40 ESSENTIALS OF BACTERIOLOGY 



Dish II, containing 5 per cent, acetic acid (i : 20), where it 

 remains one-half to one minute. The acid removes the excess 

 of stain. 



Dish III, water, to rinse off the acid. The section can now 

 be placed under the microscope, covered with cover-glass to see 

 if the intensity of the stain is sufficient or too great. A second 

 section is then taken, avoiding the errors, if any; and having 

 reached this stage, proceeded with as follows: 



Dish IV, alcohol, two to three seconds, to remove the water in 

 the tissue. 



V. A few drops of oil of cloves, just long enough to clear the 

 specimen to make it transparent (so that an object placed 

 underneath will shine through). 



VI. Remove excess with filter-paper. 



VII. Mount in Canada balsam (xylol balsam). 



CHAPTER VI 

 SPECIAL METHODS OF STAINING AND MODIFICATIONS 



Gram's Method of Double Staining (For Cover-glass 

 Specimens). I. A hot solution of anilin- water gentian- violet 

 two to ten minutes. 



II. Directly, without washing, into Gram's solution of iodin 

 potassium-iodid one to three minutes (the cover-glass looks 

 black). 



III. Wash in alcohol 60 per cent, until only a light brow 

 shade remains (as if the glass were smeared with dried blood) . 



IV. Rinse off alcohol with water. 



V. Contrast color with either eosin, picrocarmin, or Bis- 

 marck-brown. The bacteria will appear deep blue, all else 

 red or brown on a very faint brown background. 



