What are the implications and management of false positives in Serum Protein Electrophoresis (SPEP)?

False Positives in Serum Protein Electrophoresis (SPEP)

False-positive M-spikes on SPEP most commonly result from fibrinogen contamination in incompletely clotted samples, recent intravenous immunoglobulin (IVIG) administration, or polyclonal hypergammaglobulinemia mimicking monoclonal patterns.

Primary Causes of False-Positive SPEP Results

Fibrinogen Contamination

• Fibrinogen from incompletely clotted serum samples creates a spike in the beta-gamma region that mimics a true monoclonal protein 1
• This represents the most common technical cause of false-positive SPEP results in clinical practice 1
• The γ/IgG ratio can distinguish fibrinogen spikes from true monoclonal proteins: a ratio >1.13 suggests fibrinogen contamination rather than true monoclonal gammopathy 1
• Using a γ/IgG ratio cut-off of 1.0 provides 93% sensitivity and 91% specificity for identifying fibrinogen interference when elevated IgA or IgM cases are excluded 1

IVIG Administration

• IVIG therapy causes false M-spikes in approximately 1.4% of treated patients tested by SPEP 2
• The interference is most pronounced when SPEP is performed within 30 days of IVIG administration 2
• Of patients receiving IVIG who had an apparent M-spike on SPEP, 5 of 14 (36%) tested within 30 days had false positives not confirmed by subsequent immunofixation electrophoresis (IFE) or clinical follow-up 2
• The exogenous polyclonal immunoglobulins in IVIG can create a broad-based peak that may be misinterpreted as a monoclonal spike 2

Technical and Analytical Factors

• Background gamma globulin concentration significantly affects SPEP accuracy, with hypergammaglobulinemia leading to overestimation of M-protein quantitation 3
• Central-gamma migrating M-proteins have greater imprecision and lower accuracy compared to cathodal migrating proteins due to increased gamma background contribution 3
• At M-protein concentrations below 2 g/L, inter-laboratory variability increases substantially, with coefficient of variation exceeding acceptable limits 3
• The perpendicular drop (PD) method of gating M-proteins shows more overestimation than tangent skimming (TS), particularly in hypergamma samples 3

Clinical Context: SPEP Performance in Lytic Bone Lesions

• In patients presenting with radiolucent bone lesions, SPEP has only 47% positive predictive value for plasma cell neoplasms, meaning more than half of positive results do not represent true myeloma 4
• The specificity of SPEP in this clinical scenario is 83%, indicating 17% false-positive rate 4
• However, the negative predictive value is 94%, making SPEP more useful for ruling out rather than ruling in plasma cell disorders 4
• Even in patients with multiple lytic lesions (where myeloma prevalence is 44.7%), SPEP maintains only 40% positive predictive value 4

Management of Suspected False-Positive Results

Immediate Confirmatory Testing

• Never diagnose monoclonal gammopathy based on SPEP alone—always confirm with serum immunofixation electrophoresis (SIFE) or immunosubtraction 4, 2
• Obtain serum free light chain assay in conjunction with SPEP to improve diagnostic accuracy 4
• Consider urine protein electrophoresis with immunofixation, as some monoclonal proteins are exclusively or predominantly excreted in urine 4

Clinical History Integration

• Document recent IVIG administration (within 30 days) before interpreting SPEP results 2
• If IVIG was given recently and SPEP shows an M-spike, repeat testing 4-6 weeks after last IVIG dose to allow clearance 2
• Assess for conditions causing polyclonal hypergammaglobulinemia (chronic infections, autoimmune diseases, liver disease) that may create pseudo-monoclonal patterns 5

Sample Quality Assessment

• Calculate the γ/IgG ratio when a beta-gamma spike is present: ratio >1.13 strongly suggests fibrinogen contamination rather than true monoclonal protein 1
• If fibrinogen contamination is suspected, recollect sample ensuring complete clotting before centrifugation 1
• Verify total protein measurement using reverse biuret method to ensure accurate baseline for electrophoresis interpretation 3

Common Pitfalls and How to Avoid Them

Pitfall 1: Treating Based on SPEP Alone

• Do not initiate therapy for plasma cell disorders based solely on SPEP without confirmatory immunofixation 4
• The 47% positive predictive value means that treating all positive SPEPs would result in unnecessary treatment in more than half of cases 4
• Always obtain tissue biopsy when plasma cell neoplasm is suspected, regardless of SPEP results 4

Pitfall 2: Ignoring Clinical Context

• False-positive rates increase dramatically in settings of inflammation, liver disease, or recent immunoglobulin administration 2, 3, 5
• Polyclonal hypergammaglobulinemia from chronic liver disease, autoimmune conditions, or chronic infections can create broad-based peaks misinterpreted as M-spikes 5
• Always correlate SPEP findings with clinical presentation and other laboratory markers 5

Pitfall 3: Over-reliance on Small M-Spikes

• M-proteins <2 g/L have unacceptable inter-laboratory variability and should be interpreted with extreme caution 3
• At concentrations of 0.5-1.0 g/L, the coefficient of variation between laboratories can exceed 30%, making quantification unreliable 3
• Consider that therapeutic monoclonal antibodies (daratumumab, elotuzumab) used in myeloma treatment can themselves create or obscure M-spikes 3

Pitfall 4: Missing Fibrinogen Interference

• Always check for fibrinogen contamination when a spike appears in the beta-gamma region 1
• Apply the γ/IgG ratio calculation: if >1.13, recollect the sample before pursuing extensive workup for monoclonal gammopathy 1
• Ensure proper sample collection technique with adequate clotting time (30 minutes at room temperature) before centrifugation 1

Algorithmic Approach to Suspected False-Positive SPEP

1. Review medication history: IVIG within 30 days? If yes, defer interpretation and retest in 4-6 weeks 2

2. Calculate γ/IgG ratio: If >1.13, suspect fibrinogen; recollect sample with proper clotting 1

3. Assess M-spike size: If <2 g/L, recognize high imprecision and require confirmatory testing before clinical action 3

4. Order confirmatory tests simultaneously: SIFE, serum free light chains, urine protein electrophoresis with immunofixation 4

5. Evaluate clinical context: Presence of hypergammaglobulinemia, liver disease, chronic infection, or autoimmune disease increases false-positive likelihood 5

6. If confirmatory tests are negative: Repeat SPEP in 3-6 months to ensure stability; do not pursue treatment 4

7. If diagnostic uncertainty persists: Obtain tissue biopsy rather than relying on serial SPEP monitoring 4