10. Mix the contents of the loop thoroughly with the drop of water on the 3 by 1 slide. 11. Again sterilise the loop as directed in step 4, and replace it in its stand. 12. Remove a cover-slip from the glass capsule by means of the cover-slip forceps, rest it for a moment on its edge, on a piece of filter paper to remove the excess of alcohol, then pass it through the flame of the Bunsen burner. This burns off the remainder of the alcohol, and the cover-slip so "flamed" is now clean, dry, and sterile. 13. Lower the cover-slip, still held in the forceps, on to the surface of the drop of fluid on the 3 by 1 slip, carefully and gently, to avoid the inclusion of air bubbles. 14. Examine microscopically (_vide infra_). During the microscopical examination, stains and other reagents may be run in under a cover-slip by the simple method of placing a drop of the reagent in contact with one edge of the cover-glass and applying the torn edge of a piece of blotting paper to the opposite side. The reagent may then be observed to flow across the field and come into contact with such of the micro-organisms as lie in its path. The non-toxic basic dyes most generally employed for the intra-vitam staining of bacteria are Neutral red, } Quinoleine blue } Methylene green } in 0.5 per cent. aqueous solutions. Vesuvin, } _Negative Stain_ (Burri).--By this method of demonstration the appearances presented by dark ground illumination (by means of a paraboloid condenser) are closely simulated, since minute particles, bacteria, blood or pus cells etc. stand out as brilliantly white or colourless bodies on a dark grey-brown background. _Reagent required:_ Any one of the liquid waterproof black drawing inks (Chin-chin, Pelican, etc.). This is prepared for use as follows: Measure out and mix: Liquid black ink, 25 c.c. Tincture of iodine 1 c.c. Allow the mixture to stand 24 hours, centrifugalise thoroughly, pipette off the supernatant liquid to a clean bottle and then add a crystal of thymol or one drop of formalin as a preservative. METHOD.-- 1. With the sterilised loop deposit one drop of the liquid ink close to one end of a 3 by 1 slide. 2. With the sterilised loop deposit a drop of the fluid culture (or of an emulsion from a solid culture) by the side of the drop of ink (Fig. 69, a); mix the two drops thoroughly by the aid of the loop. 3. Sterilise the loop. 4. Hold the slide firmly on the bench with the thumb and forefinger of the left hand applied to the end nearest the drop of fluid. 5. Take another clean 3 by 1 slide in the right hand and lower its short end obliquely (at an angle of about 60°) transversely on to the mixed ink and culture on the first slide, and allow the fluid to spread across the slide and fill the angle of incidence. 6. Maintaining the original angle, draw the second slide firmly and evenly along the first toward the end farthest from the left hand (Fig. 69, b).