362 RADIOISOTOPES IN BIOLOGY AND AGRICULTURE 



of chromatographic separations (4, 7, 8). At the present time, however, 

 the practice is primarily based upon arbitrary experimental conditions. 

 The resolution of substances, although chiefly dependent upon partition 

 between solvents, may also be affected by surface adsorption and ion 

 exchange. Attention is called to the widely used constant Rf, which is 

 defined as follows: 



P _ movement of band 



^ movement of advancing front of liquid 



Rf is easily determined in paper chromatograms. Absolute values of Rf 

 have little meaning, since they are dependent upon solvent composition, 

 temperature, pH, presence of salts, and other factors. The primary use 

 of a listing of Rf values, determined under similar conditions, is to indicate 

 whether or not the separation of a given mixture is possible; or, if there are 

 data for several solvents, the listing permits a choice of the most suitable 

 solvent. 



GENERAL METHODS 



Chromatograph Chamber. The primary requirement is a mechanical 

 arrangement that permits one end of a filter-paper strip or sheet to be 

 immersed in the solvent. It is also necessary to keep the atmosphere 

 saturated at all times with the solvent, since otherwise the capillary flow 

 would cease because of evaporation. Apparently it makes little differ- 

 ence whether the solvent ascends, descends, or does both, as described by 

 Block et al. (8). Commercial chambers are available, as well as auxiliary 

 items such as solvent troughs and paper racks. However, homemade 

 equipment may serve ec}ually well. Figure 8-1 illustrates a simple assem- 

 bly that has been found satisfactory for strips. An ordinary glass cyl- 

 inder is placed within a cylindrical jar which can be covered with a glass 

 plate to make a reasonable seal. A clear glass pipette washer or hydrom- 

 eter jar with a flanged neck is satisfactory for the cylindrical jar. A his- 

 tological staining dish is placed on top of the inner cylinder, as shown, to 

 hold the solvent, and the paper strips are held in place, with an end sub- 

 merged in the solvent, by means of microscope slides (13). This arrange- 

 ment provides for a descending flow of solvent. However, the ascending 

 technique can be employed with a simple modification in which the inner 

 cylinder is not used and the strips are supported from a rack attached to 

 the cover plate, with the bottom of the strip dipping into the solvent. 



Figure 8-2 illustrates a simple cabinet in which sheets can be used. It 

 is necessary only to have a box large enough to accommodate the size of 

 paper to be used, with some device for holding the sheets. With this 

 apparatus, solvent troughs about 30 in. long are supported on the cleats 

 near the top of the box. The inside of the box is lined with glass plates, 



