54 FIXATION 



salt-linkages with the carboxyl side-groups of certain amino- 

 acids. 



These reactions appear not to have a strongly coagulative effect, 

 but the mercury is firmly held. This can easily be shown by test- 

 ing with mercuric potassium iodide, Hgl2.2KI, a useful reagent 

 for distinguishing between mercuric ions or loosely held mercury 

 on one hand, and the firmly held metal on the other. The reagent is 

 yellow, but gives the red colour of mercuric iodide in the presence 

 of the ions or loosely held metal. 



A different kind of combination is characteristic of weakly 

 alkaline, neutral, or weakly acid conditions down to the iso- 

 electric point of the protein. The mercuric chloride is taken up as 

 a whole molecule through subsidiary valencies, mainly by the 

 amino side-groups of lysine and arginine, and is now held very 

 loosely, so that a red reaction is given with mercuric potassium 

 iodide. The formation of this loose compound is associated with 

 coagulation. Coagulation is opposed by sodium chloride. This is 

 because the chloride ions have a stronger affinity for mercuric 

 chloride than the protein has. The anion [HgCl4]" is formed. ^^^ If 

 albumin is coagulated by mercuric chloride and washed, the clot 

 is reaily dissolved by a saturated solution of sodium chloride (or 

 potassium iodide). 



On the acid side of the iso-electric point of the protein, but still 

 within the range of weak acidity, the ion [HgClJ" combines 

 through main valencies with the amino-groups of the protein, 

 which are ionized to some extent on the acid side of the iso- 

 electric point. The compound formed is again very loose, but 

 coagulation is now promoted by the presence of sodium chloride, 

 which increases the amount of the reactive mercuric ion. 



In conditions of strong acidity mercuric chloride does not react 

 with proteins, except to form mercaptide ; there is little tendency to 

 coagulation. 



In practical microtechnique, acids and sodium chloride are 

 often added to solutions of mercuric chloride, apparently without 

 much consideration of the complex consequences. Tissues that 

 have been fixed with mercuric chloride are sometimes placed in 

 Lugol's solution. This is likely not only to decompose the mer- 

 captide with oxidation of the former sulphydryl groups to di- 

 sulphide,^^^ but also to dissolve the coagulum. 



If mercuric chloride were to block all -NHg groups by combining 

 with them, but left the -COOH groups untouched, the protein 



