Background <p>Accurate detection and monitoring of monoclonal proteins in multiple myeloma (MM) are critical for diagnosis and treatment guidance. Conventional assays such as serum protein electrophoresis (SPEP), immunofixation electrophoresis (SIFE), and serum-free light chain (SFLC) testing can yield discordant or inconclusive results, particularly in patients with low M-protein levels.</p> Objectives <p>To evaluate the concordance and discordance between SIFE and SFLC assays in MM and to assess the clinical utility of the heavy/light chain (HLC) assay as a complementary quantitative tool.</p> Methods <p>We retrospectively analyzed 1,310 MM patient samples. The concordance and discordance between SIFE and SFLC were assessed, and the analytical performance of HLC measurements was evaluated relative to conventional assays.</p> Results <p>The overall discordance rate between SIFE and SFLC was 26.9%, with SIFE–/SFLC + most frequent in light chain MM and SIFE + /SFLC– observed in patients who achieved a very good partial response (VGPR). HLC strongly correlated with total immunoglobulins (combined immunoglobulin G [IgG]/immunoglobulin A [IgA], <i>R</i>^<i>2</i> = 0.99) and SPEP (<i>R</i>^<i>2</i> = 0.94). HLC ratios demonstrated excellent concordance with SIFE (<i>κ</i> = 0.83, 95% confidence interval [CI] 0.72–0.93) but poor agreement with SFLC ratios (<i>κ</i> = 0.01, 95% CI − 0.18–0.20). Using the HLC response model, 15 patients classified as having VGPR by the International Myeloma Working Group (IMWG) criteria were reclassified as having a partial response due to a dHLC reduction of &lt; 90%.</p> Conclusions <p>This large retrospective study provides a comprehensive evaluation of SIFE and SFLC discordance in MM, offering real-world evidence of their complementary diagnostic and monitoring roles. These findings support the integration of HLC testing into existing response assessment strategies to enhance the accuracy of treatment evaluation and the interpretation of discordant results.</p>