Abstract
A new immunological method of quantitative measurement of antigens and antibodies was developed by applying the near infrared turbidimetry to the latex agglutination reaction. The turbidity of the antibody-sensitized latex (0.1 to 0.8 μm in the diameter) increases very markedly along with the immunoreaction with the corresponding antigens in the near infrared region, not in the visible or ultraviolet region. This phenomenon is supported theoretically by the Mie scattering theory which handles the light scattering by relatively large particles.
Detailed studies by the rate method in the system were done with hCG (human chorionic gonadotropin), and the system was evaluated from the view points of reproducibility, recovery and correlation with conventional RIA method. The characteristics of the system were exhibited as follows:
(1) Both within and between assay variance (C. V.) of the results obtained through the reproducibility experiment of the rate measurement were less than 5% with triplicate specimens in each of three-time experiments.
(2) Data processing was carried out according to the second order equation, log V=a+b (log A)+c (log A)2, where V represents the rate of turbidity increase at the early phase of immunoreaction, and A represents the concentration of the antigen. The calibration curve obtained with hCG in saline solution containing 0.2% of BSA showed very narrow width of confidence band.
(3) Interference by turbid serum specimens was negligible. Variance (C. V.) of the results obtained through the reproducibility experiment with clinical specimens was as small as those obtained with the standard samples dissolved in saline.
(4) The recoveries of added hCG in sera and urine were 94-106% and 94-117%, respectively, except for two cases of the lowest concentration of hCG dissolved in urine.
(5) Correlation coefficient between this technics (LPIA) and the conventional RIA was more than 0.97.
(6) The minimum detectable dose of hCG is 2 ng (30 mIU) per ml. with the measurable range up to about 200 ng (31U) per ml., when 100 μl sample is used.
(7) Measurement time required for hCG is in the range of 40 to 120 seconds.
In summary, this study has shown that the LPIA system is a promising approach to detect the minute amount of antigen by simple procedure. An overall agreeement in terms of correlation of the LPIA system with RIA procedure has been obtained with an advantage of negligible interference by any color and turbidity of clinical samples. Further works are requied to expand the system for many other kinds of antigens and to provide the computer-based LPIA with fully automated instrument hardware.
