Solid-phase activity assay for biologically active substance
OwnerRonald R. Marquardt, Guojie Wang +3inventors
InventorsRONALD R. MARQUARDT, GUOJIE WANG, ZHIQUN ZHANG, ZHIBO GAN, XIAO HAO
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Abstract
A method of detecting, via a solidphase assay the amount of biological activity, identity and/or the quantity of a biologically active substance is disclosed. The method utilizes the biological activity of the substance itself to provide the method of detection. The method provides competitive and noncompetitive assays.
Claims55
A solid-phase assay for detecting the amount of biological activity of a biologically active substance utilizing the biological activity comprising the steps of:
- a. binding to a surface a first component wherein the first component is conjugated to a first indicator;
- b. contacting a sample containing a second component having unknown biological activity to the conjugated first component in a reaction mixture under conditions such that the biological activity between the first and second component will unbind the first component;
- c. removing the sample after a fixed reaction time;
- and
- d. determining the amount of remaining bound first component wherein there is a reciprocal relationship between the amount of biological activity and the remaining bound first component.
A method for detecting via a solid-phase assay the amount of an inhibitor of biological activity of a biologically active substance utilizing the inhibition of biological activity comprising the steps of:
- a. binding to a surface a first component wherein the first component is conjugated to a first indicator;
- b. contacting a sample containing a known amount of a second component having known biological activity and an unknown amount of a third component being an inhibitor of the second component to the first component in a reaction mixture under conditions such that the biological activity between the first and second component will unbind the first component and the third component will interfere with the reaction between the first and second component;
- c. removing the sample;
- and
- d. determining the amount of remaining bound first component wherein there is a direct relationship between the remaining bound conjugated first component and amount of the third component in the sample.
A method of detecting via a solid-phase assay the identity of a biologically active substance utilizing inhibition of the biological activity comprising the steps of:
- a. binding to a surface a first component wherein the first component is conjugated to a first indicator;
- b. contacting a sample containing a second component having generally known biological activity but unknown specificity and containing a known amount of a third component being a potential inhibitor of the second component to the first component in a reaction mixture under conditions such that the biological activity between the first and second components will unbind the first component and the third component can interfere with the reaction between the first and second component;
- c. removing the sample after a predetermined time;
- and
- d. determining the amount of remaining bound first component wherein if there is a reduction of the amount of the bound first component, then the third component did interfere with the reaction between the first and second component thereby identifying the second component.
A method of detecting via a solid-phase competitive assay the quantity of a biologically active substance utilizing the biological activity of the substance comprising the steps of:
- a. binding to a surface a known quantity of a first component which binds to a biologically active second component;
- b. contacting a sample containing an unknown quantity of the second component having biological activity and a known quantity of the second component coupled to a first indicator to the first component in a reaction mixture under conditions such that the first and second components will bind to each other due to the biologic activity;
- c. removing the sample;
- and
- d. determining the amount of the second component coupled to a first indicator bound to the first component wherein a reciprocal relationship exists between the quantity of the second component coupled to a first indicator bound to the first component and the unknown quantity of the second component having biological activity in the sample.
A method of detecting via a solid-phase competitive assay the quantity of a substance binding to a biologically active substance utilizing the biological activity of the substance comprising the steps of:
- a. binding to a surface a known quantity of a first component which binds to a second component having biological activity;
- b. contacting a sample containing an unknown quantity of the first component and a known quantity of the second component coupled to a first indicator to the bound first component in a reaction mixture under conditions such that the first and second components will bind due to biologic activity;
- c. removing the sample;
- and
- d. determining the amount of second component coupled to a first indicator bound to the bound first component wherein a reciprocal relationship exists between the quantity of the second component coupled to a first indicator bound to the bound first component and the unknown quantity of the first component in the sample.
A method of detecting via a solid-phase competitive assay the quantity of a biologically active substance utilizing the biological activity of the substance comprising the steps of:
- a. binding to a surface a known quantity of a second component having biological activity;
- b. contacting a sample containing an unknown quantity of the second component and a known quantity of a first component coupled to a first indicator to the bound second component in a reaction mixture under conditions such that the first and second components will bind due to biologic activity;
- c. removing the sample;
- and
- d. determining the amount of first component coupled to a first indicator bound to the bound second component wherein a reciprocal relationship exists between the quantity of the bound first component coupled to an indicator and the unknown quantity of the second component in the sample.
A method of detecting via a solid-phase competitive assay the quantity of a substance binding to a biologically active substance utilizing the biological activity of the substance comprising the steps of:
- a. binding to a surface a known quantity of a second component having biological activity which binds to a first component;
- b. contacting a sample containing a known quantity of the first component coupled to a first indicator and an unknown quantity of a first component to the second component in a reaction mixture under conditions such that the first and second components will bind due to biologic activity;
- c. removing the sample;
- and
- d. determining the amount of first component coupled to a first indicator bound to the bound second component wherein a reciprocal relationship exists between the quantity of the bound first component coupled to an indicator and the unknown quantity of the first component in the sample.
Dependencies are inferred from the claim text.