Determination of ambient concentrations of several analytes

A method for determining the fractional binding site occupancy of a plurality of binding agents by a plurality of analytes in a liquid sample of V liters, comprising:

• (a) loading a plurality of different binding agents, each being capable of reversibly binding an analyte which is or may be present in the liquid sample and is specific for said analyte as compared to the other components of the liquid sample, onto a support at a plurality of spaced apart small spots such that each spot has a high coating density of one of said binding agents but not more than 0.1 V/K moles of binding agent are present on any one spot, where K liters/mole is the affinity constant of said binding agent for said analyte;
• (b) contacting the loaded support with the liquid sample to be analyzed, such that each of the spots is contacted in the same step with said liquid sample, the amount of liquid used in said sample being such that only an insignificant proportion of any analyte present in said liquid sample becomes bound to said binding agent specific for said analyte;
• and
• (c) thereafter contacting the loaded support with site-recognition reagents which recognize either the unfilled binding sites or filled binding sites of that binding agent, the site-recognition reagents being labelled with markers from which the fractional binding site occupancy for each binding agent is determined.

A method for detecting a plurality of analytes in a liquid sample of volume V liters, comprising:

• loading a plurality of different binding agents, each being capable of reversibly binding an analyte which is or may be present in the liquid sample and is specific for said analyte as compared to the other components of the liquid sample, onto a support means at a plurality of spaced apart small spots such that each spot has a high coating density of one of said binding agents but not more than 0.1 V/K moles of binding agent are present on any spot, where K liters/moles is the affinity constant of said binding agent for said analyte;
• contacting the loaded support means with the liquid sample to be analyzed, such that each of the spots is contacted in the same step with said liquid sample, the amount of liquid used in said sample being such that only an insignificant proportion of any analyte present in said liquid sample becomes bound to said binding agent specific for said analyte;
• contacting the support with a site-recognition reagent specific for each binding agent in a competitive or non-competitive technique, the site-recognition reagent being capable of recognizing either the unfilled binding sites or the filled binding sites on said binding agent, said site-recognition reagent being labelled with a marker;
• and
• measuring the signal from the marker of the site-recognition reagent in a particular location to detect the presence of said plurality of analytes in said sample.