New quantitative method to determine protein concentration and cell number in aqueous in vivo

We have developed a new quantitative method to determine protein concentration and number of cells in the aqueous in vivo. The principal instruments in the system were a He-Ne laser and a detection system for measuring scattered light intensity. The power of the He-Ne laser was 25 microW and the focused beam diameter was 20 microns. The laser beam was operated by an optical scanner. The sampling window, 0.3 X 0.5 mm, was fixed in the center of the laser path. For the protein concentration measurement, the laser beam was scanned for a length of 0.6 mm vertically, covering the sampling window. To minimize signal contamination by background laser scattering, scattered light intensity produced by aqueous protein (Sp) was calculated as follows. Sp = Si - (S'o + S''o)/2, where Si indicates scattered light intensity when the beam passed in the sampling window and S'o and S''o indicate scattered light intensity when the beam passed above and below the sampling window. For the cell counts, the laser beam (0.25 X 0.6 mm) was scanned over the sampling window and the detected peaks were counted. Each measurement mode took 0.5 seconds. The operation of the instrument and data analysis were performed by a personal computer. In in vitro measurements with human plasma solution in diluting factors ranging from 1/50 to 1/10(4), and bovine serum albumin solution in concentrations ranging from 1 g/100 ml to 1 mg/100 ml, significant linear correlations between the values for concentration and photon counts (/msec) were obtained in each solution (r, ranging from 0.987 to 0.998, P = 0.000). In in vitro experiments with latex particles with diameters of 2.02 or 2.95 microns, significant correlations between the number of detected peaks and latex particles were also obtained. However, as the number of latex particles decreased, the variation in the number of detected peaks was large. In 31 normal young adults, with an average age of 23 years, the average photon count was 4.1 +/- 1.0, which corresponded to 27 mg/100 ml according to the in vitro bovine albumin solution measurement, and there was no definite relation in photon counts between eyes with and without mydriatics. Twenty patients with incipient or immature cataract, whose ages averager 70 years, showed significantly higher average photon counts, 6.2 +/-2.5, than the above young adults.We concluded that this new method of determining protein concentration and cell number in the aqueous enabled us to make a noninvasive and quantitative evaluation of the inflammation in the anterior segment of the eye.(ABSTRACT TRUNCATED AT 400 WORDS)