78 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS incorporated into the hair shaft during the growth phase of the hair cycle, it should be possible to utilize fluorescein as a marker for the growing hair. This report examines the usefulness of fluorescent tracers as an approach to accurately determine the rate of hair growth. MATERIALS AND METHODS Adult male Hartley guinea pigs (250-300 g) were purchased from Camm Laboratory Animals, Inc. (Wayne, NJ). Male Syrian golden hamsters (10-12 weeks) were ob- tained from Harlan-Sprague Dawley (Farmersberg, VA). The animals were maintained at a photoperiod of 12-h light and 12-h dark. Food and water were provided ad libitum. An area 3 to 4 cm on the back of the animals was plucked manually. Three days after epilation, various doses (3 to 60 mg/kg body weight) of sodium fluorescein (Aldrich Chemical Co., Milwaukee, WI), dissolved in 0.2 cc of physiological saline, were in- jected subcutaneously on alternate days at a site distant from the epilated area. The hairs were plucked three days after the last injection, rinsed with hexane, and dry- mounted on glass slides. The incorporation of this dye into the hairs of hamsters was also evaluated using subcutaneous injections of sodium fluorescein. In another experi- ment, various doses (0.05 to 1.0 mg) of sodium fluorescein in 0.1 cc of saline were injected intradermally into epilated dorsal skin of the guinea pig. The hair of the guinea pig was classified according to the method of Dawson (6). The fluorescent bands in I and type II hair were visualized using a fluorescence microscope (Ortholux II, Leitz) equipped with a drawing attachment and integrated with a com- puterized graphics calculator (Numonics Corp., Lansdale, PA). Type I hairs are long (10-25 mm in length) and broad, with the apex ending abruptly in a sharp point. Type II hairs are shorter than type I and characterized by a very fine, wool-like apical portion. Linear growth was determined by measuring the distances between the center of two fluorescent bands. RESULTS Figures la and lb show the presence of fluorescent bands along the shafts of type I and II hairs of the guinea pig when examined under the fluorescence microscope. Although autofluorescence was present along the entire length of the shaft, the amount of fluores- cein incorporated after subcutaneous administration at a dose of 30 mg/kg was sufficient to provide a distinct fluorescent band. Some of the nonspecific fluorescence, possibly due to lipids coating the hair shaft, can be eliminated with chloroform or ether. The effects of various doses of subcutaneously injected sodium fluorescein on the inten- sity of the band in guinea pig hair are shown in Table I. Doses of 15 mg/kg and below produced barely detectable banding or no fluorescence at all. The animals tolerated the treatments well and untoward effects were not observed even at the highest dose. So- dium fluorescein was also administered intradermally at doses ranging from 0.05 mg to 1.0 mg/injection. Fluorescent bands were present even at the lowest dose (0.05 mg). However, a greater fluorescence intensity, suitable for growth measurements, was ob- served at a dose of 0.25 mg and higher.

RATE OF HAIR GROWTH VIA FLUORESCENCE 79 Cl b Figure 1. The fluorescent bands in the hair of the guinea pig after subcutaneous injection of sodium fluorescein at a dose of 30 mg/kg of body weight (25 x magnification). (a) Type I hair. (b) Type II hair. Table I The Intensity of Fluorescent Band in Type I and II Hair of Guinea Pigs Injected With Sodium Fluorescein Method of Fluorescence b administration Dose a intensity Subcutaneous 0 - injection 3 - 5 - 15 30 + + 60 ++ + Intradermal 0.0 - injection 0.05 + 0.10 + 0.25 + + + 0.5 +++ 1.0 +++ Each point represents observations derived from three animals. a Dose for subcutaneous injections is expressed in mg of fluorescein per kg of body weight. Dose for the intradermal injection is in mg of fluorescein per injection. b The degree of fluorescence was measured subjectively using a grading system: (-) no fluorescence, (+) barely detectable, (+ + ) detectable, (+ + + ) intense fluorescence. Animals within each group responded similarly. Fluorescence intensity was similar for both type I and type II hairs