804 PROFESSOR W. C. M'INTOSH AND MR E. E. PRINCE ON 



The plan followed by Kupffer (His and Braune's Archiv Anat. Ahth., 1882), Henneguy (Bull 

 de la Soc. Philom., Paris, 1879, pp. 75-77), and others involve too many processes to be adopted when 

 the species studied are numerous, and the quantity of material is large. 



Various circumstances conduce to render Teleostean eggs difficult objects for treatment — not 

 only on account of their small size, pelagic ova being rarely more than a millimetre in diameter — 

 but the tough nature of the capsule and fluidity of the contents render the removal of the former a 

 most delicate and hazardous task. If hardened before the capsule is removed, shrinking results ; 

 and if the capsule be removed before hardening, the egg is more or less disorganised, unless the 

 operator be very fortunate. The best sections are those gained by leaving the egg almost intact, 

 and by hardening, staining, and imbedding in toto, but this plan is beset by many dangers. On 

 removing the egg from the sea-water, and reference is made here to marine ova solely, the capsule 

 is carefully pierced in order to facilitate the admission of the various media into which it is to be 

 transferred. Save for this puncture, the egg is left entire, and thus it is passed through all the 

 processes of killing, hardening, staining, clearing, and imbedding. 



The paraffin method proved to be the only practical one, other methods, such as imbedding 

 in pith, which might serve for large eggs, such as those of the Salmonidae, were unsuitable 

 for eggs so small and frail as those of the Gadoids, Pleuronectidce, &c* Various forms of the 

 microtome were used in preparing the extensive series of sections of the various Teleosteans 

 considered in these pages — the rocking microtome of the Cambridge Scientific Instrument Company 

 being found very useful. The large Caldwell microtome, used in the classes of zoology at the 

 United College, and kindly lent by the authorities of the University of St Andrews, was of great 

 service ; while the Jung (Thoma's) microtome was found to be well adapted for older stages of the 

 embryos, and for adult ovaries — a series of sections being cut by Dr Scharff. The sweeping 

 motion of the last-named instrument proved very efficient in cutting through the more mature 

 skeletal and other tissues of young fishes, to which task the fixed razor of the English microtomes 

 proved unequal — refusing, in fact, to pass through the firm connective and cartilaginous elements. 



II. Killing, Fixing, and Hardening — Corrosive Sublimate. — The saturated solution is one of 

 the most efficient killing and fixing fluids available in the laboratory, and it kills, fixes, and hardens 

 so rapidly that Teleostean ova require to be left in it for a very short time. As soon as the 

 penetration of the fluid is complete, they are removed and washed in dilute alcohol, rather than in 

 distilled water. Washing must be well done, in order to prevent subsequent deposition of crystals 

 in the tissues. The desirability of staining, clearing, and cutting after treatment with this fluid is 

 too well known to require any explanation — the best preparations being found to be those in 

 which, after killing and fixing, the subsequent operations are immediately proceeded with. A mix- 

 ture of two parts corrosive sublimate and one part acetic acid was found to be most serviceable. It 

 is a powerful killing and fixing fluid, and produces the best results. For killing, two or three 

 minutes usually suffice, and washing is then done in very weak alcohol — the alcohol being 

 frequently changed until the killing medium is wholly extracted, and graduated alcohols follow, 

 viz., 30, 40, 50, and 60 per cent. 



Picro-sulpJmric Acid (Kleinenberg). This useful killing and fixing fluid does not produce 

 the best results, since it frequently causes the blastomeres in early stages to expand and burst the 

 capsule, thus entirely disorganising the embryonic structures. Whitman experienced the same 

 results (op. cit, p. 152), but occasionally this effect is not produced, and, if successfully killed and 

 hardened in this fluid, ova are often found to produce most satisfactory sections. It is, however, not 

 reliable. Creosote is added on Kleinenberg's suggestion, but apparently without much effect. If the 

 ova placed in picro-sulphuric acid maintain their normal shape, they remain four or five hours, and 

 then are transferred into 70 per cent, alcohol, which is frequently changed, as it becomes stained by 

 the yellow picric acid. When the alcohol is seen to be uncoloured, the ova are then ready to 

 be transferred to absolute alcohol, preparatory to clearing. Emery recommends this fluid for 



* Henneguy used elder-pith soaked in alcohol and covered with a layer of collodion. 



