General Aseptic Techniques

Research -Wash hands

Bacteriological Quality of Water -Clean bench area with swab soaked in methylated spirit
*Various of Procedures available to measure Bacteriological -don't touch any part of container, pipette, etc. , coming in
Quality of Water contact with the sample/ culture
-Total Plate Count at 20C and at 30C -Do not open the lid of container or caps of test tubes longer
-Presence of Coliform Bacteria as indicator of sewage than absolutely necessary
contamination - Lightly the top of test tubes ends of pipettes, necks and
-Identification of specific pathogenic bacteria stoppers of bottle before and after adding/ withdrawing
-Employing miscellaneous indicators and serological samples/ inocula
methods -Transfer culture from one tube to another using wire loop
*Identification of specific micro-organisms difficult& costly sterilised by heating to redness and cooled
*Simplified test preferred as indicators of sanitary quality of -Always follow the methods to avoid contamination!
water Enumerating bacterial Populations
-Test for coliform group of bacteria Solid Media: Total Plate Count
-Total Plate Count Liquid Media: MPN (Most Probable Number) Less accurate
Culture of Micro-organisms than Total Plate Count
*Bacteria Grown in Laboratory under controlled conditions Enumeration on solid Media
-grown medium with nutrients and energy source *Plate Count in solid media
- Temperature -Individual colonies must be counted, must well separation
-pH, salinity, oxygen *each colony should represent 1 cell
-No competing organisms *each colony represents 1 cell
- No antibacterial substances *Dilution Techniques needed for high concentrations
*Typical solid media is 'agar', extracted from seaweed -1mL sample added to 9 mL diluent.
Types of Culture Media - Diluent should be standard salt solution
*General Purpose Dilution series made Dilution techniques for plate count
*Enrichment
*Selective
Culture of Liquid Media
*In liquid Media, Bacterial Growth is demonstrated by:
-Turbidity
-Change in color of indicators
-Gas Production
*Gas can be trapped in small inverted test tubes (Durham
Tube)
*Gas Production is used to indicate Coliform Bacteria.
*Bacterial Growth with Gas Production in Liquid Culture
Medium
Culture of Solid Media
*Bacteria in Solid agar media cannot move
*Cells grow locally, formed in colonies and clusters
*Colonies Visible to the Naked eye can be counted
*Solid agar media often poured into shallow, covering
(petri) dishes to harden
*Typical Solid Media is 'agar', extracted from seaweeds.
Types of Culture Media
*General purpose
*Selective
*Enrichment
Culture of Liquid Media
- In Liquid Media, Bacterial Growth is demonstrated by:
The Killer
-Turbidity
-Change in Color of Indicators
-Gas Production
* Gas can be trapped in small inverted test tubes (Durham
Tubes)
*Gas Production is used to indicate coliform Bacteria
*Bacterial Growth with Gas Production in Liquid Culture
Medium
Culture of Solid Medium
*In Solid 'agar' medium, bacteria Cannot Move
*Cells grow locally, forming colonies or clusters
*Colonies that can be seen through the naked eye can be
counted
Solid Agar Media can be poured into shallow, covered
(petri) dishes to harden
* Samples are already in petri dishes or added in dishes
after hardening
Sterilisation
*Sterilise all equipment and materials
- Use autoclave for steam sterilisation
-120C for culture broths and dilution water, 15 mins
-oven for dry sterilisation
-170C for pipettes, 20-30 mins
*for pipettes in metal containers