging in an experiment (including biological experiment principal
investigator) should undergo a medical examination prior to handling a sample andanother medical examination within one year from the start of handling the sample at
his or her University. The results of the examinations should be used to determine
whether or not a problem exists affecting the continuance of the experiment, and thedecision should be communicated to the head of the Safety Office.
12) Inspection of BSL-2 Lab (Frequency of inspection of items such as filters in thesafety cabinet)
Inspect the exhaust filter for clogging and the safety cabinet for any issues before
and after the start of a research project. Record in the log the inspector's name and theinspection date, time, inspected items, and the results.
13) Restoration of lab after an experiment and record-keeping
After an experiment, disinfect with glutaraldehyde and 75% alcohol the locations
where infectious samples were used. Items such as the safety cabinet should bereturned to the original state. Inspect the exhaust filter for clogging and other issues. If
an issue is found, correct it to restore the original working order.
Record the name of the person who verified the status along with the verification

date, time, item, and remedies taken to restore the original condition. This log should
be presented to the Facility Utilization Director.
14) In case of an accident or an event that could lead to and accident
If a glass capillary is broken and the virus sample has leaked or is suspected ofhaving leaked, disinfect and sterilize the area suspected to be contaminated and theexperimental apparatuses without delay using glutaraldehyde and 75% alcohol. Items
that can be sterilized with high-pressure steam should be so treated. If skin came into
contact with the leaked virus sample, disinfect the skin with 75% alcohol.
If an accident occurs, immediately report it to the biological experiment principal
investigator. The principal investigator should then immediately report it to theFacility Utilization Director and the head of the Safety Office.
The biological experiment principal investigator should provide remedial actions as
outlined above in cooperation with the head of the Safety Office under the supervisionof the Biological Sample Safety Manager.
15) Safety measures for specific biological samples in case of a disaster, such as anearthquake or fireIn the case of a disaster, such as an earthquake and fire, virus samples should beplaced in a safe location so that no contamination occurs. Personnel who have been ormight have been contaminated with virus samples should be rescued without delay.
Emergency measures should be provided as prescribed in the previous section, 14) In

case of an accident or event that could lead to an accident.

Fires and injuries to personnel should be reported as prescribed in Appendix 3,

Emergency reporting.

16) Entrance of outside individuals to BSL-2 Lab
Individuals who are not involved in an experiment but would like to enter the BSL-2
Lab should follow the instructions of the biological experiment principal investigator.
The entry and exit of such individuals should be recorded.
17) Other safety considerations
The emergency contact number of **** Laboratory, XXX Department, Faculty of YYY,ZZZ University, is 0＊＊-2＊＊-9＊＊＊.

添付書類第11Appendix

BL Equipment Director:
BL 装置責任者：■田 ■太（PHS 0001）( PHS 0001)

BL Manager:

X yama Y suke, Dean, Faculty of AA,○○大学△△学部長：●山 ●助BB University

Z ichi W kawa, Chair, Department of VVV

BL担当者：▲暴 ▲ニ_（ PHS 0003）（PHS 0003）□□□学科主任教授：■川 ■一(Tel: 0**-2-**-S***)（Tel : 0＊＊-2＊＊-8＊＊＊）

Professor XXX OOO, Principal Investigator of Biological Experiment
生物実験責任者 ：○○ ○○（教授）Assistant Professor PPP UUU, AssociatePrincipal△△（助手）Investigator of Biology Experiment
生物実験副責任者：△△

Experiment conducted by:
実験従事者 ：●● ●●×× ××Experiment group management structure

実験グループ管理

Appendix
2
添付書類第２Exhaustunit
排気ユニット
排気ダクト
Exhaust duct

Existing equipment

既設機器BeamLine
ビームライン

安全キャビネット
Safetycabinet★

Storage-type hand washer
貯留式手洗器High-pressure
steam sterilizer
高圧蒸気滅菌器
実験ハッチ
Experiment
hatch

Locked refrigerator
鍵付冷蔵庫
★75%
alcohol and glutaraldehyde★；75％アルコール、グルタルアルデヒド★

Sterilizing locker
殺菌ロッカー★

ハッチ扉
Hatch door
Shutter
operation key
シャッター操作キー

作業管理室

Workcontrol room
（立入り制限区域）(restricted entrance)
ドア（記帳による入室管理）
Door (entryand exit are logged)

BL01XU 実験ハッチおよび作業管理室
BL-1XU experiment
hatch and work control room

Biological experiment inspection check list

Biological experiment inspection check list (draft)

Director, Safety Office, SPring-8
I hereby submit my report in accordance with Clause 31-1 of Regulations on Biosafety.
Principal Investigator of Biology Experiment:
Prior to experiment:
Date:
Month:
Year:
During experiment:
Date:
Month:Year:
After experiment:
Date:
Month:
Year:
Inspections performed:

Issuesdiscovered: Remarks:

If an issue is discovered, please report it to the Safety Office (ext. 2200)