Appendix. 209 



Gelatin. Agar. 



7. Warm on water bath, stirring till pepton and 

 gelatin are dissolved and not allowing the 

 temperature to rise above 60 C. 



8. To 500 g. of the meat infusion add 



500 c.c. of the three per cent agar, 

 keeping the temperature below 

 60 C. 



9. Heat over boiling water (or steam) bath for 

 30 minutes. 



10. Restore loss 'by evaporation. 



11. Titrate, after boiling one minute to expel car- 

 bonic acid. 



12. Adjust reaction to + i.o per cent by adding 

 normal hydrochloric acid or sodium hydrate 

 as required. 



13. Boil two minutes over free flame, constantly 

 stirring. 



14. Make up loss by evaporation. 



15. Filter through absorbent cotton and cotton 

 flannel, passing the nitrate through the filter 

 until clear. 



1 6. Titrate and record the final reaction. 



17. Tube, using 10 c.c. of medium in each tube. 



1 8. Sterilize five minutes in the autoclave at 120 

 degrees, or for 30 minutes in streaming steam 

 on three successive days. Put the gelatin at 

 once into ice-water till solidified. 



19. Store in the ice-chest in a moist atmosphere, 

 to prevent evaporation. 



LACTOSE (OR DEXTROSE) LITMUS AGAR. 



Lactose or dextrose litmus agar shall be prepared in the 

 same manner as nutrient agar, with the addition of one 

 per cent of lactose (or dextrose) to the medium just before 

 sterilization. The reaction shall be made neutral to 

 phenolphthalein. 



