458 



METHODS AND FORMULAS 



DS 22.8-DS 23.10 



22.8 Wallace 1931 19938, 74:369 



REAGENTS REQUIRED: A. Any DS 11.122 formula; B. 0.5% eosin Y; C. water 45, aniline 



45, abs. ale. 10, methyl violet to sat; D. ADS 12.2 Lugol (1905) ; E. aniline 30 xylene 60 

 method: [5 n sections of F 3700.1010 fixed material] -^ A, till nuclei lightly stained -^ 



wash -^ B, 30 sees. — > thorough wash — > C, 2 hrs. -^ wash — > D, 10-15 mins. — > wash 



-^ E, till differentiated —> xylene wash -^ balsam 

 RECOMMENDED FOR: basal bodies of cilia. 



23. SELECTIVE STAINS FOR SPECIFIC ORGANISMS 



There are surprisingly few stains which 

 will selectively stain a specific organism 

 compared to the number of stains which 

 have been proposed for that purpose. 

 Bacteria, for example, may be very well 

 stained by any nuclear staining tech- 

 nique; therefore here are given (DS 23.2) 

 only those methods for staining them 

 which have, by convention and custom, 

 come to be supposed to be "bacterio- 

 logical methods." 



One is on even less secure ground in 

 dealing with the first class (DS 23.1), in 

 which are given those stains which stain 

 virus, Rickettsiae, and Negri bodies, for it 

 is, of course, impossible that any individ- 

 ual virus molecule could be stained in such 

 a manner as to bring it within the range of 

 an optical microscope. But the methods 

 given in the division DS 23.11 for stain- 

 ing minute unidentified bodies may quite 

 possibly represent aggregates of virus, as 

 they were supposed to do by the inventors 

 of the techniques. 



The third division here given (DS 23.3) 

 is for the differential staining of parasites 

 in host tissues, to which much attention 

 has been devoted, and for which a number 

 of relatively specific methods have been 

 proposed. The last two divisions (DS 

 23.4, DS 23.5) contain a few staining 

 techniques of such speciaUzed application 

 that they cannot possibly be given else- 

 where in the present work. 



23.1 Virus, Rickettsiae, and 

 Negri Bodies 



23.10 typical examples 



Demonstration of Rickettsiae in the 

 scrotum of a guinea pig using the 

 magenta-thionin stain of 

 Macchiavello 1938^ 



It is presumed, in the description which 

 follows, that the technician is acquainted 



with the exceedingly dangerous nature of 

 Rickettsiae and is prepared to take all 

 those aseptic precautions through which 

 he or she may be protected against infec- 

 tion. Even then it must be emphasized 

 that Rickettsiae can only be handled in 

 properly equipped laboratories and under 

 the supervision of a skilled bacteriologist 

 or pathologist. 



The staining method here described 

 (DS 23.12 Macchiavello 1938) is the basic 

 method which has been adapted by 

 numerous people, who usually refer to 

 their own method as "Maccliiavello." 

 The volume of Macchiavello 1938 is, more- 

 over, not readily available and most work- 

 ers have derived their information as to 

 the method from the pubhcation of Cox 

 1939 (17302,53:2242) to whom this 

 method is therefore frequently attributed. 

 Rickettsiae are difficult to stain satis- 

 factorily, and though it is suggested that 

 the beginning technician attempt this 

 original technique of Macchiavello for the 

 first preparation, it is very probable that 

 he will have to vary the technique to suit 

 the materials which he is using. 



The only difficulty likely to be en- 

 countered will be by technicians who are 

 accustomed to staining bacteria by some 

 method in which every effort is made to 

 get these bodies free on the slide without 

 a great deal of cell debris around them. 

 Rickettsiae, on the contrary, cannot be 

 satisfactorily stained and differentiated 

 unless they are within a cell, hence the 

 critical stage of the preparation consists 

 in securing undamaged cells containing 

 Rickettsiae in a layer thin enough to 

 permit their examination with the highest 

 powers of the microscope. 



Rickettsiae may be recovered either 

 from the walls of the blood vessels or from 

 the subcutaneous tissues of almost any in- 

 fected animal. The guinea pig has been 

 selected for the present demonstration be- 



