ANALYTICAL ELECTRON MICROSCOPY 151 



object holder. The latter is essential to absorb the 50-60 watts of 

 power given off by the heated filament, which would tend to soften 

 the sealing compound and weaken the vacuum. 



The microscope is evacuated through a port (P, Fig. 29) by 

 means of two double-stage mercury vapor pumps in parallel employ- 

 ing a Cenco Hyvac oil forepump. A vapor trap ccioled with carbon 

 dioxide in butyl alcohol is placed between the microscope and the 

 pumps, and the glass-to-metal connection is sealed with black 

 vacuum wax. Pressures are measured with the ionization gauge of 

 Montgomery and Montgomery ( 1938) employing a No. 47 radio 

 tube of Radio Corporation of America. A portable power pack is 

 used so that it can be employed with several vacuum systems to 

 obviate duplication of expensive meters. A pressure of 10"^ mm. 

 mercury or less is sufficient for the electron microscope, and the 

 sensitivity of the pressure gauge is about 7 X 10"^ mm. mercury per 

 microampere ion current. 



Manipulation 



Preliminary experiments with salts of sodium, potassium, calcium, 

 magnesium, and iron demonstrated that very bright images were 

 formed on the screen by calcium and magnesium, weak ones by 

 sodium and potassium, and none by iron. By maintaining a cathode 

 temperature of 700-800° for an hour the sodium and potassium 

 were volatilized so that the bright image could be safely assigned 

 to calcium and magnesium. For these experiments, ashless gelatin 

 impregnated with the chlorides was hardened in 10% formalin, 

 dehydrated in alcohols and embedded in paraffin. Paraffin sections 

 ( 10 fi.) were placed directly on the prepared cathode. 



The nickel cathode is prepared for use by polishing with optical 

 rouge and washing with water and nitric acid. It is then coated 

 with a mixture of 40% barium carbonate and 60% strontium 

 carbonate in a 2% solution of nitrocellulose in amyl acetate. The 

 barium-strontium mixture serves to increase the emission of the 

 calcium and magnesium in the tissue by about 1000%, by activa- 

 tion, and at the same time the mixture emits electrons itself to give a 

 contrast background on the screen. When completely dry, a 10 /x 

 section of tissue embedded in paraffin, prepared by the freezing- 

 drying technique (see page 3), is placed on this surface and 

 smoothed down with a steel needle. The cathode is then inserted into 



