Tissue culture techniques continued

Inoculation of the explant

This is done in the inoculation room in the sterile atmosphere of a laminar flow hood. The sterile air gently streaming through the hood allows you to freely open sterile culture tubes/ bottles and do the inoculation. 

 

 If the hood has ultraviolet germicidal lamp it should be turned on at least 15 minutes before working in the hood. Turn it off while working in the hood to avoid exposure of eyes and skin to the harmful uv rays. Except for the filter the hood should be wiped down gently with a disinfectant such as 70% alcohol or isopropanol. The air flow should be turned on 15 minutes before using the hood. Wash your hands thoroughly with soap and water and wipe them with 70% ethanol or isopropanol. wear a lab coat if possible. 

Tools used for inoculation such as forceps and scalpels are first wrapped in aluminium  foil and sterilized in an autoclave  Later they are sterilized by flaming them periodically over a Bunsen burner or a spirit lamp. Alternately they can be sterilized by keeping them in a glass bead sterilizer in between use

                                                                  Glass bead sterilizer

Points to remember while using the laminar flow hood

1.Avoid any obstruction of the laminar air flow because it will change the air flow pattern and so cause contamination  Do not have items such as beakers or racks of test tubes between the HEPA (high efficiency particulate) filter and the exposed cultures

2.Work at arms length as far back in the hood as it is practical

3.Do not make wide sweeping arm movements over the work area since contaminants may drop from the arms or sleeves

The more careful you are the lesser the chance of contamination