352 TISSUE CULTURE 



some attempt should be made to work with approximately constant 

 dilutions of the embryonic juice in Tyrode solution. Fischer 

 adopts the method of removing the concentrated juice after the 

 first centrifuging and replacing it with a volume of Tyrode solution 

 equal to that of the remaining pulp, which is then well mixed with 

 a glass rod or pipette and returned to the centrifuge. The second 

 extract is particularly suitable for hanging drop cultures of rapidly 

 growing cells such as fibroblasts. A third, weaker extract, useful 

 for maintaining organotypic growth in a so-called growth restrict- 

 ing medium, is prepared by repeating the process of adding another 

 volume of Tyrode and re-centrifuging. The concentrated extract 

 is used principally in varying dilutions of Tyrode solution for flask 

 cultures. The worker must ascertain, by experiment, however, 

 which type and concentration of extract is most suitable for the 

 particular tissues to be cultivated and a clear description of the 

 method of preparing the extract should be included in the account 

 of tissue culture experiments. 



Embryonic extract does not keep well even in the refrigerator 

 and should be prepared fresh for each batch of cultures. Its 

 growth promoting principle appears to be destroyed by heating 

 over 56° C. Nevertheless heat inactivated extract is sometimes 

 used where it is desirable to suppress the growth of connective 

 tissue cells and to favovn- the migration of epithelia (see Doljanski, 

 Arch. f. exper. Zellforsch., xi, 1931, p. 261). Berkefeld sterilisation 

 of embryonic extracts is not to be recommended. 



785. Tissue Extracts. No extract from adult tissues and 

 organs has been found to possess the same power of stimulating 

 the growth of tissue cultures as embryonic extract ; but, in special 

 instances, better initial growth of an organ has resulted if the 

 medium contains also the juice of that organ (see Amoroso, 

 Archiv. f. exper. Zellforsch., xii, 1932, p. 274). A useful table 

 indicating the j^H of tissue extracts from a wide variety of 

 laboratory animals should also be consulted (Kutscherenka and 

 SoLowiEW, Biol. Gen., ii, 1920, p. 523). 



For special growth-promoting tissue extracts, see Carrel, 

 Journ. Exper. Med., xxxvi, 1922, p. 385 (leucocytes), and for 

 artificial substitutes for einbryonic extract, consult Carrel and 

 Baker, Journ. Exper. Med., xliv, 1926, p. 503. 



786. Preparation of Cultures. If possible, the setting up of 

 cultures in rooms used for other purposes should be avoided. The 

 use of glass boxes of various designs with holes for inserting the 

 arms are a hindrance to speed and to the line dissection of small 

 embryos under sterile conditions. The reader will find several 

 excellent plans for laying out a tissue culture room in the text- 

 books recommended at the end of the chapter. It should be noted 

 that the simplest possible arrangement of the apparatus combines 

 to give speed and accuracy as well as complete sterility. Select a 



