Cross-Reactivity
Definition
The capacity of an antibody raised against one analyte to bind structurally related compounds. In RIA, cross-reactivity is the main driver of false-positive screening results. For example, an antibody raised against morphine may also bind codeine or dihydromorphine at elevated concentrations. Confirmatory testing by chromatography-mass spectrometry is required to resolve cross-reactive positives.
- Field
- Immunoassay-based screening (RIA, ELISA)
- Main consequence
- False-positive screening results
- Common example
- Anti-morphine antibody binding codeine or dihydromorphine
- Resolution method
- Confirmatory chromatography-mass spectrometry
Common questions
Why can't cross-reactivity be eliminated entirely?+
Antibodies bind based on molecular shape, and structurally related compounds share enough of that shape to trigger a response. Raising an antibody with perfect specificity for one molecule and no others is not achievable with current immunoassay chemistry, which is why screening results are always treated as presumptive.
How do labs know when cross-reactivity has produced a false positive?+
They do not know from the screen alone. Any presumptive positive from an immunoassay is routed to a confirmatory method such as GC-MS or LC-MS, which separates and identifies compounds by mass rather than by antibody binding, resolving cross-reactive results.
Related terms
- ELISA (Enzyme-Linked Immunosorbent Assay)
- A plate-based immunoassay format in which a target antigen is captured by an antibody bound to a solid surface, detected by a...
- Lateral-Flow Immunoassay
- A point-of-care competitive immunochromatographic format on a nitrocellulose strip. In the absence of drug, colloidal gold-labelled antibody is captured at a test...
- Monoclonal Antibody
- An antibody produced from a single B-cell clone, recognising a single specific epitope. In forensic species identification, monoclonal antibodies offer higher specificity...
- Monoclonal Antibody (mAb)
- An antibody produced by a single hybridoma clone, so that every molecule in the preparation is identical, targets the same epitope, and...
- Precipitin Test
- A serological method in which an antiserum specific to one species' proteins is mixed with an extract of an unknown specimen. Formation...
- Prozone Effect
- A false-negative serological result caused by antibody excess. When antibody concentration is too high relative to antigen, antibodies saturate individual antigen molecules...
- Affinity
- The binding strength of a single antigen-binding site for one epitope, expressed as the equilibrium dissociation constant (Kd). A lower Kd indicates...
- Anti-Human Haemoglobin Antibody
- A monoclonal antibody raised against epitopes unique to human haemoglobin. Forms the detection and capture antibodies on the lateral-flow strip, conferring specificity...
- Antigen
- A molecule recognised by the adaptive immune system through specific binding to an antibody or T-cell receptor. Defined by immunogenicity (capacity to...
- Antiserum
- Serum collected from an immunised animal containing antibodies directed against a specific antigen or group of antigens. In species identification, antiserum raised...
- Calibration Curve
- In RIA, a set of standards containing known concentrations of unlabelled analyte that are run alongside test samples. Bound radioactivity is measured...
- CEDIA (Cloned Enzyme Donor Immunoassay)
- A homogeneous immunoassay using two inactive fragments of beta-galactosidase, an enzyme donor (ED) fragment conjugated to drug, and an enzyme acceptor (EA)...
Explained in these topics
- Antibody Structure, Classes, and SpecificityThe binding of an antibody to an antigen other than its primary target, usually because the two antigens share a structurally similar epitope. Cross-reactivity...
- Antigens: Structure, Types, and Forensic RelevanceThe ability of an antibody to bind to a molecule other than its target antigen because that molecule shares a structurally similar epitope. Cross-reactivity is...
- Complement Fixation and Mixed Antigen-Antibody ReactionsThe binding of an antibody to an antigen other than the one against which it was raised, caused by shared or structurally similar epitopes. In species identifi...
- ELISA: Principles, Formats, and Forensic ApplicationsThe binding of an antibody to an antigen other than its intended target, typically a structurally similar molecule. In forensic drug ELISA, cross-reactivity ca...
- Immunological Drug Testing: Screening and Forensic InterpretationThe binding of an immunoassay antibody to a compound other than the primary target analyte, because that compound shares a structurally similar epitope. Cross-...
- Immunological Confirmatory Tests for Human BloodThe ability of an antibody to bind an antigen other than its intended target. For human-blood lateral-flow tests, cross-reactivity with some non-human primate...
- Immunological Identification of Vaginal Secretions, Urine, and Other FluidsThe binding of an assay antibody to an antigen other than its intended target, producing a false-positive signal. In body-fluid immunoassays, cross-reactivity...
- Immunological Testing in Wildlife Crime and Food Fraud InvestigationsThe binding of an antibody to an antigen other than its primary target, typically because related species share conserved protein sequences. In species identif...
- Modern Immunological and Molecular Approaches to Species IdentificationThe binding of an antibody to an epitope on a protein from a non-target species that shares structural similarity with the target epitope. Cross-reactivity bet...
- The Precipitin Test for Species IdentificationThe property by which an antiserum raised against one species' proteins also reacts with structurally similar proteins from a phylogenetically related species....
- Radioimmunoassay: Principles, Uses, and Decline
- RSID-Blood and ABAcard HemaTraceA positive test result produced by a non-target species whose haemoglobin shares enough structural similarity to bind the anti-human antibody. For HemaTrace, d...
- Sensitivity, Specificity, and Quality Controls in Forensic ImmunoassaysThe binding of an antibody to an antigen other than its primary target because the two antigens share a structurally similar epitope. Cross-reactivity is the p...
- Sources of Error and False Results in Serological TestingThe binding of an antibody to an epitope on a molecule other than its intended target, due to structural similarity. In forensic serology, cross-reactivity can...