essential element. The reaction is now determined and adjusted to the desired end point, “standardized,” as it is called. The medium is again _thoroughly_ boiled and filtered boiling hot. The adjusting of the reaction and the boiling ordinarily cause a precipitate to form which is largely phosphates of the alkaline earths with some protein. The filtered medium is collected in suitable containers, flasks or tubes, which are plugged with well-fitting non-absorbent cotton plugs and sterilized, best in the autoclave for twenty minutes at 15 pounds pressure, or discontinuously in streaming steam at 100°. If careful attention is paid to _titration_ and to _sufficient boiling_ where indicated, the meat broth prepared as above should be clear, only faintly yellowish in color and show no precipitate on cooling. The conventional method for standardizing an acid medium is as follows: Take 5 cc of the medium, add 45 cc of distilled water and 1 cc of _phenolphthalein_ as indicator. Boil the solution and while still hot run in from a burette N/20 NaOH solution until a faint pink color appears. From the number of cc of N/20 NaOH used to “neutralize” the 5 cc of medium it is calculated how many cc of N/1 NaOH are necessary to give the desired end reaction to the volume of medium which is to be standardized. The resulting reaction is expressed as % _acid or alkaline to phenolphthalein_. If it is necessary to add to each 100 cc of the medium 1 cc of N/1 NaOH to make it neutral to phenolphthalein, the reaction is called 1% acid: if to each 100 cc of medium there is added 1 cc of N/1 alkali in addition to the quantity necessary to neutralize, the reaction is called 1% alkaline. In order to obtain a pink color when titrating with this indicator not only must the “free acid” be neutralized by the alkali but also loosely combined acid and any other substances present which will combine with the alkali rather than with the indicator so that in many media _more alkali_ is added than is necessary to neutralize the “free acid,” _i.e._, the free H ions present. It is well established that the controlling factor in the growth of bacteria in so far as “reaction” is concerned is not the _titratable substances_ present but only the “free acid,” _i.e._, the _number of free H ions_, consequently it is better to determine the concentration of H ions and to _standardize to a definite H ion concentration_. Phenolphthalein as shown above is not a good indicator for this purpose. The H ions present can be determined accurately in all cases only by electrolytic methods. The apparatus necessary is usually relatively expensive and scarcely adapted to the use of large classes of students. There are a number of indicators each of which will show color changes within rather narrow ranges of H ion concentration. Standardization by the use of these indicators, the “colorimetric method,” is recommended by the Society of American Bacteriologists and is coming into general use.