MICROSPECTROSCOPY AND MICROSPECTROPHOTOMETRY 



255 



In the method just described the specimen is illuminated simul- 

 taneously by all the radiations of the spectrum, this being incon- 

 venient from the stray-light standpoint. Nevertheless, the method is 

 suitable in some cases, e.g. when investigating a transient phenomenon. 

 Yet, whenever feasible, the methods in which the object is illuminated 

 in monochromatic hght are to be preferred. 



4. MICROSPECTROPHOTOMETRIC MEASUREMENTS 



As in conventional spectrophotometry, quantity determination and 

 identification of a substance can be made by microspectrophotometry. 



In one instance, the spectrum recorded by one of the previous 

 methods is compared with the spectra of known substances, spectra 

 derived from the selfsame instrument. When the detail investigated 

 exhibit a typical spectrum, this is readily carried out. 



d 04 



A =0-405/^ 



Fig. 10.12. Absorption curve of an erythrocyte from a newt (After Barer, Holiday 



and Jopes). 



Figure 10.12 shows, according to Barer, Hohday and Jopes, the 

 absorption (absorbance) curve of an erythrocyte (blood red cell com- 

 prising a nucleus) from a newt, showing this erythrocyte schematically 

 (Fig. 10.13) the absorbance measurement is effected, for instance, 

 along the hne ox at a specific wave-length (A = 405 /<)• The areas AB 

 and CD, in Figs. 10.12 and 10.13, correlate the cytoplasm; and the 

 area BC the nucleus, respectively. A family of curves, similar to the 

 one shown in Fig. 10.12, can be plotted for all the spectrum wave- 

 lengths. Absorption variation versus the wave-length, taking place at 

 specific points of the specimen, can also be determined. In both cases 

 the curves in Fig. 10.14 show the outcome. The curve N shows the 



