THE INVESTIGATION OF DANDRUFF 619 would be to develop a standardized method of taking samples, followed by a viable count procedure. Both steps involve serious practical difficulties. The area liable to show dandruff is considerable and the distribution of scaling is uneven, whilst the presence of hair interferes with the sampling. Furthermore, owing to entrainment in the horny scale and fatty matter, and the clumping tendency of the organisms, a true viable count is virtually out of the question. We therefore attempted to devise a reproducible, "semi-quantitative" assessment of the level of infection. A sterile plastic applicator (Fig. 7) was used to brush the scalp of each subject six times in a standardized manner. Samples of scale adhering to the teeth of the applicator were then trans[erred with light pressure to the culture medium in a 35 in petri dish. Two plates, representing two separate applicators, were obtained from each subject. The inoculated plates were then placed in a plastic box with a close-.fitting lid to prevent excessive drying of the culture medium, and incubated at 37øC for 5-7 days. Figure 8 Typical incubated plate showing units of growth of P. ovale.

620 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS The units of growth were then recorded. Each applicator had 160 teeth, so the maximum number of units (colonies) for 2 plates was 320 (Fig. 8). Colony characteristics were examined by means of a binocular microscope (x 10) and the microscopic appearance of cells was also noted. This technique was not truly quantitative, since numerous organisms on one tooth of an applicator would be classified as a single unit of growth further, we could not be sure that "no growth" always meant "no infection". Nevertheless, the method did appear to provide an approximate repre- sentation of the level of infection. P.ovale in a Clinic Panel In the 18-week period immediately preceding the test, the 19 volunteer subjects had their hair washed in the salon weekly with our standard anionic shampoo. During the treatment period, 5% cetrimide solution was used as a shampoo to avoid interaction of materials on the scalp with the anti- septic selected. Daily treatment, in most cases by the subjects themselves, was carried out with a 0.25% aqueous solution of dequalinium chloride, approximately 5 ml per application. It had previously been shown in vitro that P.ovale was inhibited by dequalinium chloride at 1: 1000. Table II Average count (units of growth) of P. ovale during dequalinium treatment Average Count Pre-Treatment Treatment Subject March-June July-October November 1962- May 1963- No. 1962 1962 April 1963 October 1963 1 238 165 26 108 5 105 210 34 51 6 182 35 8 21 10 120 248 46 76 11 55 13 1 1 14 29 25 0 32 17 39 164 12 8 18 30 69 10 14 During the pre-treatment period, the average recovery of P.ovale units of growth varied between 30 and 240 per subiect, out of 320 units possible. In the first 3 months of treatment, the P.ovale recovery decreased in some cases and increased in others. However, all cases showed a decrease in average P.ovale units during the next 6 months (November 1962 to April 1963), though some of these counts rose again to some extent whilst treatment continued (May to October 1963) data from subjects continuing the full period of investigation are given in Table II.