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We describe the use of inorganic (metal) colloidal particles as a label for immunoassays. Dose-response curves for human placental lactogen (HPL) and human chorionic gonadotrophin (HCG) were obtained with sandwich immunoassays, using conjugates consisting of antibody-coated colloidal gold or silver particles. Several techniques were used to measure the amount of bound conjugate, viz. colorimetry and carbon rod atomic absorption spectrophotometry (CRAAS). At higher antigen concentrations the results of the assay could be read by the naked eye. Using gold particles as label and CRAAS as detection method, we found a detection limit for a sandwich HPL sol particle immunoassay (SPIA) of 1,4 pmol/l, which was equal to that of an optimalized competitive radioimmunoassay. When using a colorimeter the detection limit for HPL of this SPIA was 5,4 pmol/l, which was superior to that of a corresponding sandwich enzyme-immunoassay (EIA). HPL and HCG were also simultaneously determined, using microtitration plates, coated with a mixture of anti-HPL and anti-HCG, and a mixture of silver particle anti-HPL conjugate and gold particle anti-HCG conjugate. CRAAS was used to measure the bound amount of silver and gold conjugate. This simultaneous assay requires more work in order to obtain better sensitivities.

(World Health Organization, 1975). For a number of years our laboratory has worked on the development of a new and sensitive immunochemical technique, the enzymeimmunoassay (EIA) Schuurs, 1971, 1972;van Weemen et al, 1974). This method has also been studied by several other groups of investigators, particularly by Perlmann (1971, 1972) and . The present paper describes the application of a solid-phase enzyme-immunoassay for the detection of IHBsAg. injections of purified HBsAg in complete Freund's adjuvant at two-week intervals until a reciprocal titre of antibody to HBsAg (anti-HBs) of > 512 in a standard immunodiffusion set-up was attained with both subtype ay and ad. The resulting antiserum was tested by immunodiffusion against normal human serum (undiluted and diluted from 1:2 to 1:32). The antiserum was absorbed with insolubilized normal human serum proteins until no antibodies to human serum proteins could be detected. The gammaglobulin fraction was precipitated with 14% (w/v) Na2SO4, and the precipitate was dissolved in 0-0175 M phosphate buffer pH 7-6 and dialysed against the same buffer. ANTIBODY-COATED MICROTITRE PLATESThe wells of Microtiter's plates (Cooke) were filled with 0-1 ml sheep anti-HBs gammaglobulin fraction having a protein concentration of 0 03 mg/ml. The liquid was removed from the plates after overnight incubation at 4°C, the plates were washed four times with 0'04 M TRIS-buffer pH 7 4, dried over silicagel, and stored at 4VC in the presence of silicagel.ANTIBODY-ENZYME CONJUGATE Horse-radish peroxidase (Miles; RZ 3-0) was linked to sheep anti-HB8 gammaglobulin in a protein ratio of 4:1 by the 'two-step' method of Avrameas 873 on 12 May 2018 by guest. Protected by copyright.