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ABSTRACT: Free light chains (FLC) are a natural product of B lymphocytes and, as such, represent a quantifiable biomarker of cellular proliferation. Accurate measurement of the concentrations of these components in serum and urine provides a unique means of ascertaining B cell immunoglobulin synthesis during physiologic and, especially, pathologic states, where such information has important diagnostic and therapeutic implications. Previously, use of such quantitative assays has been limited due to the lack of potent serologic reagents specific for these components. We have immunized mice with kappa- and lambda-type monoclonal human light chains (Bence Jones proteins (BJP)) and have obtained monoclonal antibodies (MoAbs) that differentiate between unbound and bound light chains. These highly specific MoAbs were used to measure by ELISA the concentrations of FLC in the serum of 22 normal individuals and in urine from 16 of these subjects. The mean serum kappa and lambda FLC concentrations were found to be 16.6+/-6.1 microg/ml and 33.8+/-14.8 microg/ml, respectively. In contrast, the values for urinary kappa and lambda FLC were 2.96+/-1.84 microg/ml and 1.07+/-0.69 microg/ml, respectively. In each case studied, the serum kappa:lambda ratio was consistently less than that of urine (mean values, serum approximately 1:2; urine approximately 3:1). That the rate of synthesis of lambda-type FLC exceeded that of kappa was evidenced in assays of culture fluid supernatants of unstimulated normal peripheral blood mononuclear cells (PBMC), where the mean kappa:lambda ratio was determined to be 1:1.4. Metabolic studies in which mice were injected with pools of kappa- and lambda-type BJP prepared in ratios of 1:1, 1:2 and 1:4 demonstrated that, regardless of the proportion, kappa FLC were preferentially excreted. Our studies provide the first evidence that lambda FLC are secreted by normal PBMC at a greater rate than are kappa FLC, as evidenced in biosynthetic studies and by measurement of their serum concentrations. Further, we posit that quaternary structural differences between the two light-chain isotypes may account for the predominance of kappa versus lambda components in urine.
Clinical & Experimental Immunology 03/1998; 111(2):457-62. · 3.36 Impact Factor
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ABSTRACT: We have developed a novel immunization protocol for the production of a panel of high-affinity murine monoclonal antibodies (MoAbs) that are specific for each of the major human kappa and lambda light chain variable-region (VL) subgroups. Mice were injected with heat-precipitated human Bence Jones proteins or VL-related fragments emulsified in monophosphoryl lipid A (MPL) and trehalose dimycolate (TDM) at two- to four-week intervals over a seven-month period. A unique direct capturing enzyme-linked immunosorbent assay (ELISA) employing biotinylated monoclonal light chains was designed to select optimally immunized animals for hybridoma preparation and to screen culture supernatants for high-affinity anti-VL MoAbs. These methods have led to the generation of MoAbs that by ELISA react specifically with each of the four V kappa subgroups--V kappa I, V kappa II, V kappa III, and V kappa IV or five V lambda subgroups--V lambda I, V lambda II/V, V lambda III, V lambda IV, and V lambda VI. These reagents have been used successfully to establish, on the basis of VL subgroup, the monoclonal nature of serum or urinary immunoglobulins as well as those found in the cytoplasm or on the cell surface of monoclonal plasma cell or B-lymphocyte populations, respectively. The availability of anti-VL subgroup-specific MoAbs will facilitate the immunodiagnosis and study of patients with multiple myeloma, AL amyloidosis, and related B-cell proliferative disorders.
Hybridoma 09/1993; 12(4):475-83.
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ABSTRACT: Hybridomas producing antihuman light chain monoclonal antibodies (MoAbs) were derived from fusion of SP2/O mouse myeloma cells with splenic lymphocytes from mice repeatedly immunized with purified kappa- and lambda-type Bence Jones proteins representative of the major V kappa (V kappa I, V kappa II, V kappa III, V kappa IV) and V lambda (V lambda I, V lambda II/V, V lambda III, V lambda IV, V lambda VI) subgroups or gene families. Monoclonal antibodies were obtained that had specificity for constant-region (CL) determinants common to all kappa or lambda light chains (C kappa and C lambda, respectively) as well as for variable-region (VL) epitopes unique to each of the V kappa or V lambda subgroups. The capability of these reagents to recognize CL and VL determinants on monoclonal immunoglobulin (Ig) molecules was demonstrated in fluid-phase antigen-capturing enzyme-linked immunosorbent assay (ELISA), solid-phase ELISA, and immunoblotting. In addition, these antilight chain MoAbs were used to establish immunocytochemically the kappa or lambda type and VL-subgroup nature of light chains expressed by the cytoplasmic Ig of monoclonal plasma cell and surface Ig of B-lymphocyte populations, respectively. These antibodies facilitated the immunohistochemical detection and characterization of light-chain-associated amyloid (AL amyloid) and other types of light-chain-related tissue deposits. Furthermore, the anti-CL-specific MoAbs were used to measure serum and urinary Ig kappa and Ig lambda concentrations. Quantification of Bence Jones protein excretion, even in the presence of other urinary proteins, was possible using the highly sensitive anti-C kappa and anti-C lambda MoAbs reactive only with free light chains. The ability to identify and characterize, through the use of these antihuman light chain MoAbs, light-chain-related epitopes at the protein, cellular, and tissue level has clinical importance in the diagnosis and treatment of patients with monoclonal plasma cell and related B-cell immunoproliferative diseases.
American Journal of Clinical Pathology 08/1993; 100(1):67-74. · 2.60 Impact Factor
