Sterilization and Disinfection 
Chris Gross, Sanaz Dovell 
 
OPEN MANUAL OF SURGERY IN RESOURCE-LIMITED SETTINGS 
www.vumc.org/global-surgical-atlas 
This work is licensed under a Creative Commons Attribution-ShareAlike 3.0 Unported License  
 
INTRODUCTION:  
This chapter will cover the topics of 
decontamination, disinfection, and sterilization of 
medical items, instruments and equipment. Due to its 
broad (and non-sequential) nature, we will break this 
chapter into four major parts: terms, theory, 
tools/techniques and practices. Our hope is that this 
overview will set a foundation for understanding the 
basics 
of 
decontamination, 
disinfection 
and 
sterilization. Note there is intentional redundancy 
within this chapter, as with other chapters within the 
Manual itself. 
 
Scope of problem:  
1. Safe surgery requires a sterile procedure to 
minimize life-threatening infections. Sterile 
procedures require sterile instruments.  
2. Global access to safe surgery is limited by a 
lack of access to sterilization.  
3. Resource-constrained settings may not have 
reliable access to reliable electricity, making 
the use of the autoclave, the WHO standard 
for sterilization, difficult or impossible. 
4.  Staff responsible for cleaning and sterilizing 
the surgical equipment may have limited 
training in sterile processing techniques.  
 
Impact of problem:  
1. Approximately five billion people around the 
world do not have access to much-needed 
surgical care, and as much as 33% of 
worldwide deaths are from surgically 
treatable conditions. 
2. Lack of proper instrument sterilization has 
led to post-surgical infection rates as high as 
46%.  
3. Disposable kits, often thought of as a fix to 
the problem, are not a viable solution because 
they generate large amounts of biowaste that 
is difficult to properly dispose of in low-
resource areas. 
 
TERMS: 
1. Cleaning is defined as removing any visible 
soil.  Cleaning is typically achieved by manually 
combining water with enzymes or detergents and 
using a brush to physically remove visible debris. 
It is important to note that clean does not mean 
sterile or disinfected. 
2. Decontamination is defined as removing 
pathogenic organisms in order to make objects 
safe enough to handle, use, or dispose of. 
3. Disinfection is the process of eliminating most 
pathogenic organisms with the exception of 
spores. This is usually accomplished with liquid 
chemicals or wet pasteurization. 
4. High-level disinfection is defined as the 
complete elimination of all microorganisms in or 
on an instrument, with the exception of a small 
number of bacterial spores. 
5. Intermediate-level 
disinfection 
is 
the 
elimination of all microorganisms, but not 
spores. 
6. Low-level disinfection kills fungi, most bacteria, 
some viruses, and no spores. 
7. Sterilization is the process of eliminating all 
microorganisms, and is commonly achieved 
through chemical and/or physical means. 
8. Germicide is an agent that can eliminate 
microorganisms. 
Agents 
with 
‘cide/cidal’ 
suffixes have some type of killing action that is 
referenced in the name (i.e fungi-cide). Note that 
germicides 
include 
both 
antiseptics 
and 
disinfectants (remember disinfectant differs 
from disinfection). 
a. Antiseptic germicides are agents that can 
be applied to living tissue (i.e skin). 
b. Disinfectant is any germicide agent that 
can be applied to non-living tissue (i.e. 
not skin). It is only used to disinfect 
surfaces of inanimate objects because of 
the risk of injury to tissue. 
9. Critical items are any items with a high chance 
of 
transmitting 
an 
infection 
if 
that 
instrument/item becomes contaminated (surgical 
instruments, urinary catheters, implants). These 
items must be completely sterile. 
10. Semi-critical items are any items/instruments 
that come into contact with non-intact skin and/or 
mucous membranes (respiratory / anesthesia 
equipment). These items should undergo 
cleaning, followed by high-level disinfection.  
Sterilization and Disinfection 
Chris Gross, Sanaz Dovell 
 
OPEN MANUAL OF SURGERY IN RESOURCE-LIMITED SETTINGS 
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11. Non-critical items are any items/instruments 
that come in contact with intact skin and do not 
come in contact with mucous membranes 
(bedpans, computers, patient furniture). It is not 
critical for these items to be sterile as long as they 
do not come in contact with non-intact skin or 
mucous membranes. 
 
THEORY: 
Before diving into the individual techniques 
for achieving sterility, it is important to discuss the 
factors that influence our ability to get an item clean, 
decontaminated, disinfected, and/or sterilized. We 
will divide these factors into those that are internal to 
the offending organism and factors that are external 
to the offending organism.   
 
Factors internal to the organism 
Organism type: The first major internal 
factor is the type of microorganism(s) present on the 
object and whether or not spores are involved. The 
major 
types of organisms to consider for 
disinfection, listed in order of most-to-least difficult 
to 
eradicate, 
are 
prions, 
bacterial 
spores, 
mycobacteria, non-lipid viruses, fungi, bacteria, and 
lipid viruses. Each type of organism has a varying 
level of susceptibility to destruction, which is related 
to its physical structure (i.e lipid envelope or protein 
capsid). For example, spores are generally resistant 
to most disinfectants while mycobacteria are 
generally resistant to alcohol, and prions are resistant 
to heat at 200 oC for as long as two hours.   
Organism quantity: Just as microbes grow at 
an exponential rate, microbes also die at an 
exponential rate.  As the number of organisms 
increases, a longer exposure time is required to 
achieve disinfection or sterility. Proper cleaning with 
water and detergent or enzymatic cleansers prior to 
disinfection or sterilization can help to reduce the 
microbial load and increase the effectiveness of the 
disinfectant or sterilant.  
Most bacteria create a biofilm on solid 
organic or inorganic surfaces when the number of 
organisms reaches a certain level. The biofilms 
contain an extracellular matrix of proteins and 
polysaccharides that encourage binding of more 
bacteria both to the matrix and to each other, 
providing stability, nutrition, and protection. 
Biofilms are known to significantly decrease the 
effectiveness of antimicrobial agents through 
different adaptations that shield one another  from 
the toxic effects of the disinfectant, slow disinfectant 
penetration, and even prevent antimicrobial agents 
from reaching bacteria at the center of the biofilm, all 
of which prevent sterility from being achieved.  
 
Factors external to the organism 
Surface factors: Smooth surfaces are easier 
to disinfect while objects with rough surfaces are 
more difficult because of the microscopic crevices 
that can house microorganisms.  In addition, the 
organic load – the presence of organic material like 
blood or tissue – inhibits the action of disinfectants. 
A high organic load can  block neutralizing agents 
from reaching the surface, and biomaterial itself can 
even inactivate agents such as bleach. Proper 
cleaning with water and detergent or enzymatic 
cleaners prior to disinfection or sterilization can 
reduce 
the 
organic 
load 
and 
increase 
the 
effectiveness of the disinfectant or sterilant. 
Similarly, biofilms created by certain organisms 
prevent agent contact with the organism itself, 
thereby requiring both a longer contact time overall, 
as well as higher agent concentration.  
Agent factors: The concentration of the 
disinfecting agent is an important factor to consider 
when attempting disinfection or sterilization, as 
higher concentrations of a chemical agent do NOT 
always increase the microbial death rate. For 
example, a 70% isopropyl alcohol solution can kill 
microorganisms in seconds because water both 
allows the isopropyl alcohol to enter and penetrate 
the entire cell and increases the contact time of the 
disinfectant on the surface by slowing the rate of 
evaporation of the isopropyl alcohol. Conversely, 
solutions with concentrations above 91% are not as 
effective because the alcohol’s action on the outer 
layer of microbes creates a protective layer of 
denatured proteins, which actually shields other 
proteins from becoming denatured. Lastly,  no 
concentration of isopropyl alcohol effectively kills 
Sterilization and Disinfection 
Chris Gross, Sanaz Dovell 
 
OPEN MANUAL OF SURGERY IN RESOURCE-LIMITED SETTINGS 
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spores, thus alcohol cannot be used to achieve high-
level disinfection or sterilization.  
Duration of exposure: Time is also 
important. The time required for disinfection is a 
function of bacterial load, concentration of the 
disinfecting agent, as well as intrinsic properties of 
the disinfecting agent. For example, 70% isopropyl 
alcohol can kill Mycobacterium tuberculosis in 5 
minutes, whereas a 3% solution of phenol requires 2-
3 hours to achieve the same effect.  
The efficacy of disinfecting and sterilizing 
agents is described by their decimal reduction time 
(D-value), or death rate curve, which is the amount 
of time it takes to kill 90% of the microbial 
population. For example, an antimicrobial agent with 
a D-value of one minute will kill 90 million cells in 
a population of 100 million bacterial cells,  leaving 
10 million cells alive after one minute. However, 
after two minutes, one million bacterial cells will still 
be alive because this agent is known to kill 90% of 
the bacterial cells in one minute. Therefore, 
sterilization, which requires eradication of all living 
cells, viruses, and spores, can be achieved by certain 
agents by increasing the duration of sterilization to 
several D-values longer than the time needed to 
theoretically reduce the microbial population down 
to one cell.  For example, glutaraldehyde-based 
solutions of ~2% can achieve high-level disinfection 
(elimination of microorganisms but not spores) with 
exposure times of 12-30 minutes, but sterilization 
can be achieved with an exposure time of 10 hours. 
Proper protocols, including concentration and 
exposure time, should be carefully considered and 
adhered to when selecting the appropriate agent for 
disinfection or sterilization. 
Physical 
factors: 
In 
general, 
most 
disinfectants work at room temperature (~20-22C), 
and, to some extent, the temperature is directly 
proportional to the activity of the disinfectant. 
Certain disinfectants require a specific pH to be 
active, while others cannot be used together because 
they can neutralize one another. Additionally, an 
agent’s physical properties must be taken into 
account. For an antimicrobial agent to effectively kill 
microbes at the concentration and amount of time 
determined by studies, the agent must remain in 
contact with the item being disinfected for the entire 
duration of time. For example, while 70% isopropyl 
alcohol may kill M. tuberculosis in 5 minutes, if the 
70% isopropyl alcohol being used evaporates within 
30-60 seconds, then disinfection will not be 
achieved. 
 
TOOLS AND TECHNIQUES 
As mentioned above, both the instrument 
type and organism type must be considered when 
determining the best method for sterilization. 
Broadly, the techniques can be divided into physical 
methods (heat) and chemical methods. In all 
situations, the instruments must be disassembled as 
much as possible and thoroughly cleaned with water 
and detergent or enzymatic cleaners to remove all 
foreign material from the surface of the equipment. 
Failure to do so can significantly limit the 
effectiveness of the sterilization process. 
 
Physical methods: 
The primary physical methods include 
applications of dry heat and moist heat, with moist 
heat being more effective due to water’s ability to 
penetrate the cells.  
Dry heat: Dry heat is used for sterilizing 
metal objects, powders and glassware (all other 
materials will melt).  It requires both higher 
temperature (160 to 180 oC) and longer exposure 
time (1-3 hours) than moist heat. Ovens are very 
common in dental offices, and due to their simplicity, 
they can be used in resource-constrained settings.   
 
Electric heat sterilizer in a dentist’s office 
 
Sterilization and Disinfection 
Chris Gross, Sanaz Dovell 
 
OPEN MANUAL OF SURGERY IN RESOURCE-LIMITED SETTINGS 
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Moist heat: Moist heat at high pressure, also 
known as steam sterilization, is the simplest and most 
frequently used method of heat sterilization. Moist 
heat alone (boiling water at 100 °C) can effectively 
kill most organisms, but spores easily survive. The 
use of moist heat at high pressure raises the boiling 
point of water to produce steam at a temperature that 
can effectively kill spores and achieve sterilization. 
Both pressure cookers and autoclaves use heat under 
the pressure of steam to eliminate organisms. Unlike 
dry heat, some hard plastics can be safely sterilized 
in this manner, such as orthopedic implants or some 
laparoscopic equipment.   
Steam sterilization has four parameters: 
steam, pressure, temperature, and time. Standard 
sterilization procedures use steam at 121 °C (250 °F) 
at 15 psi (pounds per square inch) for 20-30 minutes, 
which effectively kills all spores to achieve sterility 
in a gravity displacement autoclave.  At higher 
temperatures of 134 °C (273 °F), sterilization can be 
achieved in four minutes, if a pre-vacuum autoclave 
is used. Note that instruments packaged in kits with 
multiple layers, as well as liquids, require longer 
times in the autoclave.  
 
Minimum cycle times for steam sterilization; adapted from 
CDC (reference at end of Chapter.)  
 
 
If the release valve fails, the autoclave can be turned into a dry 
heat autoclave, which creates higher temperatures and will 
likely melt any plastics in the device (in this case sizers for a 
knee arthroplasty set.). 
 
Flash sterilization is a modification of the 
standard steam sterilization that sterilizes unwrapped 
items at 132 oC for 3-4 minutes at 27-28 psi. It is 
designed for use only in emergency situations, when 
there is not enough time for a standard sterilization 
cycle. Items to be sterilized must have already gone 
through the proper cleaning and decontamination 
process. Since items are unwrapped, contamination 
can occur as soon as the item is removed from the 
sterilizer, increasing the risk of infection. Flash 
sterilization should be avoided for any implantable 
devices, and it should not be used as a method for 
convenience.  
Filtration: Filtration can be useful if an IV 
medication used during a procedure must be 
sterilized, since heat or chemical sterilization will 
denature drug compounds. Micropore filters with 
pore sizes of 0.2 μm will remove bacterial cells, but 
not viruses. Filtration of viruses requires a pore size 
of 20 nm.  
Irradiation: Ultraviolet radiation has poor 
penetration, therefore, it can only be used to sterilize 
surfaces. Other forms of ionizing radiation, such as 
gamma rays, are associated with higher costs and 
harmful effects on equipment compared to other 
methods of sterilization. Therefore, sterilization via 
irradiation is not recommended. 
 
Chemical methods:  
The most common chemical methods are 
iodine, 
alcohols, 
chlorine 
derivatives, 
glutaraldehyde, hydrogen peroxide, peracetic acid, 
enzyme solutions, and ethylene oxide. Not all 
chemicals can be used for sterilization, even with 
increased exposure time, and careful consideration of 
proper storage and disposal of these chemicals must 
be taken when selecting the appropriate agent to use.  
Sterilization and Disinfection 
Chris Gross, Sanaz Dovell 
 
OPEN MANUAL OF SURGERY IN RESOURCE-LIMITED SETTINGS 
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The chemicals in use can be tracked and standardized using an 
erasable board such as this one, supplied by the manufacturer 
of the chemicals. 
 
Iodine: Iodine often comes in the form of a 
tincture mixed with an alcohol or as an iodophor 
(iodine mixed with a solubilizing agent). Iodophors, 
which increase the activity of iodine, are found in 
ointments, surgical scrubs, and solutions such as 
Betadine (povidone-iodine). For iodine to become 
effective, it must be properly diluted (~ 10% 
solution), and have at least 30 seconds of contact 
time with the object. It is important to note that 
iodine is not sporicidal. 
Alcohols: Isopropyl and ethyl alcohols are 
the most common type of alcohol used for low-level 
disinfection, but not sterility. Instruments are first 
thoroughly cleaned by scrubbing with water and 
detergent or enzymatic cleaner, then typically 
submerged overnight in 70% solution (ranges from 
60-90%). These agents work well against most gram 
negative and gram positive bacteria, and are a 
common option in severely resource-limited settings. 
It is important to note that these alcohol agents are 
not sporicidal and are ineffective against hydrophilic 
viruses such as polio.  
Chlorine: 
Chlorine 
and 
its 
derivative 
compounds are halogens that are commonly used  for 
disinfection, as it can kill all microbes but cannot kill 
spores. The most common chlorine disinfectant is 
sodium hypochlorite (Chlorox, or bleach) which has 
a 1:10 dilution of 5.25% concentration (0.5% to 1% 
is required for disinfection). As mentioned 
previously, organic matter can inactivate bleach, so 
proper cleaning to remove biomaterial prior to the 
use of bleach is essential for proper disinfection. 
 
Left: sodium percarbonate, tetraacetyl- ethylenediamine and 
N-alkyl(C12-14)-N-benzyl-N, N-dimethylammonium chloride 
tablets. 
Right: 
Didecylmethylammonium 
Chloride 
and 
Chlorhexidine Gluconate solution.  
 
Enzymatic solutions: Enzymatic solutions 
use various proteases to break down biomaterial at 
neutral pH. They are used after initial washing and 
scrubbing, and can reduce organic load in difficult to 
reach areas of equipment without destroying delicate 
and expensive equipment. For this reason, enzymatic 
solutions are ideal for endoscopic equipment. 
Depending on the solution, these enzymes can be 
combined with other chemical agents for instruments 
to soak. The enzymes used in these cleaners break 
down proteins on any surface it comes in contact 
with; therefore, care must be taken while using these 
cleaners to avoid skin contact, accidental ingestion, 
contact with mucous membranes, or inhalation if 
aerosols are used.  
 
Sterilization and Disinfection 
Chris Gross, Sanaz Dovell 
 
OPEN MANUAL OF SURGERY IN RESOURCE-LIMITED SETTINGS 
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Enzymatic cleaner, a mixture of protease, amylase, lipase, 
carbohydrase, and “proprietary enzymes.” This is mixed with 
(non-sterile) 
water 
according 
to 
the 
manufacturer's 
instructions- the mixture is changed daily.  
 
Glutaraldehyde: This solution, also known as 
“Cidex” is a chemical solution that can be used for 
cold disinfection. It requires a pH >7, with greatest 
efficacy at pH 9-10. Items are submerged in a ≥2% 
solution for 20-90 minutes at 20 oC to achieve high-
level disinfection or 10 hours to achieve sterility. 
This solution can be reused for 14-30 days. Note that 
this solution can be quite toxic, therefore proper PPE 
and procedural care must be taken to avoid inhalation 
or skin contact. Although it is widely used in the 
healthcare setting, especially for disinfection of 
endoscopic equipment, this solution may not be legal 
in all countries. 
 2% Glutaraldehyde solution: this is not to be diluted. .  
 
Hydrogen peroxide can be used to achieve 
high-level disinfection via a 7.5% solution for 30 
minutes at 20 oC. Sterilization can be obtained by 
increasing contact time to 6 hours. This solution can 
be reused for 21 days. 
Peracetic acid at a concentration of 0.2% at 
50-56 oC can be used to achieve sterility when items 
are submerged for 12 minutes. A solution of 7.35% 
hydrogen peroxide with 0.23% peracetic acid can be 
used to achieve high-level disinfection at 20 oC for 
15 minutes. Increasing exposure time to 3 hours is 
effective for sterilization. In addition, this solution 
can be reused for 14 days. 
Ethylene oxide: This is a poison gas that very 
effectively sterilizes laparoscopic equipment as well 
as plastic materials since it does not use heat or 
moisture. A concentration of 700 mg/L can sterilize 
at 38 oC (100.4 oF) in eight hours and at 54 oC (129 
oF) in four hours. However, this method is not readily 
available, as it requires a special ethylene oxide 
chamber, and the used gas must be destroyed either 
via a sulfuric acid scrubbing mechanism or catalytic 
oxidation. In addition, this gas is explosive. 
Therefore, in resource-limited settings other methods 
should be used for disinfecting/sterilizing certain 
equipment such as rubber, fragile plastic or 
laparoscopic cameras. 
 
PRACTICE 
Given 
that 
this 
topic 
is 
vast 
and 
sprawling,  focus will be placed on a few topics 
related to common practices: general operating room 
layout for optimal sterilization, a case-based walk 
through of general steps to be taken after an 
operation, and finally a few tips for operating 
common sterilization tools. Remember, the steps to 
achieve 
sterilization 
include 
cleaning, 
decontamination / cleaning, disinfection, and 
sterilization. 
 
Operating room and sterilization area design. 
The operating room has a network of support 
areas/rooms 
that 
allow 
it 
to 
function: 
decontamination area, packing area, sterilization 
area, and sterile supply room. Each of these distinct 
zones are necessary within your operating facility  to 
keep operations going. 
Decontamination area: This area is the first 
stop after an operation is completed. It has a sink 
used for mechanical removal of any bioburden, via 
high pressure water, air, or manual scrubbing. It also 
has an area for submersion disinfection (i.e 
enzymatic cleaner or glutaraldehyde.) It is physically 
separated from the other areas to reduce risk of cross-
contamination. 
Sterilization and Disinfection 
Chris Gross, Sanaz Dovell 
 
OPEN MANUAL OF SURGERY IN RESOURCE-LIMITED SETTINGS 
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The decontamination area is well ventilated, well drained, with 
areas for disposing of contaminated waste and washing soiled 
instruments.  
 
Packing area: This area, which is separate 
from the decontamination area, has large surfaces 
such as tables where disinfected instruments can be 
sorted, packed and wrapped. This packing area also 
houses any disposable items, such as paper and 
gloves, as well as linens. 
 
In the packing area, clean and dry instruments are wrapped for 
sterilization. In this instance, the table behind the technician 
contains instruments that are air-drying (Red arrow.) Once 
they are dry they are transferred to this area for wrapping.  
 
Sterilization area: The sterilization area is 
where the autoclaves are located. Ideally, multiple 
autoclaves would be operating to allow for a more 
efficient sterilization process, as autoclave cycles are 
time-consuming and require an adequate drying time 
before sterile packs can be handled and transported.  
 
Wall-mounted autoclaves in the sterilization area, with a trolley 
to support the rack as it is being unloaded. 
 
Sterile supply room: After sterilization has 
been achieved, the items must be properly stored. 
However, it is important that the sterile packs are 
completely dry before moving them to the sterile 
supply room. Moisture encourages microbial growth, 
and microbes can even travel to the inside of the 
sterile packs causing contamination. 
 
Instruments are stored in a cool, dry area with labeled shelves 
for easy retrieval.  
 
Case-based 
example: 
Decontamination 
to 
Sterilization to Storage 
 
1. Initial handling, cleaning of contaminated 
items, and movement  to decontamination 
area 
Sterilization and Disinfection 
Chris Gross, Sanaz Dovell 
 
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At the conclusion of an operation, all 
instruments (including unused instruments) must be 
handled as if they are contaminated and moved to the 
decontamination 
area. 
Any 
tubing, 
suction 
containers, and/or fluid-filled basins must also be 
placed on a cart, covered with a sheet, and then 
moved to the decontamination area. Any fluids from 
this cart can be offloaded into a fluid hopper once in 
the decontamination area. 
Once in the decontamination area, proper 
personal protective  equipment (PPE) should be 
donned; this includes face and eye protection, a thick 
apron, thick gloves, as well as a long-handled brush. 
If possible, forceps should be used to empty the 
instruments from the cart or, more simply, 
instruments can be dumped out onto a towel. Care 
should be taken to never reach into any container or 
tray without looking first; it should always be 
assumed that each tray or container has sharp items 
capable of breaking skin and causing infection. Any 
biohazard or soiled linens should be placed in the 
appropriate receptacle. 
 
In this instance, the scrub technician has separated the 
instruments that were not used during the surgery. They are 
placed in an enzymatic solution to soak in case any unknown 
contamination occurred during the surgery. 
  
 
Soiled linens are soaked in this decontamination solution 
before being taken to the laundry for washing.  
 
A note about instruments: Stainless steel 
is  not actually stainless and can experience corrosion 
from biological and chemical liquids. As a rule of 
thumb, any soiled instruments should be cleaned 
within twenty minutes of contamination. If 
immediate cleaning is not possible, organic matter 
such as blood or tissue can be prevented from drying 
on the surface of the instrument by submerging dirty 
instruments in water containing enzymatic detergent. 
During a surgery, instruments can be submerged in 
sterile, distilled water to clean and remove blood or 
other visible debris while not in use.  
The leading cause of pitting/stained surgical 
equipment is moisture (pus, blood, cleaning 
solutions). Use of housekeeping cleaning solutions, 
laundry or dish detergent, and iodine based solutions 
can also lead to staining and pitting of surgical 
equipment. Make note of any instrument in disrepair 
and set it to the side.  
 
2. Cleaning while in decontamination area 
All reusable instruments, tubes, suction 
devices, and packing trays used during an operation 
must be cleaned thoroughly prior to disinfection or 
sterilization. The presence of biomaterial on the 
surface of equipment hinders the sterilizing 
capability of autoclaves and can inactivate chemical 
sterilants and disinfectants. The general workflow is 
to disassemble and sort, spray/soak, scrub, rinse, dry, 
lubricate, and then disinfect or sterilize.   
Sterilization and Disinfection 
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Disassembly: All equipment or devices with 
removable parts should be disassembled according to 
the manufacturer’s guidelines to facilitate removal of 
all organic debris and allow better access to all parts 
of the instrument by the disinfectant or sterilant. 
Disassembled parts should be kept together for easy 
reassembly later on. Sorting equipment into groups 
that will be processed in the same way helps to 
streamline the overall process. 
Spray/Soak: After disassembly and sorting, 
items are placed in the sink and sprayed with water 
(preferably high pressure) and/or compressed air to 
remove large chunks of debris. Items can then be 
soaked in an enzymatic cleaning solution with a 
neutral pH for 10-20 minutes to help remove debris 
from hard-to-reach places and make the cleaning 
process easier and more effective. Extensive 
soaking, such as overnight soaks, can damage 
equipment and should be avoided.  
 
Near the washing area, contaminated instruments are placed in 
the enzymatic solution before being scrubbed.  
 
Scrub: If available, an ultrasonic cleaner 
should be used to mechanically scrub items after 
spraying down and soaking them. Manual scrubbing 
may be used if this is not an option. It is important to 
note that low-mineral water (distilled or reverse 
osmosis water) is essential for cleaning instruments 
because damage can be caused by high mineral 
content water. Saline should also never be used in the 
cleaning process. A nylon brush can be used to 
manually scrub organic debris from instruments. If 
debris is still present, a stainless steel brush can be 
gently used, with care being taken to prevent scoring 
or scratching of metal (which creates crevices for 
organisms to hide, grow, and form biofilms).  
 
As seen here, the technician has donned an impermeable gown 
and thick gloves (as well as a mask and eye protection.) The 
needle holder in her left hand has been opened and she is 
scrubbing the rough surface inside its jaws with a heavy brush. 
 
 
Even if all removable sharps (scalpel blades, needles) have 
been removed, the technician must take special care because 
some instruments can still cause injury. Examples include 
penetrating towel clips (shown here) osteotomes, and other 
sharp instruments.  
 
Rinse: All equipment/devices should be 
thoroughly rinsed after scrubbing to remove any 
residual detergent. Residual detergent can react with 
sterilizing solutions and disinfectants and hinder the 
disinfectant or sterilization process.  
Sterilization and Disinfection 
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Dry: After thoroughly rinsing, equipment 
must then be allowed to completely dry, either by 
hand or air-dried, for maximal efficacy of chemical 
disinfectants. 
 
Here, cleaned but still wet instruments are passed through a 
window from the decontamination area to the sterilization area 
to dry before being wrapped for sterilization.  
 
Lubricate: After the instrument has been 
cleaned of visible debris, a process called gross 
decontamination, surgical instrument lubricant 
should be applied to any mobile joints or hinges 
before sterilization.  
 
Do not use industrial lubricants like WD40 or any 
oil-based lubricants. Instead, water soluble 
lubricants that are steam-permeable should be 
used.  
 
At this point, semi-critical patient care items, such as 
endoscopes, should undergo high-level disinfection. 
Critical medical and surgical items should be 
packaged for sterilization.   
 
3a. High-level disinfection.  
Semi-critical patient care items that have 
been properly cleaned, rinsed, and dried should be 
soaked in a high-level disinfectant for the minimum 
effective time and concentration that has been 
determined for that particular chemical disinfectant. 
Lengthy submersion may cause damage, especially 
to finer instruments. 
 
Laparoscopic instruments, after being scrubbed, rinsed and 
dried, are soaked in glutaraldehyde solution for the time 
prescribed by the manufacturer.  
 
It is important to note that unwrapped 
instruments must be used either immediately after 
sterilization, or placed in a dry, covered sterile tray, 
where they can be stored safely for up to one week. 
 
After soaking in glutaraldehyde, laparoscopic instruments are 
rinsed in sterile saline and allowed to dry using strict sterile 
technique. A technician then loads them into a sterile container.  
 
Sterilization and Disinfection 
Chris Gross, Sanaz Dovell 
 
OPEN MANUAL OF SURGERY IN RESOURCE-LIMITED SETTINGS 
www.vumc.org/global-surgical-atlas 
This work is licensed under a Creative Commons Attribution-ShareAlike 3.0 Unported License  
 
 
Laparoscopic instruments that have been treated as above and 
placed in a sterile container. When needed, they are retrieved 
using sterile technique and placed on the surgical table.  
 
 
Common disinfectants and sterilants and their concentrations 
and exposure times.  
 
Avoid using any benzyl ammonium chloride 
solutions with instruments that have tungsten-
carbide inserts.   
 
If your surgical theater is set up with a dedicated 
decontamination and clean area, ensure that soiled 
items are loaded from the decontamination side 
and then (after washing) clean instruments are 
removed from the clean side. 
 
3b. Packing and sterilization 
After decontamination and disinfection, the 
items can now be sterilized by equipment such as an 
autoclave. First, a pack must be created.  
A thoughtful strategy when making packs 
can help reduce the “wear and tear” on instruments. 
By creating smaller packs that can be added together 
to make a larger unit, unnecessary future processing 
can be avoided, which in turn minimizes repairs. 
When packing surgical instruments into 
sterilization pouches, ensure that the instruments are 
open and in their UNLOCKED position, which 
allows steam to reach all active surfaces, and 
prevents cracks from heat expansion. Locking 
jaws/blades will prevent proper sterilization and may 
damage the  instrument’s box joints during the 
heating sterilization process. Ensure that any  sharp 
tips are covered, yet are covered in a way that still 
allows steam to penetrate the covering.  
Use disposable paper: Packs can be created 
out of individual instruments or entire sets of 
instruments, which will then be placed in the 
autoclave to achieve sterilization. Pouches should 
not be overpacked with instruments, and there should 
be adequate room in the pouch for steam to  reach the 
surfaces of the instruments.  
 
The technician here is using a combination of paper and cloth. 
He folds the instrument tray on all four sides, covering it 
completely.  
 
 
Here, he has secured the wrapping with both tape and string. 
The instrument set is labeled as below.  
 
Sterilization and Disinfection 
Chris Gross, Sanaz Dovell 
 
OPEN MANUAL OF SURGERY IN RESOURCE-LIMITED SETTINGS 
www.vumc.org/global-surgical-atlas 
This work is licensed under a Creative Commons Attribution-ShareAlike 3.0 Unported License  
 
 
The smaller strip of tape with the date on it is autoclave tape: 
the fine white stripes on it will turn black when exposed to 
sufficient temperature and pressure to sterilize the instruments. 
The date of sterilization and expiry are noted in the upper part 
of the photo. The name of the instrument tray is noted on the 
lower part.  
 
Using the autoclave: After the cleaned and 
decontaminated, but non-sterilized packs have been 
created, the pouches may be placed into the 
autoclave to be sterilized. These packs, however, 
should not be stacked because it will block steam 
from circulating throughout the chamber. Once 
properly packed, the autoclave may be started. 
Depending on the model of the autoclave, once the 
autoclave has cycled, the door may be opened 
slightly (~ 1cm) to allow any steam to escape. The 
door should not be opened fully as condensation can 
form on the instruments and pouches as cold air 
rapidly enters the autoclave. After running the dry 
cycle, sterile tongs should be used to remove the dry 
and sterile items. 
Storage after sterilization: The sterilized 
packs should be completely dry on the wire racks 
before moving to sterile storage. If instruments are 
wrapped, they can be stored indefinitely in a warm, 
dry and closed space such as a cabinet  (provided the 
pack remains intact and dry).   
 
Sterilization Tips and Procedures for Various 
Appliances 
 
1. Commercial or Domestic Pressure Cookers 
 
A simple electric pressure cooker autoclave. Source: Viv Rolfe, 
CC BY-SA 4.0  
https://creativecommons.org/licenses/by-sa/4.0, 
via Wikimedia Commons 
 
As discussed earlier, steam sterilization is 
more effective than dry heat, and use of steam under 
high pressures is effective at killing spores, provided 
the appropriate parameters of temperature, pressure, 
and time are used. Most commercial pressure 
cookers can sterilize effectively since they can 
achieve the pressure, temperature, and time 
parameters of 15 psi, 121 oC, and 30 minutes that are 
needed for sterilization.  Store-bought, or home 
pressure cookers are relatively low-cost; however, 
most are unable to reach the temperature and 
pressure requirements needed to kill spores. One 
study demonstrated sterilization using an “Instant 
Pot,” home pressure cooker, but it required a duration 
of 150 minutes to kill the spores in the test sample. 
The extended time requirement has to be taken into 
account when operating in low-resource settings 
where electricity may not be reliable. One company 
has created a pressure cooker, the EcoClave, that can 
sterilize instruments in 30 minutes using burning 
wood as the fuel source, making it a potentially  ideal 
option for sterilization in very low resource settings 
(source.) 
When operating any pressure cooker, all 
manufacturing instructions should be followed 
closely for proper safety. First, all instruments must 
be cleaned and decontaminated as discussed 
previously. A barrier, such as a trivet or bowl, should 
be placed between the inner surface of the pressure 
cooker and the items being processed. Items to be 
sterilized can also be placed into sterilization packs 
Sterilization and Disinfection 
Chris Gross, Sanaz Dovell 
 
OPEN MANUAL OF SURGERY IN RESOURCE-LIMITED SETTINGS 
www.vumc.org/global-surgical-atlas 
This work is licensed under a Creative Commons Attribution-ShareAlike 3.0 Unported License  
 
or bags, just like when using an autoclave. Autoclave 
tape can also be used over the packs to ensure that 
the proper temperature for sterilization has been 
achieved. When placing items inside the cooker, 
adequate spacing should be maintained to allow 
proper circulation (just like an autoclave). Enough 
distilled water should be added to reach a height of 
2.5 cm from the bottom of the pot for steam 
production. The cover should then be placed and 
properly sealed. At this point, the external heat 
source should be applied to the pressure cooker, 
either wood fire, gas stove, or electric burner. (For 
the Instant Pot, the device must be plugged into an 
electric outlet and operating instructions followed for 
a total cook time of 150 minutes). After heating has 
started, steam should be allowed to exhaust for about 
four minutes before closing the outlet.  
The pressure gauge should be watched 
carefully. When the pressure has reached the green 
area, or 15 psi, sterilization has begun. The 
instruments should be allowed to process for as long 
as 35 minutes. After sufficient processing, the 
pressure cooker should be removed from its heating 
source and depressurized by opening the steam 
outlet. No skin should be in contact with the hot 
steam that escapes through the valve or serious burns 
can occur. Once the gauge has reduced to zero, the 
pressure cooker can be opened by lifting the lid in a 
direction that is away from any person’s face or 
body.  
 
Never open a pressure cooker under pressure. 
 
Once items are removed, they must be dried 
properly before storing.  Items can be placed either 
on a wire rack (covered with a fly net) or in an oven 
to dry (~200 degrees for 6 hours). Once dry, these 
items can be placed in a dated and labeled ziplock 
bag.  
 
Some home pressure cooker units cannot get hot 
enough 
or 
attain 
adequate 
pressures 
for 
sterilization. In fact, they may fail and rupture. 
However, the Instant Pot brand was shown to 
effectively achieve sterilization after 150 minutes 
of cook time. 
 
2. Large Autoclaves 
 
A large electronically controlled autoclave in a hospital in a 
resource-rich country.  
 
When using autoclaves, one of the most 
important components of maintenance is regular use 
and testing. When preparing to use an autoclave, 
appropriate PPE should be used, and overcrowding 
of the packs should be avoided.  Autoclave pouches 
that can be penetrated by steam should be used, and 
the packs should not be stacked on top of one 
another. If glass is being placed in the autoclave, it 
should be inspected for cracks to avoid shattering. If 
liquids are to be sterilized, a secondary container 
should be used to house the primary container to 
catch any spills. Autoclave tape is often used as a 
visual marker to ensure that adequate temperature for 
sterilization has been achieved  during the autoclave 
cycle.  
 
Considerations for special equipment 
This chapter has focused on general 
instructions for routine sterilization of common 
surgical instruments. However, there are special 
considerations for other tools such as endoscopic 
equipment.  
Endoscope 
sterilization: 
Endoscope 
sterilization is a unique process, requiring different 
steps than a metal instrument like a clamp. These 
Sterilization and Disinfection 
Chris Gross, Sanaz Dovell 
 
OPEN MANUAL OF SURGERY IN RESOURCE-LIMITED SETTINGS 
www.vumc.org/global-surgical-atlas 
This work is licensed under a Creative Commons Attribution-ShareAlike 3.0 Unported License  
 
instruments also generally have a much higher 
bioburden and microorganism load than other items 
due to the nature of their use (i.e colonoscopy). 
Therefore 
extra-care 
must 
be 
taken 
to 
properly/maximally disinfect/sterilize the equipment 
while also maintaining its function. An additional 
chapter will be dedicated to processing of special 
equipment.  
 
Further Reading: 
United States Centers for Disease Control (CDC) 
Guideline for Disinfection and Sterilization in 
Healthcare Facilities (2008). 
https://www.cdc.gov/infectioncontrol/pdf/guidelines
/disinfection-guidelines-H.pdf  
 
The Eco-Clave wood burning autoclave by MedAid 
International:  
https://medaid.co.uk/wp-
content/uploads/2021/11/EcoClave-Booklet-
2021.pdf  
 
Chris Gross 
University of Florida College of Medicine 
Florida, USA 
 
Sanaz Dovell, PhD 
University of Florida College of Medicine 
Florida, USA 
 
The authors wish to acknowledge the work of Dr. 
Bruce Steffes in preparing this material.  
