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A total of 31 individual ovine milk samples, ranging between 30 and 2600 x 10(3) cells/mL, were divided into 8 aliquots/milk with the objective of studying the overall accuracy of 2 rapid somatic cell count (SCC) counters, one based on cytometry on disk (Fossomatic 360) and the other on flow cytometry (Fossomatic 5000), under 4 types of preservation (without preservation, bronopol, sodium azide, and potassium dichromate) and 2 analytical temperatures (40 and 60 degrees C). All analyses were carried out in duplicate. In addition, each sample was analyzed in quadruplicate by reference microscopic method using Pyronin Y-methyl green as a stain. A second experiment using 13 samples divided into 20 aliquots/ sample, enabled repeatability to be studied depending on the values obtained in the SCC and on the SCC equipment used. Comparison of the methods was based on repeatability and accuracy studies (means comparison and regression studies vs. reference method). Both counters gave adequate repeatability and accuracy values in ovine milk, though the SCC obtained by Fossomatic 5000 was closer to the reference method and was somewhat more repeatable than Fossomatic 360. In the regression study, slope and intercept values were statistically different from their theoretical values (1.00 and 0.00, respectively) in the unpreserved samples but not in the preserved ones. In all cases, correlation coefficients very close to 1.00 were obtained. The preserved milk analyzed by flow cytometry gave optimal repeatability values (s(r) = 16.3 to 19.7 and s(r)% = 1.9 to 2.4), and their logSCC means (5.62 to 5.64) were not different from the reference value (5.63). Bronopol was the optimal preservative for the Fossomatic method. Analytical temperature did not contribute significantly to SCC variation, although disk cytometry gave slightly more repeatable SCC at 40 degrees than at 60 degrees C.

In studying butter, Redfield ~ has proposed a routine microscopic method of counting yeasts and oidia as correlated in some degree at least with the quality of the cream used in its manufacture. This method is based upon the examination of the curd-whey mixture separating out when the sample of butter is melted at 45°C. One-cubic centimeter samples of this mixture are removed to sterile containers and smears are made by transferring 0.01 co. samples of this mixture to slides with the Breed type of pipette, for microscopic study. The results shown in table 1 are selected from experiments made to test the accuracy of this method. These results based on the number of yeasts found in 10 microscopic fields, check as closely as those usually obtained by the official plate method, and therefore indicate that yeasts are fairly evenly distributed throughout the butter. Counts of this kind, check fairly closely when the totals are high as in these samples. 1V~uch greater differences occur in successive examinations of samples containing very low numbers of yeasts. These counts were based on numbers found in 100 microscopic fields. Each cell found was counted. The results indicate that yeasts are somewhat more evenly distributed than are the oidia. The mycelial character of Oidium lactis tends to result in more uneven distribution of the

Summary The results of this statistical analysis of counts made by the direct microscopic and agar plate methods show a high degree of variability in both methods, as applied comparatively to rating raw market milk supplies. In other words, there is such a relatively large experimental error in both methods that sanitarians interested in the bacteriological control of milk supplies must avoid the danger of making too fine distinctions in establishing the maximum numerical limits for the various, so-called, grades of milk. Variations of 200 to 300 per cent have been frequently reported in counts made in well controlled laboratories. While it is true that some technicians, under exceptionally well controlled conditions, have been able to obtain smaller percentage variations, it should not be forgotten that we must be governed by limitations arising through practical application where many samples of milk are daily prepared for counting by technicians who do not have time to exercise that extreme care which is possible only in research laboratories where there always exists ample time in which to plate the half dozen or so samples. The direct microscopic count of individual cells per cubic centimeter varied from 4.8 to 6.2 times greater than the count of the groups of bacteria under the microscope, and from 3 to 4.5 times greater than the agar plate colony count. The plate count on either plain or lactose agar was greater than the corresponding microscopic group counts. This relationship is to be expected in most samples due to the breaking up of the groups into smaller components during the process of plating the milk. The individual cell count averaged 4.5 times greater than the plain agar plate count but where lactose was added this difference was reduced to 3 times. There was likewise a reduction in the variability as well as a noticeable increase in the correlation. Undoubtedly the addition of lactose to the medium would materially increase the accuracy of the plating method. Although the microscopic group count was more variable than either the count of individual cells by the microscopic method or the agar plate count, yet the probable error of the coefficients of variability showed little or no significant difference. There was a better correlation between the counts of the individual bacterial cells and groups of cells per cubic centimeter by the direct microscopic method than between the individual cell count and the agar plate count. The poorest correlation obtained was that between the microscopic group counts and the agar plate counts. This is undoubtedly largely due to the variations introduced in the former count by the arbitrary manner in which separate groups are determined and by the counting of dead organisms and in the latter by variations in the adaptability of different organisms to growth in cultural media, and also by variations in the breaking up of colonies during plating. A few excessively high counts may have a marked effect upon the degree of correlation. For example, the elimination from 439 counts in all, of nine which were excessively high and which represented over-crowded microscopic fields and agar plates, resulted in a markedly lower correlation throughout (see table 7). It is conceivable that with raw milk uniformly low in bacterial content that there might be very little if any correlation between the two methods. Although, while showing more or less lack of correlation as applied to individual samples of milk, yet a close agreement may, in general, be expected in the results obtained by the two methods in rating the whole supply. Arithmetic averages of bacteria counts are apt to be misleading as a measure of the central tendency. One high count among several that were uniformly low would give an erroneous idea of the numerical bacterial quality of a given milk supply. The median, which is the middle number of a series of counts arranged in sequence, is a much fairer measure. One method gives as reliable a picture of the bacterial condition of any given raw milk supply as does the other. There is no reason for believing that one method is significantly more accurate than the other; nor that one lends itself more advantageously to the dividing of a raw milk supply into classes than the other. The plating method, however, is more applicable to the counting of bacteria in milk where there are only a few thousand present per cubic centimeter. Irrespective of the wide variations and possible sources of error that may be pointed out, both methods, if properly administered, are sufficiently accurate to insure a marked reduction in the amount of carelessly handled milk, without inflicting an undue hardship upon anyone.

Several procedures were developed and evaluated to find a satisfactory method for electronic counting of somatic cells in milk. Best results were obtained when milk was prepared for counting by a) diluting 1:50 with 0.85% saline, b) centrifuging 10ml of the diluted milk at a relative centrifugal force (RCF) of 1,020 for 15min at 4C, c) discarding the top 5ml of material (diluted milk and milk fat), and d) re-suspending the sedimented somatic cells by shaking in a vortex mixer. A Model B Coulter Counter fitted with a 100- μ -aperture tube was used to count the somatic cells in the diluted milk, with instrument threshold set to count all particles above 195 μ 3 in volume (approximate diameter 7.2 μ ). This setting was obtained by determining the best agreement of triplicate electronic counts with the direct microscopic method. Statistically designed experiments on 111 samples of milk counted in duplicate by both the electronic and microscopic methods showed that when milk samples had a somatic cell count of 300,000 per milliliter or over, counts by the two methods were not different at α = 0.05. The linear correlation index was 0.997. When milk samples contained less than 300,000 somatic cells per milliliter, however, the electronic method gave significantly higher counts than did the microscopic procedure.

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During recent years the methylene blue reductase test has been advocated as a practical method to use for determining the approximate number of bacteria in milk. The test has been used quite extensively in some European countries and recently to some extent in this country at dairy manufacturing plants and milk receiving stations. The well established plate count method and the more recent direct microscopic count method have been adopted as official by the American Public Health Association. These tests are more expensive and complicated and demand more care and skill than is required in operating the reductase test. But the latter test has not been used in this country to such an extent that its place in the dairy industry has been definitely established. During the months of July, August, and September, 1924, a number of dairy manufacturing plants and milk receiving stations were visited by the writer and a sample of the milk delivered by each producer was tested for bacteria by both the methylene blue reductase test and the direct microscopic method. The obiect of the work was first, to determine the grade (A, B or C) of each producer's milk on the day it was tested and, second, to learn how the results obtained by the reductase test checked with those obtained by the direct microscopic count method.

Summary A brief review of the theories concerned with the crystallization of lactose in ice cream has been presented, including a discussion of the properties of supersaturated lactose solutions, isomeric forms of lactose, and the rate of diffusion in ice cream at low temperatures. Four methods of detecting crystallized lactose in ice cream were studied. The microscopic method has been given in detail because of its employment in this research. The following factors and their influence on lactose crystallization in ice cream were discussed: Composition of the mix, freezer operation, rate of hardening, method of packaging, development of crystals throughout the mass of ice cream, temperature fluctuations, mechanical shocks, presence of seed or nuclei, mix pasteurization temperature, presence of nut meats and extent of fat globule clumping.

The catalase activity of milk was determined by a disc flotation method designed for measuring the activity of catalase preparations. The method consists of absorbing milk on a 0.5-in. disc of highly absorbent paper and dropping the disc in a 16 by 125 mm test tube containing 5 ml of 3% hydrogen peroxide. The time required for the disc to float to the surface is recorded. Statistical analysis of leucocyte counts made on milk by the direct microscopic method, and catalase activities by the disc flotation method gave a correlation coefficient of −0.75. Forty-six individual quarter samples of milk were examined to determine the relationships among leucocyte counts, disc flotation time, and California mastitis test reactions. The leucocyte counts and disc flotation times were in better agreement than leucocyte counts and California mastitis test reactions. The disc flotation test is simple, rapid, accurate, and requires very little equipment. It can be used to detect catalase activity in bulk milk, milk from individual cows, or from quarters of the udder.

The usefulness of the Wisconsin Mastitis Test in an abnormal milk program is dependent, in large measure, on the degree of its correlation with the direct microscopic leucocyte count. This correlation is dependent to a large degree on the accuracy with which the microscopic count is performed. One of the most likely sources of error in the microscopic count is the criteria for recognizing leucocytes in milk. Wisconsin test results were compared with the microscopic method using counting criteria: whole leucocytes only, whole leucocytes plus damaged cells at least 50% intact, and whole leucocytes and all cell fragments. The results of 117 milk sample comparisons indicated that microscopic Procedures 1, 2, and 3 showed a good correlation with the Wisconsin test (r 1 = 0.88, r 2 = 0.82, r 3 = 0.79), but Procedure 1 was best for predicting Wisconsin test values.