Standard Operating Procedure
Liquid Nitrogen
This is an SOP template and is not complete until: 1) lab specific information is entered into the
box below 2) lab specific protocol/procedure is added to the protocol/procedure section and
3) SOP has been signed and dated by the PI and relevant lab personnel.
Print a copy and insert into your
Laboratory Safety Manual and Chemical Hygiene Plan.
Refer to instructions for assistance.
Department:
Date SOP was written:
Date SOP was approved by PI/lab supervisor:
Principal Investigator:
Internal Lab Safety Coordinator/Lab Manager:
Lab Phone:
Office Phone:
Emergency Contact:
(Name and Phone Number)
Location(s) covered by this SOP:
(Building/Room Number)
Type of SOP:
☐ Process
☒Hazardous Chemical
☐ Hazardous Class
Purpose
Cryogenic fluids are liquids that exist at a very low temperature (below −150 °C). Common
examples of cryogenic liquids are liquid nitrogen and helium. These fluids are used as coolants
for various instruments and experiments.
Physical & Chemical Properties/Definition of Chemical Group
Class: Toxic
Liquid Nitrogen
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CAS #: 7727-37-9
Formula: N2
Form: gas
Color: colorless
Potential Hazards/Toxicity
Emergency Overview
OSHA Hazards
Compressed Gas
GHS Classification
Gases under pressure (Compressed gas)
GHS Label elements
Signal word: Warning
Hazard statement(s)
H280 Contains gas under pressure; may explode if heated.
Precautionary statement(s)
P410 + P403 Protect from sunlight. Store in a well-ventilated place.
Personal Protective Equipment (PPE)
Respiratory Protection
Respiratory protection is not required. Where protection is desired, use multi-purpose combination (US) or
type ABEK (EN 14387) respirator cartridges Use respirators and components test ed and approved
under appropriate government standards such as NIOSH (US) or CEN (EU).
Hand Protection
For prolonged or repeated contact use protective gloves.
NOTE: Consult with your preferred glove manufacturer to ensure that the gloves you plan on using are
compatible.
Refer to glove selection chart from the links below:
http://www.ansellpro.com/download/Ansell_8thEditionChemicalResistanceGuide.pdf
OR
http://www.allsafetyproducts.com/glove-selection-chart-chemical-breakthrough-ratings.html
OR
http://www.showabestglove.com/site/default.aspx
OR
http://www.mapaglove.com/
Wear insulated gloves.
Eye Protection
ANSI approved safety glasses or goggles are recommended.
Skin and Body Protection
Lab coat, long pants, and closed-toe shoes are required.
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Hygiene Measures
After working with chemical, immediately remove gloves, wash hands and arms with soap and water.
Engineering Controls
Work with chemicals in a certified fume hood.
First Aid Procedures
If inhaled
Remove to fresh air. If not breathing, give artificial respiration. If breathing is difficult, give oxygen. Call a
physician
In case of skin contact
Immediately flush skin with plenty of soap and water for at least 15 minutes while removing contaminated
clothing and shoes. Get medical attention immediately
In case of eye contact
Immediately flush eyes with copious amounts of water for at least 15 minutes while removing
contaminated clothing and shoes. Get medical attention immediately
If swallowed
Wash out mouth with water provided person is conscious. Never give anything by mouth to an
unconscious person. Call a physician.
Special Handling and Storage Requirements
Conditions for safe storage
Do not keep in closed container. Contents under pressure.
Spill and Accident Procedure
Chemical Spill Dial 911 and x59797
Spill – Assess the extent of danger. Help contaminated or injured persons. Evacuate the spill area.
Avoid breathing vapors. If possible, confine the spill to a small area using a spill kit or absorbent material.
Keep others from entering contaminated area (e.g., use caution tape, barriers, etc.).
Small (<1 L) – If you have training, you may assist in the clean-up effort. Use appropriate personal
protective equipment and clean-up material for chemical spilled. Double bag spill waste in clear plastic
bags, label and take to the next chemical waste pick-up.
Large (>1 L) – Dial 911 (or 310-825-1491 from cell phone) and EH&S at x59797 for assistance.
Chemical Spill on Body or Clothes – Remove clothing and rinse body thoroughly in emergency shower
for at least 15 minutes. Seek medical attention. Notify supervisor and EH&S at x59797 immediately.
Chemical Splash Into Eyes – Immediately rinse eyeball and inner surface of eyelid with water from the
emergency eyewash station for 15 minutes by forcibly holding the eye open. Seek medical attention.
Notify supervisor and EH&S at x59797 immediately.
Medical Emergency Dial 911 or x52111
Life Threatening Emergency, After Hours, Weekends And Holidays – Dial 911 (or 310-825-1491 from
cell phone) or contact the Ronald Reagan UCLA Medical Center (emergency room) directly at x52111
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(located at 757 Westwood Plaza, enter from Gayley Avenue). Note: All serious injuries must be reported
to EH&S at x59797 within 8 hours.
Non-Life Threatening Emergency – Go to the Occupational Health Facility (OHF), x56771, CHS room
67-120 (This is on the 6th floor, 7th corridor, room 120. Enter through the School of Dentistry on Tiverton
Drive and proceed to the “O” elevator to the 6th floor.)Hours: M - F, 7:30 a.m. to 4:30 p.m. At all other
times report to Ronald Regan UCLA Medical Center (emergency room) at x52111. Note: All serious
injuries must be reported to EH&S at x59797 within 8 hours.
Needle stick/puncture exposure (as applicable to chemical handling procedure) – Wash the affected
area with antiseptic soap and warm water for 15 minutes. For mucous membrane exposure, flush the
affected area for 15 minutes using an eyewash station. Page the needle stick nurse by dialing 231 from a
campus phone, enter 93333 when prompted and then enter your extension. Hours: M – F, 8:00 a.m. to
4:00 p.m. At all other times report to Ronald Regan UCLA Medical Center (emergency room) at x52111.
Note: All needle stick/puncture exposures must be reported to EH&S at x59797 within 8 hours.
Decontamination/Waste Disposal Procedure
Lab coats must be decontaminated before they are removed for laundering. This may be accomplished
by washing the affected area in small container of soap and water. Dispose of the soap and water as
hazardous waste.
Laboratory work surfaces and equipment shall be decontaminated at the conclusion of each procedure
and at the end of each day. Use a soapy, wet paper towel to clean the affected areas and dispose of the
paper towel as hazardous waste.
General hazardous waste disposal guidelines:
Label Waste
 Affix an on-line hazardous waste tag on all waste containers using the WASTe Online Tag
Program https://ehs.ucop.edu/waste as soon as the first drop of waste is added to the container
Store Waste
 Store hazardous waste in closed containers, in secondary containment and in a designated
location
 Double-bag dry waste using transparent bags
https://www.ehs.ucla.edu/hazwaste/management/containers
 Waste must be under the control of the person generating & disposing of it
Dispose of Waste
 Dispose of regularly generated chemical waste within 90 days
 Call EH&S at x61887 for questions
 Empty Containers
o Dispose as hazardous waste if it once held extremely hazardous waste (irrespective of
the container size) https://www.ehs.ucla.edu/hazwaste/types/extremely-hazardous
o Consult waste pick-up schedule
https://www.ehs.ucla.edu/hazwaste/management/pick-ups
Prepare for transport to pick-up location
 Check on-line waste tag
 Write date of pick-up on the waste tag
 Use secondary containment
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Safety Data Sheet (SDS) Location
Online SDS can be accessed at http://msds.ehs.ucla.edu.
Protocol/Procedure (Add lab specific Protocol/Procedure here)
Quantities covered by this SOP:
0-200 ml
Conditions covered by this SOP:
<150°C – 30°C
Specific examples
1. Containers for cryogenic liquids must not be closed completely. Although liquid nitrogen are stored in
vacuum-jacketed vessels (dewars), there is always some heat leak into the dewar such that there is
boil-off of the liquid at all times. Pressure will build up and if no exit is available to the gas, the
container would ultimately explode. A pressure relief valve is attached to the dewar to keep the
internal pressure very low. Make sure you are aware of the presence of a protective pressure relief
valve on any dewar that you handle.
2. If a crack in the metal between the liquid and the vacuum space of a dewar were to occur, a rapid
pressure build-up in this relatively confined space will occur since it would no longer be insulated by
vacuum. For this reason, all metal dewars must be constructed such that a pressure relief valve or
rupture disk connected to the vacuum space will relieve excess pressure prior to sufficient build-up to
cause explosion. For the same reason, all glass dewars must be wrapped with tape or netting to
prevent flying glass in the event of an explosion.
3. Do not leave openings to cold dewars wide open to the atmosphere for any longer than is absolutely
necessary for the manipulations required for transferring liquids. The temperature of liquid nitrogen at
atmospheric pressure is -196°C. Air (and its contents) will condense into the dewar and can cause
blockages that are potentially dangerous and that will almost certainly interfere with some aspect of
the liquid transfer or with the operation of the instrument in the long run. The freezing point of liquid
oxygen is -183°C, i.e., above that of these cryogenic fluids. The collection of liquid oxygen into these
colder liquids is an explosion hazard!
4. The very cold temperatures of cryogenic fluids necessitate complete avoidance of contact with the
skin. Frostbite, i.e. “burns” from extreme cold, will occur very quickly upon contact, especially if
clothing, shoes, etc., hold the liquid tightly to the skin. Take care about spills and use appropriate
hand protection while transferring these liquids. Use thermally insulated gloves; thin gloves will not
help. Take care to avoid contact with the portions of the transfer line that have been inserted into the
dewars during the transfer as they will be cold enough to cause cold “burns” for quite a while after
removal from the dewars.
Flash Freezing Protocol:
 Put on proper Personal Protective Equipment (safety glasses, cryogenic gloves)
 Inspect the dewar and gloves for any signs of visible damage.
 If there are any signs of damage, discontinue work and report the problem to the supervisor.
 If there are no signs of damage, proceed with flash freezing by setting the dewar down in a stable
position and putting 20-200 ml liquid nitrogen in the dewar
 Use forceps or an equivalent extension tool to hold the sample in the dewar. (Never place hands
in the dewar).
 Never place the sample higher than your head when checking to see if the sample is completely
frozen.
 When finished, remove safety glasses and gloves and return to the designated storage location.
 Allow the liquid nitrogen to safety evaporate into a well-ventilated environment.
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Protocol for Cryopreservation of cells:
 Put on proper Personal Protective Equipment including safety glasses, and cryogenic gloves.
 Transfer cryovials of cells from -80C freezer to dry ice and put next to dewar for easy transfer to
liquid nitrogen.
 Locate box to transfer cryovials to in the liquid nitrogen inventory book.
 Place blue, rectangular ice bucket in the sink.
 Carefully remove rack from dewar. Be sure excess nitrogen is allowed to drain off back into the
dewar.
 Move rack to the ice bucket in the sink.
 Remove the appropriate box and place cryovials in the appropriate places.
 Return box to rack.
 Slowly and carefully return rack to the dewar.
 Pour excess liquid nitrogen from the ice bucket back into the dewar and return dewar lid.
 Be sure to update liquid nitrogen inventory book.
NOTE
Any deviation from this SOP requires approval from PI.
Documentation of Training (signature of all users is required)

Prior to conducting any work with liquid nitrogen, designated personnel must provide training to
his/her laboratory personnel specific to the hazards involved in working with this substance, work
area decontamination, and emergency procedures.

The Principal Investigator must provide his/her laboratory personnel with a copy of this SOP and a
copy of the SDS provided by the manufacturer.

The Principal Investigator must ensure that his/her laboratory personnel have attended appropriate
laboratory safety training or refresher training within the last one year.
Principal Investigator or Lab Supervisor SOP Approval
Print name_________________________Signature___________________________
Approval Date
I have read and understand the content of this SOP:
Name
Liquid Nitrogen
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Signature
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