694 GENERAL TECHNIQUES FOR CLASSES OF PLANTS 



p. 287 (flagellates); Skinner, Plant Physiol, vii., 1932, p. 533 (soil 

 algae). 



Pure cultures are frequently essential, especially for the simple 

 unicellular and colonial forms. Numerous methods have been 

 devised. Due regard must be paid to the sterilisation of all 

 vessels, media, etc., but it is usually difficult to obtain such 

 cultures free from admixtures of bacteria. 



See Beijerinck, Bot. Zeit. xlviii., 1890, p. 725; Chodat and 

 Grintzesco, C. R. Congr. int. Bot., Paris, 1900, p. 157; 

 Grintzesco, Bull. Herb. Boissier, ser. 2, v., 1902, p. 225; Klebs, 

 p. 184 (above); Richter, Ber. Deutsch. bot. Ges. xxi., 1903, p. 493 

 (diatoms) ; Wettstein (above). 



Reference should be made to the work of Klebs and others for 

 methods of inducing the reproductive phases. 



1378. Fixation. Most are adapted to methods applicable to 

 filamentous specimens. Imbedding should be by very gradual 

 stages. Mats of filaments, crusts of filamentous and unicellular 

 types on sticks and stones, and epiphytes and endophytes should 

 be fixed and stained undisturbed. Tease, scrape from the sub- 

 stratum and dissociate as necessary just before mounting. 



Shrinkage of the protoplast in filamentous forms is most 

 troublesome. To eliminate, vary the amounts of the various 

 constituents of the fixative, increasing the acetic acid until 

 shrinkage is overcome. Handle unicellular and other small forms 

 by sedimentation methods. Or, place a drop of suspension on an 

 albumened slide, fix with the vapour of osmic acid for a few seconds 

 to one minute. Dry the drop and stain. If the specimens are 

 rare, pick them out with a fine pipette. Chamberlain (1932, 

 p. 228) handles small amounts of planktonic forms rolled up in a 

 tube made of a strip of epidermis from an inner scale of an onion ; 

 the ends of the tube are tied. Cut off the ends when ready to 

 mount. Material can be imbedded and sectioned in this way. 

 See also Conger, J. Roy. Micr. Soc, 1925, p. 48. 



Bold {Bull. Torrey. Bot. Club, Ix, 1983, p. 241) fixes Protosiphon 

 (coenocytic), grown on agar, in situ, by placing drops of fixative 

 on the plants with a pipette. Then pour melted agar, near the 

 point of gelation over the fixed plants. When the agar has set, 

 cut out blocks containing the alga and imbed in paraffin. 



Handle massive forms (Siphonocladiales, etc.) like soft tissues 

 of higher plants. Decalcify if calcareous. 



Fix in 4 per cent, formalin, chromo-acetic, Bouin or formalin- 

 acetic-alcohol for habit and grosser cell structure. G. H. C. 

 {Turtox News, iii, p, 45) recommends Rawlin's formol-acetic- 

 alcohol (see Rawlins, p. 14), in which Cladophora alone sometimes 

 shows plasmolysis. Evan's, Keefe's or Wood's {Science, Ixx, 

 1929, p. 637) fluids will preserve green algae in their natural colour. 

 West {ex Nieuwland, Bot. Gaz., xlvii, 1909, p. 237) fixes pre- 



