Office of Research

Disinfection, Sterilization, Incineration and Waste Management


This protocol was developed by the Infection Control Committee and reviewed by the Institutional Biosafety Committee. It shall be reviewed annually by the ICC; all changes should be reviewed by the IBC.


The Scope of the Infection Control Protocol is defined by both regulatory and safety requirements. The requirement for an Infection Control Committee comes from SC Regulation 61-105 (R61-105), South Carolina Infectious Waste Management (June 2010): According to the regulation:

Each generator must have a designated infection control committee with the authority and responsibility for infectious waste management. This committee must develop or adopt a written protocol to manage the infectious waste stream from generation until offered for transport.

This protocol has been expanded beyond the narrow definition of infectious, which is limited to human pathogens, and addresses the management of biohazardous waste, defined below. This management protocol applies to activities at all University facilities except the Veterinary Diagnostic Laboratory, which has a separate permit, and the Patewood Bioengineering Center, whose waste is managed by Greenville Health System.

Biohazardous waste is waste that requires inactivation of the biological material in an approved manner prior to final disposal. It includes the following:

Sharps Waste. All hypodermic needles, syringes with needles attached, IV tubing with needles attached, scalpel blades, broken glass contaminated with a biohazard and lancets that have been removed from the original package.

Cultures and Stocks of Etiologic Agents and Associated Biologicals. This includes, but is not limited to, specimen cultures, cultures and stocks of etiologic agents and agents requiring biosafety level (BSL) 1, 2 and 3 containment, wastes from production biologicals and serums, and discarded live and attenuated vaccines.

Human Pathological Waste. This includes human tissues and anatomical parts that emanate from surgery, obstetrical procedures, autopsy, teaching and research laboratories. This does not include extracted teeth, hair, toenails, fingernails, human corpses, remains and anatomical parts that are intended for interment or cremation.

Human Body Fluids. This includes, but is not limited to, blood and blood products, serum and plasma, cerebrospinal fluid, synovial fluid, pleural fluid, peritoneal fluid, pericardial fluid, and amniotic fluid.

Infected Human Body Substances. This includes wastes that have come into contact with human body fluids or tissues from humans infected with, or isolated to protect others from, highly communicable infectious diseases.

Non-Human Primate Waste. This includes, but is not limited to, non-human primate blood, carcasses, tissues, body fluids, and bedding.

Animal Waste. This includes, but is not limited to, animal carcasses, body parts, and bedding of animals that are known to be infected with, or that have been inoculated with, pathogenic microorganisms infectious to humans; animals requiring Animal Biosafety Level (ABSL) 2 and 3 containment.

Recombinant DNA. All contaminated liquid, solid, carcasses, and animal wastes from research activities involving RG-2 agents requiring BSL-2 and ABSL-2 containment. 
All contaminated liquid, solid, carcasses, and animal wastes from research activities involving RG-3 agents requiring BSL-3 and ABSL-3 containment. NOTE: Although RG1-associated DNA is not included in the definition of infectious waste; all materials that have been in contact with recombinant DNA must be decontaminated prior to disposal in the sewer or trash. These items are not subject to the logging requirements described for autoclaves and chemical disinfection but they must still be decontaminated.

Laboratory Waste Which Has Come in Contact with a Biohazard (as listed above). This includes, but is not limited to, disposable laboratory personal protective equipment (gloves, gowns, shoe covers, masks), disposable laboratory plastic ware (culture dishes, plates and flasks, pipettes, and pipette tips), blood specimen tubes, devices used to transfer, inoculate and mix cultures; and paper and cloth which have come into contact with cultures and stocks of etiologic agents.

Additional wastes may be added at the discretion of the Infectious Waste Committee, in consultation with the Institutional Biosafety Committee (IBC).

Note that although not defined as biohazardous waste, rDNA experiments involving plants have NIH imposed requirements for waste. For both BL1-P and BL2-P, experimental organisms shall be rendered biologically inactive by appropriate methods before disposal outside of the greenhouse facility. See Appendix H for more information.


Disinfection. Disinfection is generally a less lethal process than sterilization. It eliminates nearly all recognized pathogenic microorganisms but not necessarily all microbial forms (e.g., bacterial spores) on inanimate objects. Disinfection does not ensure an “overkill’’ and therefore lacks the margin of safety achieved by sterilization procedures. The effectiveness of a disinfection procedure is controlled significantly by a number of factors, each one of which may have a pronounced effect on the end result. Among these are: the nature and number of contaminating microorganisms (especially the presence of bacterial spores); the amount of organic matter present (e.g., soil, feces, and blood); the type and condition of instruments, devices, and materials to be disinfected; the temperature.

Sterilization. Any item, device, or solution is considered to be sterile when it is completely free of all living microorganisms, endospores and replication competent viruses. The definition is categorical and absolute (i.e., an item is either sterile or it is not). A sterilization procedure is one that kills all microorganisms, including high numbers of bacterial endospores. Sterilization can be accomplished by heat, ethylene oxide gas, hydrogen peroxide gas, plasma, ozone, and radiation (in industry). From an operational standpoint, a sterilization procedure cannot be categorically defined. Rather, the procedure is defined as a process, after which the probability of a microorganism surviving on an item subjected to treatment is less than one in one million (10-6). This is referred to as the “sterility assurance level.”

Incineration is complete combustion of the waste and packaging to carbonized or mineralized ash. 

Biohazard Bag is a red or orange disposable, leak-proof bag having enough strength to prevent ripping, tearing, breaking, or bursting under normal use. These red or orange bags are marked with the biohazard symbol and the word "Biohazard." Biohazard bags are provided by Research Safety.

Biohazard Bin is semi-rigid, leak resistant secondary container that is impervious to moisture, marked with the biohazard symbol plus the word “Biohazard” and lined with a biohazard bag. Biohazard bins are provided by Research Safety. The bin may be plastic or cardboard.

Biohazard Bottle is a poly leak-proof, rigid poly container labeled with the biohazard symbol and the word "Biohazard." Biohazard bottles are provided by Research Safety.

Sharps Container is a red leak-proof, rigid, puncture-resistant, durable plastic container labeled with the biohazard symbol and equipped with a tight-fitting lid for use during handling and transport. Sharps containers are provided by Research Safety.

Unmarked autoclave bag is a clear (not red or orange) bag that does not bear the biohazard symbol.

All other terms used herein shall have the meaning defined by R61-105.



Individual principal investigators and/or departmental managers/supervisors are responsible for identifying the biohazardous waste generated by their activity, segregating it into the appropriate waste stream, managing it in accordance with this protocol and providing appropriate training to research personnel under their supervision. The second level of supervision is the Biological Safety Officer (BSO) who is responsible for ensuring, through training and inspections that all laboratory personnel are knowledgeable of and comply with procedures outlined in this protocol. Research Safety is responsible for picking up those biohazardous wastes requiring offsite treatment from laboratories in a timely manner, final packaging, manifesting and shipping them for incineration and recordkeeping in compliance with SC R61-105.


SC Regulation 61-105 requires that any material or surface that comes in contact with an infectious substance must be disinfected prior to reuse. Disinfection is permitted by appropriate use of an EPA registered disinfectant used according to the label instructions at the tuberculocidal (TB) strength. Clemson’s IBC requires that the disinfectant be registered for HIV-1 and HBV as well as TB. The list used for disinfection at Clemson is EPA Disinfectant List E (HIV-1, HBV, TB); see   The most familiar disinfectants on this list are Clorox Bleach (active ingredient sodium hypochlorite) at a concentration of 10% (1 part household bleach mixed with 9 parts water, hitherto called 10% bleach), Cavicide, and 70% isopropyl alcohol.

Spills shall be disinfected immediately with 10% bleach. As diluted bleach degrades rapidly, this solution must have been prepared within 8 hours of use. See Chapter 8 for detailed spill procedures.

Reusable containers that have been used to contain biohazardous waste must be disinfected immediately after being emptied.


SC Regulation 61-105 requires that infectious waste be treated by chemical disinfection, incineration or steam sterilization before disposal.  Clemson University requires that all biohazardous waste be treated with 10% bleach (liquids only), sterilized by autoclave or packaged for offsite contract incineration.


Sharps are deposited in a sharps container. All sharps waste is treated offsite by incineration; disposal is by the University’s Infectious Waste Contractor. Pretreatment is discouraged.

Re-usable sharps are safely segregated and contained in leak-proof, rigid, puncture-resistant containers while awaiting cleaning, decontamination, and sterilization before re-use.

Liquid biohazardous waste is chemically treated with Bleach and disposed via sanitary sewer or autoclaved and disposed via sanitary sewer or contained in a biohazard bottle which is placed in a biohazard bin. Biohazard bins are treated offsite by incineration; disposal is by the University’s Infectious Waste Contractor.

Solid biohazardous waste is either autoclaved in an unmarked autoclave bag and disposed of as ordinary trash or placed in a biohazard bin. Biohazard bins are treated offsite by incineration; disposal is by the University’s Infectious Waste Contractor.  Red or orange bags, or bags marked as biohazard cannot be disposed as ordinary trash, regardless of whether they were autoclaved.

Infectious animal body parts or carcasses are contained in biohazard bags, kept in designated freezers or refrigerators until pickup by Research Safety. They’re placed in a biohazard bin before removal from the lab.  Biohazard bins are treated offsite by incineration; disposal is by the University’s Infectious Waste Contractor.


Chemical Treatment Standards

Household bleach is 5 - 10 % sodium hypochlorite. The appropriate concentration of sodium hypochlorite for disinfecting liquid waste, e.g. supernatants from cell culture, is 5000 ppm, approximately 0.5%. A 1:9 (v/v) dilution of bleach (EPA registration number 5813-50) to liquid biological waste is required. A minimum contact time of sodium hypochlorite with liquid waste is 20 minutes. After 20 minutes of contact, disinfected liquid waste is poured into the sanitary sewer. Bleach should be stored between 50 and 70°F. According to Clorox, undiluted household bleach has a shelf life of six months to one year from the date of manufacture, after which bleach degrades at a rate of 20% each year until totally degraded to salt and water (exposure to sunlight or additional heat increases the degradation rate substantially), and a 1:9 bleach solution has a shelf life of 24 hours. However, this protocol requires that the dilution of bleach be mixed within the previous 8 hours.

All wastes that meet the definition of biohazardous waste and are treated chemically or by steam sterilization MUST be logged before disposal. The date and amount of waste treated and disposed must be included. A monthly log must be sent to the BSO at the beginning of each month for the previous month.

Autoclave Standards

All steam sterilizers must,

(a) use the biological indicator Bacillus stearothermophillus placed at the center of a load processed under standard operating conditions to confirm the attainment of adequate sterilization conditions. Indicator organisms must be used monthly at a generator facility in each steam sterilizer;

(b) record the temperature and time during each complete cycle to ensure the attainment of a temperature of 121 degrees Centigrade (250 degrees Fahrenheit) for 45 minutes or longer (refer to Appendix I of the Biosafety Manual for large quantities) at fifteen (15) pounds pressure, depending on quantity and density of the load, in order to achieve sterilization of the entire load; (Thermometers shall be checked for calibration at least annually.)

(c) have a gauge that indicates the pressure of each cycle.

(d) use heat sensitive tape that is lead-free or other device for each container that is processed to indicate that the steam sterilization temperature has been reached. The waste will not be considered appropriately treated if the indicator fails.

(e) maintain records of the procedures specified in (b) above for a period of not less than three (3) years.

(f) assure that treatment residues are disposed of in accordance with applicable State and Federal Requirements.