Microtome Sectioning of Material in Paraffin 43 



have a grainless or very fine-grained texture, should be free from 

 bubbles and opaque spots, and should contain no grit or other debris. 



PROPER INFILTRATION 



Improperly infiltrated material breaks out of the paraffin block. 

 Examine the cut face of the material with a hand lens or a binocular 

 dissecting microscope. Crumbling within the tissues may indicate 

 inadequate penetration by paraffin during infiltration or may be 

 the result of excessive hardness or brittleness of the tissues. Breaking 

 out of entire sections from the paraffin ribbon indicates poor ad- 

 hesion between external surfaces of the piece of tissue and the 

 paraffin. Inadequate infiltration may be due to incomplete dehydra- 

 tion or excessively rapid infiltration. The remedy lies in rcinfiltration. 



ORIENTATION OF THE MOUNTED MATERIAL 



The paraffin around the piece of material should be trimmed 

 rectangular, with the material approximately centered laterally in the 

 paraffin. If the tissue is not vertically centered in the paraffin, the 

 thicker layer of paraffin should be at the top, affording support against 

 the pressure of the cutting action (Fig. 6.2 F) . Trim the upper and 

 lower edges of the wax so that the sections are close enough to each 

 other on the slide for efficient use of the slide and cover glass. See 

 Fig. 6.6 for efficient placement of sections. The edge which ap- 

 proaches the knife should be parallel to the knife (Fig. 6.3 C) . For 

 most paraffin sectioning the knife is placed at right angles to the 

 vertical motion of the paraffin block. The other angle to be con- 

 sidered is the declination, or the tilt of the flat face of the knife 

 toward the tissue (Fig. 6.3 A, B) . This angle must be determined by 

 trial. 



RIGIDITY OF MOUNTING 



The piece of tissue should be firmly attached to a mounting block 

 or disk and supported, especially on the edge away from the knife, 

 by a generous layer of paraihn (Fig. 6.2 E) . The mounting block, the 

 knife, and the knife carrier must be firmly clamped into place. Inade- 

 quate rigidity of the tissue mounting or of the knife results in alter- 

 nate sections of unequal thickness. This can often be recognized in 

 the ribbon but usually becomes evident during staining. The thicker 

 sections will be more deeply stained than the alternating thin ones. 

 In sections of a large stem there may be alternate deeply stained and 

 lightly stained bands in each section. 



