Non-denaturing Lysis Reagent for Immunoassay Interference

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Solution Overview

Problem

Current immunoassays for immunosuppressant drugs like tacrolimus and cyclosporine face interference from human anti-animal antibodies, requiring separation steps that complicate the measurement of these drugs in blood samples, especially when using antibodies from different species.

Innovation Solution

A non-denaturing lysis reagent containing ethylene glycol or propylene glycol, along with mouse immunoglobulins, is used to create a homogeneous mixture that allows direct binding of capture antibodies to analytes without pre-existing interference, eliminating the need for separation steps and reducing human anti-mouse antibody interference.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Measurement precision

If separation steps are used to remove binding proteins and interfering antibodies, then measurement accuracy is improved, but device complexity and loss of time increase

Engineering Contradiction:

Improvemeasurement accuracyVSAvoidassay complexity

Core Design Contradiction:

Measurement precisionVSDevice complexity

Solution Approach 1:

The patent combines the lysis function and the interference neutralization function into a single reagent formulation. The lysis reagent contains both the lysis components (detergent, salt) and excess immunoglobulins of the same animal species as the capture antibody, eliminating the need for separate separation steps while maintaining measurement accuracy.

Inventive Principle:

Principle #5Merging (Combining)

Solution Approach 2:

The lysis reagent is prepared in advance with excess immunoglobulins already included at concentrations sufficient to neutralize interfering antibodies. This preliminary preparation ensures that when the reagent is added to the sample, both cell lysis and interference neutralization occur simultaneously before the assay proceeds, without requiring additional processing steps.

Inventive Principle:

Principle #10Preliminary action

2Measurement precision

If separation steps are used to remove interfering proteins, then measurement accuracy is improved, but loss of time increases

Engineering Contradiction:

Improvemeasurement accuracyVSAvoidassay time

Core Design Contradiction:

Measurement precisionVSLoss of time

Solution Approach 1:

Inventive Principle:

Principle #5Merging (Combining)

Solution Approach 2:

The lysis reagent performs multiple functions continuously in a single step: it lyses cells to release bound analyte, solubilizes proteins, and neutralizes interfering antibodies through competitive binding. This continuous multi-functionality eliminates the need for sequential processing steps, reducing total assay time while maintaining measurement accuracy.

Inventive Principle:

Principle #20Continuity of useful action

3Quantity of substance

If detergents are used to lyse cells, then analyte release is improved, but interference from denaturation increases

Engineering Contradiction:

Improveanalyte releaseVSAvoiddenaturation interference

Core Design Contradiction:

Quantity of substanceVSObject-generated harmful factors

Solution Approach 1:

The patent modifies the lysis reagent composition by including excess immunoglobulins of the same animal species as the capture antibody at concentrations of 1-100 mg/mL. This parameter change enables the reagent to neutralize interfering antibodies through competitive binding while maintaining the lytic function, thereby releasing analyte without the harmful effects of protein denaturation.

Inventive Principle:

Principle #35Parameter changes

Solution Approach 2:

The patent converts the potentially harmful effect of antibody interference into a beneficial competitive binding scenario. By adding excess immunoglobulins that compete with interfering antibodies for binding to the capture antibody, the reagent neutralizes the harmful interference while maintaining the desired analyte detection function.

Inventive Principle:

Principle #22Blessing in disguise (Convert harm into benefit)

Applied Scientific Principles

This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.

Function Achieved in This Case

This method enables accurate and efficient measurement of immunosuppressant drugs by forming a solid phase-affixed immune complex, reducing interference and simplifying the assay process, allowing for precise quantitation without centrifugation or detergent use.

Implementation Method 1

contacting a test sample comprising cells with a lysis reagent to form a homogeneous lysis mixture

Methodology Applied

Scientific EffectCell lysis:

Implementation Method 2

the lysis reagent comprising a glycol selected from the group consisting of ethylene glycol, propylene glycol, and a glycol having from two to six carbon atoms

Methodology Applied

Scientific EffectGlycol solution effect:

Implementation Method 3

the immunoglobulins compete with the capture antibody for binding to the solid phase-affixed binding partner

Methodology Applied

Scientific EffectAntibody-antigen binding:

Implementation Method 4

the immunoglobulins compete with the capture antibody for binding to the solid phase-affixed binding partner, whereby the capture antibody and analyte form a solid phase-affixed immune complex

Methodology Applied

Scientific EffectCompetitive inhibition:

Implementation Method 5

the capture antibody and analyte form a solid phase-affixed immune complex

Methodology Applied

Scientific EffectImmune complex formation:

Implementation Method 6

contacting the lysis mixture with a solid phase comprising a solid phase-affixed binding partner for the capture antibody and with the capture antibody, wherein the capture antibody is not pre-bound to the binding partner, and said contacting is carried out under conditions suitable for the capture antibody to bind to the analyte

Methodology Applied

Scientific EffectAntibody-specific binding:

Data Source

PatentEP2118657B1Non-denaturing lysis reagent for use with capture-in-solution immunoassay

Publication Date: 2014.05.21 ABBOTT LAB INC

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AI summary

The invention provides a lysis reagent and method for preparing a test sample for use in an assay, wherein the method yields a homogeneous lysis mixture suitable for use in automated pipetting systems without the need for a centrifugation step. The lysis reagent includes a glycol and non-specific animal immunoglobulins. Other aspects of the invention include related immunoassays and test kits.