22 



STERILIZATION BY FILTRATION 



another piece of glass tubing also bent at a right angle ; the vertical branch 

 should reach a few centimetres below the bung, while the horizontal arm 



has two constrictions with a fairly tight plug of wool C between 



them. 



The apparatus thus fitted up is heated in the autoclave at 



120 C. for 20 minutes. When cool, it is examined to see that the 



bung still fits tightly, and the apparatus is then ready for use. 



Technique. Stand the filter F in a glass cylinder E which must be 

 rather larger than the filter, and fill up the cylinder with the un- 

 filtered liquid. Connect the horizontal limb of the tube D by 

 means of pressure-tubing with a water pump (Chap. VI.) and exhaust. 

 The liquid is thus aspirated through the filter into the bottle A. 



When the liquid has all passed through, turn off the water and 

 disconnect the tubing connecting the bottle and the pump (the air 

 which will then enter the bottle is filtered through the wool plug C 

 between the constrictions). Flame the neck of the bottle, and replace 

 the bung either by a previously sterilized wool plug or by another 

 bung so arranged that the fluid can be manipulated as described later. 

 The liquid thus sterilized by filtration can be kept sterile indefinitely 

 in the bottle. 



There is always a little liquid left in the filter, and if necessary this 

 can be collected by disconnecting the tube B from the nozzle and 

 aspirating the fluid into a long sterile bulb pipette (fig. 21). 



FIG. 21.- Bulb 

 pipette. 



B 



Distribution of the filtrate. Having obtained a sterile filtrate, 

 it follows that the subsequent manipulations must be so devised 

 as not to contaminate it. The methods to be employed will now 

 be described. 



The bung used during filtration (p. 21) must be replaced by another fitted 

 in the following manner. Take an india-rubber bung perforated with two 

 holes of the same size as the one to be replaced. Through one hole pass a 

 piece of glass tubing A (fig. 22) bent at a 

 right angle and having a cotton-wool plug 

 between constrictions in the horizontal arm. 

 Through the other hole pass another piece 

 of glass tubing B bent in the form of an 

 inverted open U, one limb of which should 

 reach nearly to the bottom of the bottle 

 while the other, which will be outside the 

 bottle, is drawn out to a fine capillary point 

 and sealed . Wrap the bung with its glass tubes 

 in situ in paper, and autoclave at 120 C. 



Flame the neck of the bottle, remove the 

 paper covering from the sterilized bung, and 

 hold the latter by its upper part in the left 

 hand. Take out of the bottle with the right 

 hand the bung used for filtration, and replace 

 it by the other as quickly as possible in order 

 to prevent dust falling into the bottle. Care 

 must of course be taken that during these 

 manipulations the new bung with its tubes 

 comes in contact with nothing likely to soil it. 



To withdraw the fluid, it is only necessary to connect an india-rubber 

 syringe to the tube A, and after flaming the capillary end of B to break off 

 the point with a pair of sterile forceps. By squeezing the syringe a few 

 times the liquid will be forced out through B. When the quantity required 



FIG. 22. Distribution of the filtrate. 



