Laboratory Outline for General Bacteriology 



When ready to use mix with 

 Ferrous sulphate, saturated 



aqueous solution 5 cc 



Filter. 



Make stain by adding 2 cc of saturated alcoholic solution of 

 fuchsin to 8 cc of a 5% solution of carbolic acid. 



Use Mordant V$> 1 minute, wash with water. 



Carbol fuchsin 1/2 1 minute, wash with water. 



MORPHOLOGY 



The size, form, arrangement, etc., of vegetative cells is deter- 

 mined from twenty-four hour old agar slant cultures. Watery 

 fuchsin stain is used as it has been found satisfactory with most 

 bacteria. 



The Gram stain is made at the same time. This stain is for 

 differential purposes only. Some bacteria keep the stain and some 

 do not. When the bacteria remain dark blue they are recorded 

 as Gram positive, otherwise as Gram negative. 



Preparation of films. The method of preparing films for these 

 two stains is as follows : 



Place cover glasses in forceps without touching them with the 

 fingers. 



With sterile loop place a drop of water on each of two cover 

 glasses. 



With a cool sterile needle or loop transfer from edge of culture 

 in lower part of tube to the drops of water on the cover glasses. 

 Use only enough to cloud the water slightly. Burn any excess 

 of bacteria from the loop and spread the drops over the surface of 

 the cover glasses. If a drop will not remain spread but rolls 

 together again it shows that the cover glass is greasy and should 

 be discarded. 



Air dry without heat. 



Fix film to the cover glass by passing it three times through 

 the gas flame. Keep the film side up. Do not hold in flame longer 

 than you could hold your finger in it without burning. 



They are now ready to be stained. For staining methods see 

 page 35, 



After staining drying may be hastened by blotting with filter 

 paper. When dry mount on slide film side down in drop of cedar 

 oil. 



Label slides carefully. 



Never pile slides together. Place them in slide boxes. 



36 



