What is the difference between immunoassay (IA) and immunochemistry (IC) in clinical practice, and when should each be used for diagnosing and monitoring diseases?

Immunoassay vs Immunochemistry: Key Distinctions and Clinical Applications

Immunoassay (IA) and immunochemistry (IC) are overlapping but distinct concepts: immunoassay refers specifically to quantitative or semi-quantitative laboratory techniques that measure analyte concentrations in biological fluids, while immunochemistry encompasses the broader field of antibody-antigen interactions including tissue-based visualization techniques like immunohistochemistry (IHC) and immunofluorescence (IF).

Core Definitions and Technical Distinctions

Immunoassay (IA)

• Primarily used for quantitative measurement of proteins, hormones, drugs, antibodies, and other analytes in serum, plasma, or other body fluids 1
• Detection methods include: radioimmunoassay (RIA), enzyme immunoassay (EIA/ELISA), chemiluminescence immunoassay (CLIA), electrochemiluminescence immunoassay (ECLIA), and fluorescent immunoassay 2, 3
• Key characteristic: Produces numerical values with defined sensitivity and specificity for analyte concentration 2
• Examples in practice: Measuring testosterone levels (LC-MS/MS or immunoassay after extraction preferred over direct immunoassay for higher accuracy), detecting anti-HCV antibodies (3rd generation EIA with 97.2-99% sensitivity), or quantifying IgG levels in autoimmune hepatitis 2

Immunochemistry (IC)

• Primarily used for localization and visualization of antigens in cells and tissues 4, 5
• Main techniques: Immunohistochemistry (IHC) on tissue sections and immunofluorescence (IF) on cells or frozen sections 4
• Key characteristic: Provides spatial information about antigen distribution and cellular morphology, typically qualitative or semi-quantitative 5
• Examples in practice: Determining NSCLC subtype using TTF1 and p40 markers, identifying tumor origin in metastatic cancer using tissue-specific markers, or detecting immune cell populations in tissue specimens 2

Clinical Decision Algorithm: When to Use Each Method

Use Immunoassay When:

1. Quantitative measurement is required for diagnosis or monitoring

   • Measuring specific antibody titers to protein/polysaccharide antigens in primary immunodeficiency evaluation 2
   • Monitoring IgG levels to assess treatment response in autoimmune hepatitis (normalization correlates with remission) 2
   • Quantifying testosterone for PCOS diagnosis (LC-MS/MS shows superior performance with pooled sensitivity 0.74, specificity 0.86) 2

2. Screening large populations or confirming serological status

   • Anti-HCV screening using EIA/CLIA (sensitivity 97.2-99%, specificity 99.8-100%) 2
   • HCV RNA quantification using real-time PCR (detection limit 12-15 IU/mL) for treatment monitoring 2

3. Serial monitoring of disease activity or treatment response

   • Immune response monitoring in cancer immunotherapy trials (ELISPOT for functional antigen-specific cells) 2

Use Immunochemistry (IHC/IF) When:

1. Tissue diagnosis requires morphological context

   • NSCLC subtyping when morphology alone is insufficient: Use limited IHC panel (TTF1 for adenocarcinoma, p40 for squamous cell carcinoma) to reduce NSCLC-NOS rate to <10% 2
   • Identifying primary site in metastatic cancer of unknown origin using tissue-specific marker panels 2

2. Predictive or prognostic biomarker assessment in tissue

   • HER2 testing in breast cancer for treatment selection 5
   • Surrogate markers for molecular alterations (IDH1, ATRX in brain tumors) 5

3. Detecting rare cells or spatial distribution patterns

   • Identifying tiny foci of tumor cells inconspicuous on H&E staining 5
   • Evaluating immune cell infiltration patterns in tissue specimens 2

Critical Technical Considerations

For Immunoassays:

• Method selection impacts accuracy: LC-MS/MS and immunoassays after extraction demonstrate superior performance over direct immunoassay for testosterone measurement 2
• Standardization is essential: Central laboratories recommended for multi-center trials to minimize variation 2
• False positives occur: Anti-HCV false positives can occur in autoimmune diseases; confirm with HCV RNA testing 2
• Timing matters: Average seroconversion time for anti-HCV is 8-9 weeks; early acute infection may show negative antibody with positive RNA 2

For Immunohistochemistry:

• Tissue conservation is paramount: Use only two tissue sections for NSCLC subtyping; avoid excessive IHC investigation 2
• Markers are not uniformly specific or sensitive: Avoid large series of immunohistochemistry markers; communication with pathologist is essential 2
• Proper fixation required: FFPE tissue processing and antigen retrieval methods are critical for reliable results 5
• Indirect immunofluorescence preferred for autoantibody detection in autoimmune hepatitis (using freshly frozen rodent substrate including kidney, liver, stomach) 2

Common Pitfalls and How to Avoid Them

Immunoassay Pitfalls:

• Using direct immunoassay for testosterone: Results in lower diagnostic accuracy; use LC-MS/MS or immunoassay after extraction instead 2
• Interpreting isolated antibody positivity: Anti-HCV persists after recovery; differentiate current from past infection using HCV RNA 2
• Ignoring immunosuppression effects: Severely immunosuppressed patients (hemodialysis, HIV, transplant recipients) may have negative anti-HCV despite active infection; test HCV RNA directly 2

Immunochemistry Pitfalls:

• Over-ordering IHC panels: This wastes precious tissue; select markers based on clinical context and morphology 2
• Misinterpreting NSCLC-NOS: If both TTF1 and p40 are negative, diagnosis remains NSCLC-NOS; do not continue adding markers indiscriminately 2
• Using wrong substrate for autoantibody detection: For autoimmune hepatitis, IFL on rodent tissue is superior to ELISA and remains the gold standard 2

Complementary Use in Clinical Practice

Both methods often work synergistically: In primary immunodeficiency evaluation, immunoassays measure serum immunoglobulin levels and specific antibody responses, while immunochemistry techniques (flow cytometry with fluorescent antibodies) enumerate lymphocyte subsets 2. In cancer diagnosis, IHC determines tissue origin and subtype, while immunoassays monitor tumor markers and treatment response 2.