Sterilization Methods and Techniques Quiz

Sterilization, Disinfectants, and Antiseptics

• Sterilization is the killing of all forms of microbes, including spores.
• Disinfectants are antimicrobial agents applied to non-living objects to destroy microbes (reduce the number of pathogenic microorganisms).
• Antiseptics are antimicrobial substances applied to living tissue (skin) to reduce the risk of infection or decay. Note that not all disinfectants are antiseptics because antiseptics must not harm living tissue.

Sterilization Methods

• Sterilization methods are categorized into:
  • Chemical sterilization methods
  • Physical sterilization methods

I. Chemical Sterilization Methods

• This method is typically used for devices sensitive to high heat (steam sterilization) or damage from irradiation (e.g., rubbers, plastics).
• Subdivided into:
  • Chemical liquid sterilization
  • Chemical vapor sterilization

II. Physical Sterilization Methods

• These methods use physical agents or processes to eliminate or inactivate all microorganisms (bacteria, viruses, spores) from surfaces, equipment, and materials.
  • Autoclaving (Moist Heat Sterilization): Uses pressurized steam at high temperatures (typically 121°C or 250°F) for 15-20 minutes to sterilize media, instruments, and biohazard waste. Kills most microorganisms, including spores, via heat and moisture.
  • Dry Heat Sterilization: Involves heating materials to high temperatures (usually 160-180°C or 320-356°F) for 1-2 hours in a dry heat oven. Suitable for sterilizing metal instruments, glassware, and powders by oxidation of microbial cells.
  • Filtration: Removes microorganisms from liquids or air by passing them through filters with small pores (typically 0.2 microns). Used for heat-sensitive liquids (e.g., vaccines) and in HEPA filters for air sterilization. Effective for removing bacteria and larger viruses.
  • Radiation (Ionizing Radiation): Uses gamma rays or electron beams to sterilize materials by damaging the DNA of microorganisms. Penetrates deeply, effective for heat-sensitive materials.
  • Ultraviolet (UV) Radiation: Uses UV-C light (wavelength around 254 nm) to damage the DNA of microorganisms, preventing their replication. Commonly used for surface sterilization.
  • Boiling: Boiling water (100°C or 212°F) for 10-15 minutes kills most bacteria and viruses but may not eliminate all spores
  • Incineration: Destroys contaminated materials by burning at very high temperatures; suitable for disposal items.

Disinfectant Classification

• High-level disinfectants: Used for large numbers of spores after prolonged exposure. e.g. hydrogen peroxide (3-6%)
• Intermediate-level disinfectants: Used for a few number of spores. e.g., 70% ethyl alcohol, isopropyl alcohol
• Low-level disinfectants: Used for mainly vegetative bacteria, some fungi, and a narrow range of viruses. e.g. Providone-iodine (5 and 10%)

Chemical Vapor Sterilization

• Chemical Vapor under pressure (Chemiclave): Uses a mixture of chemicals (alcohol, formaldehyde, ketone, acetone, and water) heated under pressure to form a sterilizing gas. Requires 20 minutes at 131°C and 20 lbs pressure.
• Fumigation: Potassium permanganate and formalin are mixed in a specific ratio to produce a fumigant for sterilization.

Bacterial Diagnosis

• Microscopy:
  • Gram Staining: Classifies bacteria as Gram-positive or Gram-negative based on cell wall structure. Aids in initial identification.
  • Acid-Fast Staining: Primarily used to detect Mycobacterium species (e.g., tuberculosis) which have unique cell walls.
• Culture Methods: Use selective or differential media (e.g., agar plates, broth cultures) to promote bacterial growth, enabling colony morphology and color observations for pathogen identification.
• Biochemical Testing: Tests (e.g., catalase, coagulase, oxidase, or urease tests) determine the presence or absence of enzymes to differentiate bacterial types. Automated systems speed identification.
• Molecular Methods: Polymerase Chain Reaction (PCR) is a highly sensitive method for detecting bacterial DNA or RNA, allowing rapid identification of pathogens.
• Blood analysis: Evaluates blood components, detects infections, blood disorders, etc. Specific tests include:
• Complete blood count (CBC): Automated analyzers or manually using microscopy
• Hemoglobin and hematocrit: Measures hemoglobin concentration and proportion of red blood cells.
• Blood coagulation tests (PT and PTT): Evaluate blood clotting function to diagnose bleeding disorders or monitor anticoagulant therapy.
• Platelet Function Tests: Evaluate platelet adhesion and aggregation to diagnose platelet disorders and bleeding risks.
• Flow Cytometry: Detailed analysis of blood cell populations using tagged cells with fluorescent antibodies.
• Bone Marrow Analysis: Provides information on blood cell production and aids in diagnosis of hematologic cancers, anemias, and bone marrow disorders
• Clinical Chemistry: Measures various chemical components in blood, to determine organ function, metabolic state, etc.
  • Spectrophotometry/Enzymatic Tests: Measure the absorbance of light by substances or use specific enzyme reactions that produce color changes for identifying components like glucose, cholesterol, and proteins, etc.
  • Electrolyte Analysis: Measures electrolytes (sodium, potassium, chloride).
  • Immunoassays: Uses antibodies to detect specific proteins or hormones (e.g. insulin, TSH, troponin).
  • Chromatography (GC and HPLC): Separates compounds in blood or urine, and useful for drug testing, toxicology, or monitoring therapeutic drug levels.
  • Electrophoresis: Separates proteins based on charge and size in protein analysis, detecting abnormal protein levels or patterns.
  • Automated Analyzers: Modern labs use automated analyzers to measure biochemical markers like glucose, lipids, liver enzymes, and kidney function markers.

Laboratory Safety and Accident Prevention

• First Aid: Keep first aid kits, eyewash stations, and emergency showers accessible and train staff on how to handle injuries.
• Biochemical Hazards: Use proper storage, labeling, and personal protective equipment (PPE) for chemicals. Work in fume hoods and dispose of waste safely.
• Biological Hazards: Follow biosafety protocols (e.g., biological safety cabinets), use proper handling techniques, and disinfect surfaces.
• General Safety: Follow Standard Operating Procedures (SOPs), conduct regular safety checks, and maintain safety equipment. Always report any hazards.