Section F P2 Flashcards
which wavelength is UV radiation most germicidal
UV radiation is most germicidial at 254 nm
. This is the wavelength at which the UV radiation is absorbed by DNA and RNA causing changes in their structure and rendering them unable to replicate.
Centrifugation separates the sample into:
pellet ( sedimented material)
supernatant (liquid that may be decanted from the tube.)
Sedimentation of a sample also depends on time and centrifugal force (or gravitational force.)
Factors affecting centrifugal force
- speed of rotation, expressed in rotation per minute (rpm).
- Radius of rotation (r), distance of the particle from the centre of rotation.
HORIZONTAL or SWING ROTOR (swinging or hanging bucket)
Mainly low speed centrifuges
In high speed ultra centrifuges
Tubes rotate from a vertical to horizontal position
Buckets return to vertical as centrifuge speed decreases (decelerates)
Advantages: produces well-formed pellets, versatile, rotor – buckets easy to change.
Disadvantages: longer path length, slower speed, so longer for separation; moving parts wear with extended use
Pelllet forms at the bottom tip
Fixed Angle Rotors
Advantages:short path length, so cells pellet faster
no moving parts so no major mechanical failures – wear only.
Disadvantage: cells may be damaged by being forced against wall of tube.
Pellet a little on the side
Centrifuge Maintenance Log
detail the number of runs age of rotors date of servicing rotation speed per run rotors send out for service name of person doing the maintenance or servicing comments
Maintenance and Quality Assurance of centrifuge
Clean inside of the centrifuge daily a 1/10 dilution of hypochlorite solution or an equivalent disinfectant
Clean bowl with a germicidal disinfectant.
Autoclave rotor heads and buckets
Remove all broken glass or plastic disposed of appropriately
Hazardous materials must be capped or in sealed buckets to prevent the release aerosols.
BROKEN TUBES in the CENTRIFUGE:
Turn off centrifuge.
If in sealed buckets, go to step 7; in unsealed cups, inform others in the vicinity and leave the immediate area for at least 30 minutes to allow aerosols to settle.
Slowly open centrifuge lid, remove all broken tubes, buckets, rotors, etc to a container of disinfectant which is non-corrosive; allow to stand for a time appropriate for the disinfectant used. Some items may be autoclavable.
Place any unbroken capped container in disinfectant for 60 minutes and then remove and rinse.
Wipe down the bowl of the centrifuge twice with disinfectant, rinse with water, then air dry.
Place towels or cloth used for the wipe down into autoclave bags in the biohazard disposal area.
Remove sealed bucket to the biological safety cabinet.
Leave broken tubes in the bucket, replace the lid loosely and autoclave the entire contents or place in disinfectant (see steps 3 & 4).
partial immersion thermometer
Has a line or ridge (called an immersion ring) on the stem (76 or 35 mm up from the bulb tip).
Designed for the bulb and a portion of the stem (up to the immersion ring) to be immersed
Waterbaths
Total immersion thermometer
Does not have an immersion line.
Is designed such that the bulb and the entire stem (liquid column-not the entire length of the thermometer) are immersed or exposed to the temperature being measured
Some manufacturers inscribing TOTAL or TOTAL IMMERSION on the reverse of the thermometer
Hot air ovens
Can be used in fridges , freezers and incubators
Complete Immersion Thermometers
Entire thermometer is exposed to the medium of the temperature is being measured
Designed for inside incubators, fridges, freezers
Calibration of liquid thermometers
Usually every three months - yearly
Traditional Method: of thermometer calibration
Measure the freezing point of water
Slushy mixture of ice and water-0º C (distilled water)
Wait least 3 minutes
Insert the thermometer, to a depth sufficient to cover the 0oC (32oF) graduation (total immersion), or to the immersion line (partial immersion).
Mark the point indicated when it comes to equilibrium
Take successive readings at least one minute apart. Readings should agree within one tenth of one graduation.
or do it with boiling water - 100D
Bi- Metallic Expansion
Principle- different metals expand at different rates when heated.
A bimetallic strip consists of two different metal strips fastened edge to edge. ( example: brass and steel)
Used in Refrigerator & Freezer, household thermostats
Electrical Continuous Reading Device:
Records the temperature inside the unit (freezer, fridge)
24 hours a day, on a sheet of paper
Some Immuno-Hematology (Blood bank) fridge. Built-in chart recorder or -70 Celsius freezer
Freezer and Refrigerator - Clean with disinfectant
Daily = highly used refrigerator and freezer, door handles
Weekly =less frequent used refrigerator and freezer, door handles
Monthly = (In addition to daily or weekly), frozen and refrigerated packs/gel packs ,thermometers and temperature monitors
Annually = Coils and motors(done by outside source) Thorough cleaning of : refrigerators, freezers, walk in cooler
water bath up keep
Drain, cleaned and re-filled water bath on a weekly basis to avoid build up of salts and contamination.
Cleaned with a mild laboratory detergent using a sponge or soft cloth.
Scale build up can be removed using a mild descaler and soft brush
Disinfectants like phenolic detergents are are added to the water bath to prevent bacterial contamination
Dont use bleach, Sodium azide- explosive, strong acid or salt
DISINFECTANT
chemical or physical agent that is applied to inanimate(non-living) objects to kill microbes, Example: Bench tops
ANTISEPTIC:
A non- toxic chemical agent that is applied to living tissue to kill microbes
May not be as effective at killing microbes as disinfectants
Can be used on human or animal tissue e.g. the skin
Not good for sterilization because of their slow action on some viruses, bacterial endospores and Mycobacterium tuberculosis
Bactericidal:
An antimicrobial that kills bacteria.
Bacteriostatic:
ntimicrobial that inhibits bacterial growth but does not kill all the bacteria.
Prolonged application of bacteriostatic agents can often result in bacterial death
The more organisms present the harder it is to disinfect.It als depends on the kinds of microorganism present-Endospores,M.tuberculosis,viruses etc.
Vegetative Bacteria
bacterial cells that are in the growing phase ie they can reproduce.
Endospore:
extremely resistant dormant cell structure produced by some bacterial species. If you break down the term endospore, ‘endo-‘ means ‘inside’ and ‘-spore’ refers to the ‘dormant structure,’ so the endospore is a structure formed inside the cell. It is a non-reproductive phase.
ECHANISMS OF ACTION OF CHEMICAL DISINFECTANT AND ANTISEPTIC AGENTS
Protein denature-disrupting the Hydrogen and Disulfide bonds
Membrane disruption
Nucleic acid damage-blocks metabolism
Inhibition of metabolism-e.g. lipids
Cell wall - chemical to use
Formaldehyde
Phenols
Chlorine releasing agents
Cytoplasmic coagulation
Chlorhexidine
Glutaraldehyde
Hexachlorophene
Cell membrane: membrane potential or electron transport
Hexachlorophene Phenols Parabens Weak acids used as food preservatives such as benzoic, sorbic and proprionic acids QACs (Quaternary ammonium compounds)
Leakage of cell components
Phenols
Chlorhexidine
Alcohols
Quats (Quaternary ammonium compounds)
Nucleic acids
Alkylating agents such as ethylene oxide gas
Spore core -Bacterial endospores:
Glutaraldehyde
Formaldehyde
Spore cortex -Bacterial endospores:
Chlorine releasing agents
Glutaraldehyde
Nitrous acid
Nitrates/nitrates act as food preservatives by preventing germination of endospores
Envelopes-surrounds the capsid
Chlorine releasing agents
Quats (Quaternary ammonium compounds)
Chlorhexidine
Viral nucleic acid
Chlorine releasing agents
Capsid-protein shell
Glutaraldehyde Quats (Quaternary ammonium compounds) Chlorine releasing agents Iodine Phenols Alcohols
Cell membrane of fungus
Chlorhexidine
Alcohols
Quats (Quaternary ammonium compounds)