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Clear identification of κ and λ molecules became possible with the use of antibodies specific for each type of protein. Immunodiffusion was initially used [65], followed by immunoelectrophoresis in 1953 [66], radial immunodiffusion and ultimately nephelometry and turbidimetry. The first successful attempt to measure FLCs in serum was in 1975. Size-separation column chromatography [67][68][69], was used to isolate them from intact immunoglobulins, prior to analysis. Although the results were accurate and showed the potential use of serum analyses, these assays were clearly impractical for routine use. However, serum immunoassays for Bence Jones protein (serum FLCs; sFLCs) remained unattainable because the antibodies could not distinguish between sFLCs and the overwhelming amounts of light chains bound in intact immunoglobulin molecules.

Subsequent assays focussed on the use of antibodies directed against “hidden” epitopes on FLC molecules. These are located at the interface between the light and heavy chains of intact immunoglobulins and become detectable when the FLCs are unbound. Radioimmunoassays and enzyme immunoassays using polyclonal antisera against FLCs were used to analyse urine samples, but specificity remained inadequate for serum measurements [70][71] and variations in FLC polymerisation caused measurement errors [72][73].

The use of monoclonal antibodies was an obvious approach to improving specificity, but satisfactory reagents were difficult to develop and their use was initially restricted to radio-immunoassays and enzyme immunoassays [74][75][76][77]. Early attempts to develop turbidimetric [78][79] Chapter 16 and latex-enhanced nephelometric assays [80] using polyclonal antibodies could not detect normal sFLC concentrations, and cross-reactions with intact immunoglobulins were unacceptable. In 2001, immunoassays based on polyclonal antibodies were finally developed that could measure FLCs at normal serum concentrations [81]. Their utility was quickly made apparent when monoclonal FLCs were detected in the sera of most patients classified as having nonsecretory MM (Chapter 16) [15]. Furthermore, as described in The Lancet, all of 224 patients with light chain MM who had Bence Jones proteinuria also had elevated sFLC concentrations (Chapter 15) [83]. The serum tests were also better at detecting residual disease than urinalysis (Chapter 24).

These results and many others, heralded the widespread use of sFLC immunoassays, and the incorporation of Freelite® assays into International Myeloma Working Group guidelines (Chapter 25).

References