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Microscopy & Analysis Suite
SAFETY BOOKLET
October 1993
Latest Revision May 2012
Microscopy & Analysis Safety Booklet
Page 1
1.
Introduction
2.
Working Practices
3.
Safety Information and Risk Assessment
4.
Specimen Preparation - Physical Sciences
4.1 Solvents
4.2 Acids
4.3 Electropolishing
4.4 Materialographic polishing and embedding
5.
Specimen Preparation - Life Sciences
5.1 Fixation
5.2 Dehydration and Embedding
5.3 Microtomy and Section Staining
5.4 Coating Grids and Negative Staining
5.5 Freeze Drying
5.6 Critical Point Drying
6.
Oil Hazards
7.
Liquid Nitrogen
8.
Vacuum Evaporation and Sputtering
9.
Lasers
10.
Emergency Procedures
11.
Working Outside Open Access Hours
Appendix A
Procedure in the Event of Contact With Hazardous Chemical
Appendix B
Plan of Centre for Electron Optical Studies
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1.
INTRODUCTION
This document has been prepared for the guidance of staff and students using the Microscopy &
Analysis Suite which includes the Bioimaging Suite (MAS) and should be read by all new users of
the facility. It outlines the working practices to be followed and gives information on the way in
which general and specific hazards are to be dealt with.
2.
WORKING PRACTICES
Training on equipment and specimen preparation procedures will be given to all new users and no
person will be permitted to use any part of the MAS until they have been accepted as
competent. Undergraduates will not be allowed to work unsupervised.
A wide range of biological and other materials are handled and prepared for microscopy in MAS
and it is essential that staff are made aware of any hazards arising from specimens received.
Consequently, before any work is undertaken, new users of MAS will be requested to register
details of their specimens and any hazards associated with them.
Laboratory coats are to be worn at all times in the specimen preparation laboratories (2.11 and
2.16F). When working in 2.11 & 2.16F be particularly careful about hygiene and ensure hands are
washed thoroughly on leaving (following removal of your laboratory coat).
Do not remove chemicals or apparatus from any of the laboratories without permission.
Before any programme of work is undertaken a member of the MAS Staff must be consulted and
the user should become familiar with any hazard by reading the appropriate sections of this
booklet and any other relevant data, see section 3.
Small spillages must be dealt with promptly by the perpetrator, but in the case of a major incident
the matter should be reported immediately to a member of staff and the laboratory evacuated.
Stored under the bench in 2.16F is a container of dresser-dri absorbent granules (suitable for
dealing with most spillages) together with sodium carbonate (for neutralising acid spills). Broken
glassware should be swept up and deposited in the yellow bin provided in 2.11, not in laboratory
waste bins.
Most equipment within MAS is of a commercial design with built-in fail-safe operation. There are
potential radiological hazards with the electron microscopes but regular checks have shown that
during normal operation dose rates are well within the acceptable limits. However, the
transmission electron microscope should never be operated with the condenser aperture
removed or with one or more of the condenser lenses switched off. Neither should they be used
with the camera boxes removed as these constitute some of the x-ray shielding.
In the transmission electron microscope a beryllium specimen holder can be used when x-ray
analysis is required. As beryllium is extremely toxic in powdered form care should be taken
that particulate material is not created and that it does not come into contact with an open
wound.
Any faults experienced on the electron microscopes or ancillary equipment should be reported to a
member of staff. Repairs are to be effected only by MAS staff, company engineers or in certain
Microscopy & Analysis Safety Booklet
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circumstances individuals who have been delegated to carry out a specific task. Non authorised
staff should not enter laboratories where the servicing of electron microscopes is taking place
because of dangers associated with exposed high voltage electrical circuits.
3.
SAFETY INFORMATION AND RISK ASSESSMENT
Before beginning any experimental work it is essential that the risks to health are fully assessed.
For many routine procedures e.g. electropolishing or fixation of biological tissue safety protocols
have been prepared but for some specialised activities this is not feasible. Then, in the first
instance, it is the responsibility of the person carrying out the experiment to assemble information
about the chemicals being used.
Available for consultation in the biological specimen laboratory are the "Electron Microscopy
Safety Handbook" and "Hazards in the Chemical Laboratory" together with a folder of safety data
sheets. In 2.16F there is a similar folder of data sheets relevant to chemicals used in preparing
specimens for the physical sciences. Another useful reference source is at
http://physchem.ox.ac.uk/MSDS/
Guidelines to help reduce risks are given below.
a.
Minimise the quantity of chemical being used. This is important when dealing with
chemicals where exposure limits are expressed in MELs (maximum exposure limits) and
OES (occupational exposure standards). Generally, limits are expressed in terms of the
safe exposure over an eight hour period but if a short term exposure (10 minute MEL) is
specified, efforts should be made to replace the chemical with a less harmful substitute.
b.
Consider the properties of any compound likely to be formed - is it more harmful than the
reactants themselves?
c.
Think about handling procedures - should gloves or goggles be used? - is a fume cupboard
required - are any special precautions necessary when transporting substances?
d.
How are waste products to be disposed of and what actions should be taken in the event of
a spillage?
e.
Are there any special first aid procedures you need to be aware of, should an accident
occur?
Having assembled all the facts discuss the experiment with your supervisor and a member of MAS
staff and if it is considered necessary a written risk assessment will be produced.
4.
4.1
SPECIMEN PREPARATION - PHYSICAL SCIENCES
Solvents
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Preparation and cleaning of samples prior to observation in scanning and transmission electron
microscopes routinely involves the use of solvents. Although the quantity of solvent used is
generally small it is important that certain basic precautions are taken.
Many solvents are extremely flammable and can form explosive mixtures in air. Furthermore, they
may react violently with certain common materials. The following protocol must therefore be
followed at all times.
a.
Look up the data sheet of the solvent you are proposing to use and ensure that it will not
react violently with any material with which it will be in contact.
b.
If the solvent is flammable ensure that there are no naked flames in the laboratory or any
other source of ignition.
c.
Use gloves and glasses or goggles for the eyes - contact with solvents can cause drying of
the skin and dermatitis, whilst solvents such as acetone can cause severe damage if
splashed into the eyes.
d.
Remove flammable solvents from the yellow, fire-proof cabinet one at a time and return
them immediately after use, making sure that the cabinet doors are closed afterwards.
e.
To avoid breathing harmful vapours pour out volatile solvents in the fume cupboard.
f.
Make every effort to minimise the quantity of solvent on a bench at any given time.
g.
Use solvents immediately where possible. Do not leave beakers on benches. The fume
cupboard may be used for temporary storage.
h.
If using the ultrasonic cleaning bath the solvent should be contained in 100 ml or 250 ml
beakers. During use the bath lid should be in place and the laboratory door closed to reduce
noise.
i.
Do not pour solvents down sinks: Dirty solvents should be poured into the waste
containers stored in the fume cupboard (2.16F). There are containers for chlorinated and
non-chlorinated solvents. Make sure that you use the appropriate one. For your guidance
the list below includes the majority of solvents stored in 2.11 and/or 2.16F
Non-Chlorinated (Non-Halogenated) Solvents
Acetone (CH3)CO
Ethanol C2H5.OH
Isobutyl Methyl Ketone - alternative name 4-methylpentan-2-ONE (CH3)2CH.CH2CO.CH3
Methanol CH3OH
Diethyl Ether (C2H5)2O
Methylated Spirits
Propan-2-ol (Alcohol)
Propylene Oxide
Hexamethyl Di-Silizane
Chlorinated Solvents
Chloroform CHC3
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If a major spillage occurs or there is a strong smell of solvent leave the laboratory and inform a
member of staff immediately. A minor spillage may be mopped up with absorbent tissues (used
tissues being placed in the fume cupboard).
In case of solvent contact with skin or eyes see Appendix A
4.2.
Acids
The acids stored in 2.11 and 2.16F, mostly in concentrated form, are:
Acetic
Hydrochloric
Orthophosphoric
Nitric
Sulphuric
Use protective clothing at all times when handling acids - wear goggles or glasses to protect the
eyes and rubber gloves to ensure that acids do not come in contact with the skin.
Look up the data sheet of the acid you intend to use and make sure the acid does not react violently
with any other chemical involved in the work.
When diluting sulphuric acid always add the acid to water.
Acids are stored separate from other chemicals. Do not leave Winchesters of acid on benches.
Although used in the past, perchloric acid is not now recommended for use for electropolishing,
except in a specialized fume hood with a wash-down system to prevent the build-up of explosive
perchlorates in the ductwork. As the fume cupboards in the MAS laboratories are not of this type,
perchloric acid should not be used in the laboratory, particularly at concentrations above
72.5%. This acid can be highly explosive and there is a long history of accidents associated
with it. Alternative methods of specimen preparation should be employed.
In the event of minor spillages of the following acids - Hydrochloric; Orthophosphoric, Nitric or
Sulphuric - spread soda ash (sodium carbonate) liberally over the spillage and mop up with plenty
of water. In the case of Acetic acid absorb on dresser-dri absorbent - shovel into a bucket and
remove from the laboratory.
If the spillage is serious or if there are noxious fumes leave the laboratory closing the door behind
you and inform a member of staff immediately.
In case of acid contact with skin or eyes see Appendix A
4.3.
Electropolishing
Inform MAS staff before attempting electropolishing. Chemicals used in electropolishing can be
dangerous unless certain rules are strictly adhered to. Users of electropolishing equipment in the
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MAS Laboratory must acquaint themselves with the properties of all the chemicals that they use
and know the remedial first aid in case of accidents. Data sheets are available in the laboratory
outlining hazards associated with commonly used solutions. Be sure to ask for this information
before any solutions are made up.
The electropolishing equipment, the 'Tenupol', is manufactured by Struers Ltd and we strongly
recommend that only the Struers solutions are used. Electropolishing solutions other than those in
the Struers range should not be made up without prior consultation with a member of the MAS
staff. Only after a thorough examination of the risks involved will permission be given for a nonstandard solution to be used.
When mixing polishing solutions always wear protective nitrile gloves. Either a plastic face guard
should be employed or the solution should be made up in a fume cupboard with the window
partially closed. Any electropolishing solution that contains a strong oxidising agent eg nitric acid
mixed with an organic solvent such as methyl alcohol can explode during mixing if the exothermic
oxidation reaction is allowed to accelerate. The following rules should be observed:
a.
To prevent overheating, reagents should be cooled with running water and the mixing
should be slow with effective stirring. Use a thermometer to monitor the solution's
temperature.
b.
Never put warm or newly mixed solutions in tightly stoppered or screwtop bottles.
c.
Except where specified otherwise, the oxidising agent should be added to the reducing
agent.
d.
Methyl alcohol is preferable to ethyl alcohol if used with nitric acid. The latter forms
more
unstable solutions which can decompose violently. After approximately one
month, or when they have become contaminated with metal ions that may act as a
catalyst, even methyl alcohol - nitric acid solutions should be discarded. There is also a
hazard with methyl alcohol solutions if evaporation losses are allowed to occur.
Consequently they should be placed in stoppered bottles.
e.
Care should be taken when mixing sulphuric acid and water because of the heat
generated.
Always add acid to water.
Operation of the electropolishing unit is detailed in the TENUPOL manual which should be
carefully studied before use. The TENUPOL is designed to operate at temperatures down to -50°C.
If sub-ambient temperatures are required the electrolyte may be cooled by careful addition of small
amounts of liquid nitrogen. Use a small Dewar to decant the liquid nitrogen and monitor the
temperature in the tank with a thermometer. Close the fume cupboard door after each liquid
nitrogen addition to ensure efficient removal of volatiles. Normal precautions for liquid nitrogen
use must be observed.
When electropolishing is completed, ensure that the TENUPOL is thoroughly flushed through with
water. Electrolyte needed for future use may be stored in stoppered bottles (if clearly labelled with
the contents, date of mixing, hazards etc.) and placed in the appropriate cupboard. Electrolyte
disposal should be discussed with a member of staff to determine the most suitable method.
Containers exist for temporary storage of harmful liquids not suitable for running directly to waste.
If in doubt about any of the safety information described, always ask. A member of the MAS staff
will answer any questions you may have.
4.4
Materialographic polishing and embedding
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Hazards associated with the process are chemical, please ensure you have read the MSDS for the
chemicals and mounting medium you intend to use. Polishing involves the use of fine nanoparticle
diamond or alumina in an liquid carrier and if the solutions becomre dry they are an irritant. Please
wear dust masks provided. If your material is toxic make sure MAS staff are aware so precautions
can be taken. Please refer to the life science section for more information on the embedding
process and the chemical hazards.
5.
SPECIMEN PREPARATION - LIFE SCIENCES
The preparation of biological specimens for scanning or transmission electron microscopy
involves the use of many hazardous chemicals and MAS staff must be informed before specimen
preparation is attempted. Preparation consists of stages at which different chemicals and
procedures are used, for example: fixation - dehydration - drying or embedding. These stages are
listed below with details of the hazardous chemicals involved. The majority of steps in the
preparation procedure are carried out within a fume hood and therefore any spillage of hazardous
chemical is likely to occur within the fume hood minimising exposure to it. The wearing of a lab.
coat and disposable nitrile gloves is mandatory when preparing biological samples. Eye protection
is necessary with certain chemicals, and this is indicated below.
After use of a hazardous chemical within a fume hood it is essential that any items that have come
into contact with the chemical such as - glassware, pipettes, syringes and filters, paper towel, or
gloves are thoroughly rinsed under running water in the fume hood sink before removal from the
fume hood for:
(1) washing and re-use (place in bucket under sink)
(2) disposal in - (a) laboratory waste bins (plastic pipettes, syringes, filters, paper towel, gloves),
(b) sharps bin (syringe needles, razor blades).
(c) yellow glass bin ( broken glassware).
Action to be taken in the event of a minor spillage of a hazardous chemical within a fume hood
is outlined for each chemical below.
In the event of a spillage of hazardous chemical outside the fume hood the action is the same in
all cases - evacuate the laboratory, shut the door and inform MAS staff immediately.
5.1.
A.
Fixation
Buffers
Routinely used buffers are phosphate, cacodylate and PIPES. These are used to dilute fixatives and
to wash tissue, and will usually be made as solutions of 0.4M concentration (or less). As far as is
known, neither phosphate or PIPES buffer constitute a health hazard; sodium cacodylate buffer
does, however, as it is an arsenic compound (skin absorbed, cumulative toxin). This chemical
should be handled as follows:
1. Make sure laboratory door is closed to minimize air currents.
2. Weigh sodium cacodylate powder on the balance in fume hood - minimize draft by opening
cupboard to arrows.
3. Add powder to volumetric flask using a funnel and water in the fume hood.
4. All subsequent use of sodium cacodylate buffer is carried out in a fume hood. A lab coat,
gloves and eye protection should be worn.
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5. Waste sodium cacodylate is collected in the labelled waste bottle (at back of fume hood) for
disposal by the University Waste Service.
6. The pH of the buffer solution requires alteration using an acid solution. Set up the pH metre
inside the fume hood and refer to section 4.2 for details of the safe handling of acids.
Cacodylate buffer: Toxic by inhalation, absorption and if swallowed. Prevent contact with skin
and eyes. Poison.
MEL 0.1mg.m-3
In case of contact with skin or eyes see Appendix A
Action in event of minor spillage within the fume hood: Wear a lab. coat, gloves and eye
protection. Sodium cacodylate may be mopped up with paper towelling which is then rinsed with
plenty of water in the fume hood sink. Flush contaminated area with plenty of water. Rinse any
contaminated items (e.g. glassware, or gloves) before removing from the fume hood.
B.
Fixatives
Fixatives are diluted in buffer to concentrations of below 10%. The most commonly used fixatives
are glutaraldehyde, formaldehyde and osmium tetroxide. Acrolein (acrylaldehyde) and
ruthenium tetroxide are used only as a last resort - when samples are not adequately fixed by the
other chemicals. They are used only under supervision. All fixatives must be used in the
designated fume hood. Lab. coats and nitrile gloves must always be worn and, in addition, eye
protection when working with osmium tetroxide, ruthenium tetroxide and acrolein. Waste fixatives
are collected in labelled waste bottles within the fume hood and disposed of by MAS staff or the
University Waste Service.
Glutaraldehyde: Toxic by inhalation & ingestion. Causes burns. Sensitisation by inhalation &
skin contact. Gluteraldehyde can cause respiratory & skin allergies. Sensitised individuals may
suffer reactions upon exposure to even small quantities leading to dangerous asthma attacks.
See University Occupational Asthma leaflet & poster.
Very toxic to aquatic organisms. Dangerous to the environment
In case of vapour inhalation or contact with skin or eyes, see Appendix A.
Action in event of minor spillage within fume hood: Wear a laboratory coat, blue nitrile gloves
and eye protection. Mop up spillage with paper towel, and rinse towel & flush area with plenty of
water from the sink in fume hood. Rinse well any contaminated items ( glassware, pipettes, gloves)
before removing from the fume hood.
Formaldehyde: Harmful by inhalation and if swallowed. Avoid contact and vapour. Mutagen,
teratogen, possible carcinogen. May cause dermatitis. MEL 2.5mg m-3.
In case of vapour inhalation or contact with skin or eyes, see Appendix A.
Action in event of minor spillage within fume hood: Wear a laboratory coat, nitrile gloves and
eye protection. Mop up spillage with paper towel, and rinse towel & flush area with plenty of
water from the sink in fume hood. Rinse well any contaminated items ( glassware, pipettes, gloves)
before removing from the fume hood.
Acrolein (Acrylaldehyde): Very toxic by inhalation, in contact with the skin and if swallowed.
Acrolein is a poison - prevent inhalation of vapour, contact with eyes and skin. Lachrymator, OES
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0.25mg. m-3. Causes burns. In case of vapour inhalation or contact with skin or eyes, see Appendix
A.
N.B. Acrolein is extremely flammable - cool bottle to 0oC before opening inside fume hood should only be used under supervision.
Action in event of minor spillage within fume hood:. Wear a laboratory coat, blue nitrile gloves
and eye protection. Pipette the spilt fixative into the waste bottle if possible or mop up with paper
towel, and flush area with plenty of water from the fume hood sink. Thoroughly rinse with water
any items (towel, pipettes, gloves, glassware) that have come into contact with acrolein before
removing them from the fume hood!
Osmium tetroxide: Very toxic by inhalation, in contact with skin and if swallowed. Causes
burns. Prevent inhalation of vapour, and contact with eyes and skin. Mutagen and teratogen, OES
0.002 mg.m-3. A solution of osmium tetroxide and its vapour will rapidly fix lipid in biological
tissues which turn black. This will happen to skin or the cornea of the eye if they are exposed to it.
In case of vapour inhalation or contact with skin or eyes, see Appendix A.
Action in event of minor spillage within fume hood: Wear a laboratory coat, nitrile gloves and
eye protection. Pipette the spillage into the waste osmium tetroxide bottle if possible, or mop up
spillage with paper towel, and flush area with plenty of water from fume hood sink. Thoroughly
rinse with water any items (pipettes, paper towel, gloves, glassware) that have come into contact
with osmium tetroxide before removing them from the fume hood.
5.2
Dehydration and Embedding
Fixed samples of tissue are dehydrated using a graded series of acetone or ethanol before
embedding in either epoxy or acrylic resin. Propylene oxide is occasionally used as an
intermediate agent between ethanol/acetone and epoxy resin.
Acetone: Highly flammable. Prevent contact with eyes. Avoid breathing vapour.
OES 1210 mg.m-3.
Ethanol: Highly flammable. Avoid breathing vapour in high concentrations. OES 1900 mg.m-3.
Use both solvents in a fume cupboard or extraction hood where possible. Always keep solvents in
the solvent cabinet and remove only briefly for use. Do not use acetone near any possible sources
of ignition. Waste non-chlorinated solvents should be placed in the waste bottle provided in the
fume hood in lab 2.15F.
In case of eye contact, see Appendix A.
Action in event of a spillage: Shut off possible sources of ignition. Wear laboratory coat, gloves
and eye protection. Mop up small amounts with paper towel and place these in a fume hood to
allow solvent to evaporate. Ventilate area well. For a large spillage producing noxious fumes
evacuate the laboratory closing the door behind you and inform MAS staff.
Propylene Oxide: Extremely flammable. Toxic by inhalation, in contact with skin and if
swallowed. Avoid breathing vapour. Avoid contact with skin and eyes. Mutagen, teratogen and
carcinogen, OES 50mg.m-3. Always open the bottle and use in a fume cupboard, wearing lab. coat,
blue nitrile gloves and eye protection. Pipette waste into non-chlorinated solvent bottle.
In case of vapour inhalation or contact with skin or eyes, see Appendix A.
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Action in event of minor spillage within fume hood: Shut off all possible sources of ignition.
Wear lab. coat, blue nitrile gloves and eye protection. Pipette spillage into waste non-chlorinated
solvent bottle if possible, or mop up with paper towel. Wash area well with plenty of water and
detergent. Rinse well with water and detergent any items (pipettes, gloves, paper towel or
glassware) that have come into contact with propylene oxide before removing from the fume
hood.
Action in event of a spillage outside fume hood: For a large spillage producing noxious fumes
evacuate the laboratory closing the door behind you and inform MAS staff.
Resins: Are used to embed fixed and dehydrated tissue in order to facilitate sectioning. A wide
range of resins is available - acrylic resins (LR White, Technovit, Lowicryl) - epoxy resins (Taab
Premix, Durcupan etc.).
Resin mixtures cause dermatitis and allergic reactions on inhalation or if they come into contact
with the skin and they may also depress the central nervous system causing headaches. Resin
accelerators are toxic.
When handling resins a lab. coat and gloves should be worn. Open resin bottles only in the
designated fume hood. Resins are heat polymerised in the oven situated within the fume hood
only. Use disposable plastic beakers with resin as it is difficult to remove resin from glassware!
Items that become contaminated with resin such as pipettes, paper towels or gloves are placed in
the wire basket provided within the fume hood. Resin-contaminated gloves must be removed
before handling anything outside the fume hood. Waste resin collected in plastic beakers is
placed in the tray provided at the back of the fume hood to await polymerisation before disposal by
MAS staff.
Any accidental spillage of resin onto the skin may be removed using a viscous resin cleaner found
by the sink; do not remove using a solvent as this drives the resin into the skin.
Spillage disposal: Bottles of resin mixtures are only to be opened and used within the fume hood
and as such a spillage should be minor. Wear a laboratory coat, safety glasses and blue nitrile
gloves. Where possible pipette any spillage into a disposable plastic beaker and clean a small area
within the fume hood with paper towels and acetone. For large spills, close fume hood window and
inform MAS staff.
5.3
Microtomy and Section Staining
Polymerised blocks are cut down with razor blades prior to sectioning using an ultramicrotome
fitted with a glass knife. Care should be taken when using razor blades or when handling glass. A
red plastic guard is available for use with single-edge razor blades to protect fingers, and cut
resistant gloves are also available.
A Sharps bin is provided next to the microtomes for disposal of razor blades, and a yellow bin
under the bench for disposal of glass knives and slides only.
Hardened resin is thought to be less harmful than its non-polymerised components, but care should
be taken to keep the microtome clear and free from resin chips. Filing and sawing of resin blocks
should also be kept to a minimum and face masks are provided to prevent inhalation of resin dust.
Resin chips and dust are cleaned up with damp paper towelling and placed in a laboratory bin.
Semi-thin sections are taken from blocks and stained with toluidine blue or epoxy tissue stain for
study with the light microscope. Heating sections in stain must be done on a hot plate inside a
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fume hood.. Wear gloves and a lab coat. Waste stain solution is placed in the waste bottle in the
fume hood
Toluidene blue & Epoxy Tissue Stain: Harmful by inhalation, in contact with skin and if
swallowed. Avoid breathing vapour. Avoid contact with skin and eyes. Exposure limits not
assigned.
Methylene blue: Harmful by ingestion. Stains may irritate eyes and skin. Avoid contact with skin
and eyes. Exposure limits not assigned.
In case of contact with skin, eyes or vapour inhaled see Appendix A.
Minor Spillage disposal within the fume hood: Wear a laboratory coat and gloves. Pipette the
spillage into the waste bottle and clean the area with plenty of water.
Staining.
Thin sections and particles are mounted on grids and stained with heavy metal stains. Ammonium
molybdate, methylamine tungstate, lead citrate, uranyl acetate and phosphotungstic acid. These
stains are made up as 1-6% solutions and normally used in small amounts. The greatest hazard
from heavy metal stains is breathing dust into the lungs. Wear laboratory coat and blue nitrile
gloves when performing staining routines. Do not allow these stains to dry out as the resulting dust
may be dispersed by air currents. Weigh heavy metal stain powders on balance in the fume hood –
open door to arrows to reduce draft. Uranyl acetate is radioactive as well as very toxic. It is stored
in a metal tin in the fume hood. Pick up the bottle of uranyl acetate using fingertips to decrease
exposure of hand to radioactivity. Waste or used uranyl acetate solution is placed in the labelled
waste bottle. Items that come into contact with stains such as - glassware, wax, pipettes, syringes
and filters, or gloves should be thoroughly rinsed in running water before removal from the fume
hood.
Lead citrate: Toxic by inhalation, absorption and if swallowed. Danger of cumulative effects.
Avoid breathing dust. Mutagen, and teratogen, MEL 0.15 mg.m-3.
In case of contact with eyes or skin: wash thoroughly with plenty of water.
Uranyl acetate: Very toxic by inhalation and if swallowed. Danger of cumulative effects. Avoid
breathing dust or vapour. Mutagen, terratogen and carcinogen, OES 0.2 mg.m-3. Radioactive.
In case of contact with eyes or skin, or inhalation of dust see Appendix A.
Minor spillage disposal within a fume hood: Wear lab. coat, gloves and eye protection. Pipette the
spilt stain into the waste bottle if possible, and/or mop up with paper towel flushing the area well
with water. Rinse thoroughly any contaminated items (glassware, paper towels, gloves) before
removing from fume hood.
5.4
Coating Grids
Grids are coated with plastic dissolved in chloroform for casting thin films. The main hazards
arise from the use of chloroform to dissolve the plastic and the initial casting of the films onto
glass slides in a chloroform vapour environment. All procedures should be performed in the
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extraction hood. Waste chloroform is placed in the chlorinated waste solvents bottle stored in the
fume hood in laboratory 2.16F.
Chloroform (trichloromethane): Harmful by inhalation, absorption or ingestion. Avoid
breathing vapour and contact with eyes. Suspected carcinogen, mutagen & teratogen. Serious
damage to health may result from prolonged exposure. OES 50 mg.m-3.
In case of contact with eyes or inhalation, see Appendix A.
Minor spillage disposal within fume hood: Wear laboratory coat, eye protection and gloves.
Pipette spillage into appropriate waste bottle, if possible, or mop up with paper towels. Site of
spillage should be washed thoroughly with water and soap or detergent. Rinse thoroughly any
contaminated items (glassware, pipettes, paper towel and gloves) before removing from the fume
hood.
Action in event of a spillage outside fume hood: For a large spillage producing noxious fumes
evacuate the laboratory closing the door behind you and inform MAS staff.
5.5
Freeze drying
The main hazard encountered here is the use of liquid nitrogen for initial freezing. Eye protection
should always be worn when handling liquid nitrogen. These are provided next to the liquid
nitrogen container. Always use correct Dewars for dispensing liquid nitrogen ( see section 7).
Never use in a confined space.
In case of contact with skin or eyes seek medical advice to deal with resulting burns. In case of
inhalation of vapour see Appendix A.
Phosphorus Pentoxide: Used as a drying agent in the freeze dryer. Causes severe burns. Prevent
inhalation of dust or contact with eyes and skin. Violent exothermic reaction with water. To clean
the metal phosphorus pentoxide holder after use - place in fume hood sink and flush with running
water until reaction is complete and all chemical is removed. Dry container thoroughly with paper
towels before recharging with phosphorus pentoxide in the fume hood. Exposure limits not
assigned.
In case of contact with eyes or skin, or if dust inhaled, see Appendix A.
Minor spillage disposal within fume hood: Wear lab. coat, eye protection and gloves. Add a small
quantity at a time to a large volume of water. When reaction complete, run to waste diluting
greatly with running water. Wash site of spillage with water.
NOTE: The freeze dryer operates under vacuum and the implosion guard must always be placed
over the bell jar when it is evacuated.
5.6
Critical Point Drying
Specimens dried using this technique are first fixed and dehydrated (see sections on fixation and
dehydration). They are subsequently placed in the critical point drying apparatus in 100% acetone.
Pressures of up to 14000 lb/u2 are attained using this apparatus, which must never be left
unattended during the heating process otherwise substantially higher pressures may be generated.
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Following drying, the specimen is mounted on stubs or planchettes using a conducting graphite
paste. These pastes contain a solvents normally iso-propyl alcohol and should be handled with
care in a fume cupboard, if possible.
Propan-2-ol (iso-propyl alcohol): Flammable. Irritant - irritating to eyes.
OES 99 mg.m3
6.
OIL HAZARDS
Various types of oil are used extensively throughout the laboratories as vacuum pumping fluids
and as lubricants. Oils and greases should not, in general, need to be handled by students nor
should they pose any significant health and safety hazard. However, they may cause dermatitus
and prolonged or repeated contact with the skin should be avoided by using impervious plastic
gloves or by washing ones hands thoroughly after use.
Oil mist fumes from vacuum pumps present a more serious problem in that they may be
carcinogenic or cause irritation of the lungs. Every effort has been made to eliminate such fumes
in the laboratories by using the University's extract system to remove vapour from rotary pumps or
to fit the pump with an oil mist filter. If oil vapour is ever smelt in the laboratories it should be
reported immediately to a member of the MAS Staff.
7.
LIQUID NITROGEN
Liquid nitrogen is widely used as a cryogen for energy dispersive detectors, anti-contaminators and
for cooling specimens. If liquid nitrogen comes into contact with the skin cold burns will result. It
is therefore essential that eye protection is used. Insulating gloves are available for dispensing
liquid nitrogen.
Follow the instructions given below.
Do not dispense liquid nitrogen into domestic thermos flasks, they are not designed to withstand
the thermal shock that will be encountered. Use only MAS Dewars.
Do not place Dewars containing liquid nitrogen on the edges of benches or in exposed positions on
the floor where they may be knocked over.
Remember not to touch metal objects cooled by liquid nitrogen as you may suffer frost bite.
Never attempt to pressurise a liquid nitrogen container.
If a major spillage of liquid nitrogen occurs in a confined space evacuate the laboratory and allow
nitrogen gas to disperse.
Do not travel with liquid nitrogen in a lift. If a spillage occurs in a confined space you may be
overcome by oxygen starvation - such a situation is potentially life threatening.
8.
VACUUM EVAPORATION AND SPUTTERING
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The glass bell jar of a vacuum evaporator or sputter coater may implode when
(a)
there is a flaw or crack in the glass
(b)
struck by a hard object
(c)
excessive heat is generated within the chamber by the evaporation source.
On implosion glass may scatter in all directions. Consequently a metal or perspex safety shield
must always be fitted over the bell jar before evacuation is commenced.
Handle the glass bell jars carefully at all times and when they are removed from the coating units
place them in a safe place where they will not be knocked over.
Never look directly at the source during evaporation, the intensity of light may cause eye damage.
Use the welding goggles provided.
9.
LASERS
Three continuous wave, helium-neon lasers are used in the laboratories and all of these are low
power class 2 lasers. Two of them are used to terminate the ion beam in the ion milling equipment
and the third is fitted to the Nanoscope III SPM. None of them are likely to be harmful provided
that the laser beam or laser light scattered from shiny surfaces does not enter the eye. Because of
the design this should not occur but never look directly into the path of a laser beam. All of
these lasers are sealed units and in the event of malfunction must be returned to the manufacturer
for repair.
In the Bio-imaging suite, the two confocal microscopes utilise Class 3B lasers. This laser class is
not covered under the standard Generic Risk Assesment procedures. Individual training in the safe
and effective use of these instruments is mandatory and a Special Hazard Assessment record
sheet must be completed (see MAS staff). As with the lower power lasers mentioned above, it
should not be possible to look directly into the path of the laser beam. These microscopes also
utilise Ultraviolet light bulbs, and again it should not be possible to look directly at these sources.
10.
EMERGENCY PROCEDURES
Fire
In the event of a fire break the glass in the fire alarm point situated outside MAS on the wall at the
head of the stairs. When at a place of safety call Security on x666 and describe the problem.
There are three fire extinguishers within MAS, at either end of the main corridor and in the
Physical Science Preparation Laboratory (2.16F) immediately inside the door on the right hand
side. All are CO2 extinguishers and should not be directed at a person's body. Extinguishers
containing water are provided just outside the main MAS doors. Instructions for use are given on
the extinguishers themselves. A fire blanket is fitted to the wall of the corridor and can be used to
smother burning clothing. All these items are marked on the plan in Appendix B.
Exiting the building
In an emergency you should leave MAS immediately. There are three exits:
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


Along the MAS corridor, out of main doors, through the 3 West foyer and out to
south side of University.
Through the door marked EMERGENCY EXIT at one end of the MAS corridor, into
Physics & left into corridor and exit to the north side of building.
From Bioimaging Suite through 2.16D and out through the window fire escape;
Routes are detailed in Appendix B
First Aid
First Aid Boxes are available in the Life Science Preparation Laboratory (2.11) and the Physical
Science Preparation Laboratory (2.16F). A shower unit is fitted to the sink in 2.16F for cases
where a major spillage of acid or other chemical has occurred on a person's body or clothing. A list
of first aiders in 3W is displayed on the notice board outside room 2.15A .
11.
WORKING OUTSIDE OPEN ACCESS HOURS
It is recognised that on occasions it may be necessary to operate equipment outside open access
hours (9:10am - 5.15pm Monday - Friday). In this case permission must be sought from a member
of MAS staff. When entering and departing from building 3 West the individual is required to sign
the register available in 3 West foyer. On leaving, equipment must be left in a safe condition.
A security system operates on the main MAS doors outside normal working hours (before 09:10
and after 17:10) – the doors will lock automatically and access is only available with a registered
swipe card. To exit MAS a ‘door release’ button is located to the right of the main doors and will
release the lock.
Working outside open access hours will only be permitted if it is considered that there is
insufficient instrument time available during the day or an overnight experiment is to be
performed. The potential user must be an experienced and responsible operator of the equipment.
Access to specimen preparation laboratories will not normally be permitted unless a member of the
MAS Staff is available to supervise.
Anyone working abnormal hours should familiarise themselves with the emergency shut-down
procedure for the item of equipment they are operating, with the location of the nearest emergency
telephone and with fire exit locations. At least two people must be present at all times if work of a
hazardous nature is being undertaken.
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APPENDIX A
PROCEDURE IN THE EVENT OF CONTACT WITH HAZARDOUS CHEMICALS
In Case of Skin Contact:
1.
Flood the splashed surface thoroughly with a large quantity of running water, and continue
for at least 10 minutes, or until satisfied that there is no chemical left in contact with the
skin. Clean up with soap. Note a shower unit is fitted to the sink in 3W 2.16F
2.
Remove all contaminated clothing, taking care not to further expose the skin in the process.
3.
Report the incident to a First Aider and if necessary refer for medical advice to the medical
centre; telephone 6655. Provide as much information on the chemical responsible as is
possible. In serious cases make an emergency call
In Case of Eye Contact:
1.
Flood the eye thoroughly with sterile water from the bottles available next to the First Aid
boxes in 2.15 & 2.16F.
2.
Ensure that water bathes the eyeball by gently prising open the eyelids and keeping them
apart until the treatment has been completed.
3.
Inform a First Aider as soon as possible and supply as much information on the chemical
responsible as you can. All eye injuries from chemicals require medical advice. In serious
cases make an emergency call.
Inhalation of Gases:
1.
Remove the casualty out of the danger area (having first ensured your own safety).
2.
Seal off the area affected if possible.
3.
Inform a First Aider immediately and provide them with details of the gas responsible. In
serious cases make an emergency call
EMERGENCY CALLS
In any case of accident requiring urgent medical attention dial 666 from any University
telephone. Give the operator your name and extension number, the location of the injured
person and, if possible, brief details of the nature of the injury.
REPORTING ACCIDENTS
All accidents occurring in the MAS should be reported to the Technical Manager who will arrange
for an accident report form to be completed.
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