MICROTOME 



209 



MILK 



solved material by plasmolysing the 

 cell to various degrees and thereby al- 

 tering the water content of the solution. 

 This method permits comparison of the 

 optical density of the vacuole at various 

 volumes and provides a direct test of 

 Beer's law. In the case of solid struc- 

 tures such as the nucleus the osmotic 

 method is probably invalidated by the 

 fact that the absorbing material is not 

 freely dissolved throughout the struc- 

 ture. 



When the cytological object contains 

 oriented absorbing material in signifi- 

 cant amounts, considerable departure 

 from the Beer-Lambert relationship is 

 to be expected (Commoner, B., Science, 

 1949, 110, 31-40). It may be possible 

 to avoid this difficulty by the use of a 

 polarized incident light beam. 



6. The effect of lateral inhomogeneity. 

 This problem has been discussed from 

 a theoretical point of view (Glick, D., 

 Engstrom, A. and Malmstrom, B. G., 

 Science, 1951, 114, 253-258; Danielli, 

 J. F., Cold Spring Harbor Symp., 1949, 

 14, 32-39). It is apparent from these 

 discussions that large errors may arise 

 from this effect but at present there is 

 no experimental way of determining 

 their magnitude. In general it would 

 seem essential that objects studied be 

 homogeneous at least with respect to 

 their microscopic appearance. 



6. The determination of absolute quan- 

 tities. In some instances (Caspersson, 

 T. and Schultz, J., Proc. Nat. Acad. 

 Sci., 1940, 26, 507-515) investigators 

 have calculated the absolute content of, 

 say, nucleic acid in a nucleus from meas- 

 urements of the optical density of the 

 nucleus and the specific extinction of 

 nucleic acid solutions. Such a proce- 

 dure has never been validated by the 

 customary tests of the analyst. 



7. The effect of ultraviolet light on the 

 optical properties of the cell. One of the 

 most serious difficulties encountered in 

 the microspectrophotometric work is 

 the fact that exposure to ultraviolet 

 light is damaging and frequently lethal 

 to the object. At the same time irra- 

 diation seriously alters the absorption 

 properties of the cell. It was first 

 shown by Brumberg, E. M. and 

 Larionov, L. P. (Nature, 1946, 158, 

 663-664) that in living cells undamaged 

 by ultraviolet the optical density of the 

 nucleus does not exceed that of the 

 cytoplasm. When the cell has been 

 killed by ultraviolet radiation the opti- 

 cal density of the nucleus rises sharply 

 whereas the absorption of the cytoplasm 

 drops. Similar changes have now been 

 observed by a number of other workers 

 (Bradfield, J. R. G., Discussions of the 



Faraday Society, 1950, No. 9, 481-490; 

 Walker, P. M. B. and Davies, H. G., 

 Discussions of the Faraday Society, 

 1950, No. 9, 461-470). These observa- 

 tions show that 1) absorption measure- 

 ments of living cells must be made on 

 cells which actually survive the ex- 

 perimental procedure, 2) fixation pro- 

 cedures may seriously alter the apparent 

 distribution of ultraviolet absorbing 

 material and 3) the structural ar- 

 rangement peculiar to the living nu- 

 cleus probably has a significant effect 

 on its absorption properties. 



Microtome. The freezing, rotatory and 

 sliding microtomes are well known and 

 advertised. The high speed micro- 

 tome required for cutting especially 

 thin sections for Electron Microscopy 

 is essential for research. See account 

 by G. H. Scott in McClung's Micro- 

 scopical Technique, 1950, p. 720; also 

 FuUam, E. F. and Gessler, A. E., Rev. 

 Sci. Inst., 1946, 17, 23 and Gessler, A. E. 

 and Fullam, E. F., Am. J. Anat., 1946, 

 78, 245. Geren, B. B. and McCuUoch, 

 D. (Exp. Cell Research, 1951, 2, 97-102) 

 have made adjustments in the Minot 

 rotary microtome by which sections of 

 tissues, prepared by the methacrylate 

 embedding technique of Newman, S. 

 B., Borysko, E. and Swerdlow, M. 

 (Science, 1949, 110, 66) can be cut 1/20 

 n in thickness. The edges of some sec- 

 tions are much thinner. Glass knives 

 with water trough (Latta, H. and Hart- 

 mann, J. F., Proc. Soc, Exp. Biol, and 

 Med., 1950, 74, 436) are a great improve- 

 ment over steel ones. 



Microtome Knife, sharpening. There is no 

 easy method. Care and long practice 

 are essential. (See Bensleys, p. 57.) 

 For the usual oil and water stones a 

 ground glass is now sometimes substi- 

 tuted (Uber, F. M., Stain Techn., 1936, 

 11,93-98). 



Micrurgical Technique (Gr. micros, small 

 + ergon, work) is referred to under the 

 heading of microdissection. 



Mikado Yellow G (CI, 622)— Stilbene Yel- 

 low — a direct dye of light fastness 3. 

 Similar to Sun Yellow but lighter in 

 color (Emig, p. 46). 



Milk, bacteria in, a modification of Newman 

 technic (Broadhurst, J. and Paley, C, 

 J. Am. Vet. Med. Assoc, 1939, 94, 

 525-526). To prepare stain add 0.4 cc. 

 cone. H2SO4 to 54 cc. 95% alcohol. 

 Mix with 40 cc. technical tetrachlor- 

 ethane in flask and heat to 55 °C. but no 

 higher. Add about 1.0-1.2 gm. methy- 

 lene blue while mixture is still hot. 

 Shake until dye goes into solution. 

 Then add 8.0 cc. 1% basic fuchsin in 

 95% alcohol. Mix, cool, filter and put 

 up in glass stoppered bottle. Spread 



