﻿IX, B. 4 Barber: The Pipette Method 321 



If there is difficulty in making a small drop, it is usually because 

 the cover is too dry or the pipette opening too large, especially 

 if large with very irregular edges. It is often possible to gauge 

 the size of the droplets by raising the pipette up and down with 

 a slight stippling movement, at the same time blowing gently 

 into the rubber tube. Liquid will come out more easily if the 

 tip is brought into contact with one of the larger drops of 

 condensed moisture. This is usually unnecessary, unless the 

 opening in the tip has a margin so even that it becomes closed on 

 contact with the cover glass. 



If no particular individual bacterium is wanted, a simple way 

 to isolate one organism is to take up some dozens of them, eject 

 them on the cover, and add broth from the pipette. Then fill 

 the pipette with this dilution and make a series of fine drops, 

 in one or several of which a single cell will appear. Where the 

 original mixture is not too thick, it is often easy to take any 

 isolated organism alone into a fine pipette. If several enter, it 

 is a small disadvantage, since they may be separated immediately. 



The droplet is made small so that one can easily assure himself 

 that it contains but one organism. There is no danger of error 

 if the droplet measures 25 microns or less in diameter. In such 

 a droplet, for example, in peptone solution or any clear fluid, 

 one can make sure of the presence of but one actively motile 

 cholera vibrio. There is no need for India ink or any addition 

 to the fluid to make the bacterium more conspicuous. With many 

 organisms, much larger droplets will meet the requirements 

 (Plate I, figs. 1 and 2). If necessary, one may introduce the 

 pipette cautiously to the edge of the droplet and remove the 

 liquid, leaving the organism against the cover where it becomes 

 more conspicuous. The isolation may be done with the low power 

 and the droplets examined with the higher powers afterwards, 

 or the whole process may be carried out under a high power, the 

 oil-immersion lens if desired. For the isolation of bacteria of 

 ordinary size, a ^-inch objective will suffice. 



If drops of moisture so large as to interfere with the making 

 of the fine droplets have collected on the cover glass, a suitable 

 area with fine droplets can almost always be found in the neigh- 

 borhood of a larger drop of agar or broth. If a particle of 

 doubtful nature is seen with the isolated bacterium in the droplet, 

 it is usually easy to pick up either it or the bacterium and place 

 each in a separate droplet. One may easily pick up the isolated 

 bacterium and transport it to another part of the cover glass. 



The isolation of bacteria can be done very quickly by one 

 experienced in the technique. I have performed the whole pro- 



