ANNALS OF CLINICAL AND LABORATORY SCIENCE, Vol. 6, No. 1 Copyright @ 1976, Institute for Clinical Science

Bence Jones Proteins: Nature, Metabolism, Detection and Significance MICHAEL J. DEEGAN, M.D. Clinical Immunology Laboratory, Department of Pathology, The University of Michigan Ann Arbor, MI 48104

ABSTRACT The nature of Bence Jones proteins is developed by a review of the pertinent literature. This is followed by a discussion of their metabolism and catabolism under normal and abnormal clinical conditions. Various methods for the detection of Bence Jones proteins are critically reviewed and the significance of these pro­ teins in various disorders is assessed. Nature more than a century after the initial descrip­ tion. In 1845 Henry Bence Jones, a physician The modern era of Bence Jones (BJ) pro­ and chemical pathologist at St. George’s tein analysis began with a report by Hospital in London, received a specimen of Edelman and Gaily16 that BJ proteins shared urine from Dr. Thomas Watson. The urine extensive antigenic cross-reactivity with was from a patient with an illness now myeloma proteins and normal gamma glob­ known to be multiple myeloma. Accom­ ulins. Putnam35 compared peptide maps of panying the specimen was a note the text of pooled human gamma globulin, seven which is reproduced here.11 purified pathologic globulins and eight BJ “The tube contains urine of very high proteins. On the basis of these studies, he specific gravity. When boiled it becomes concluded that there were two types of BJ highly opaque. On the addition of nitric acid protein and that they represented incom­ it effervesces, assumes a reddish hue and be­ plete or aberrant polypeptide chains of comes quite clear; but as it cools assumes the normal gamma globulin. Mannik and consistence and appearance which you see. Kunkel,27 using Ouchterlony immuno­ Heat reliquefies it. What is it?” diffusion methods, demonstrated that ap­ This challenge to Bence Jones resulted in proximately 60 percent of normal gamma a series of publications in which he clearly globulin molecules carried one group of an­ described many of the physicochemical fea­ tigenic determinants and 30 percent carried tures of the urinary protein which later came a second distinctive group of antigenic to bear his name. In his laboratory investi­ characters. Thus, the normal kappa to gation, Bence Jones placed considerable em­ lambda ratio of two to one for serum im­ phasis on the thermal properties of this uri­ munoglobulins was initially described. nary substance and these features were used Schwartz and Edelman39 used two-dimen- as the basis for defining the presence or sional, high-voltage electrophoresis and absence of this protein in patient’s urine for compared the tryptic hydrolysates from the

38 BENCE JONES PROTEINS 3 9 light chain of a myeloma globulin and BJ great deal of diversity. In fact, several protein from the same patient. They found subgroups of the VL portion of both kappa that the two were identical. Based on this and lambda chains have been charac­ and other data they suggested that BJ pro­ terized.37,43 They undoubtedly play an im­ teins were not aberrant or incomplete portant role in the antigen binding site of polypeptide chains but rather were entire which this portion of the molecule is an inte­ light (L) chains which had not been incor­ gral part. porated into the autologous myeloma protein. Further evaluation2 of the tryptic peptide Metabolism maps of a larger number of BJ proteins from The intracellular events necessary for the both antigenic groups (kappa and lambda) production and secretion of an immuno­ revealed that a common set of peptides exist globulin molecule are quite similar to those for each type of BJ protein, but that the required for other proteins. Heavy (H) peptides common to the kappa group of BJ chains and light chains are synthesized on proteins are completely different than the separate polyribosomes and are then joined common peptides of the lambda group. in the cisternae of the endoplasmic re­ These observations led Putnam36 to propose ticulum.9,40 Under normal conditions (un­ that each BJ protein or L chain was com­ associated with malignancy) one H and one posed of a constant and a variable portion as L chain form an HL intermediate that com­ defined by their amino acid sequence. This bines with a second HL molecule to form a proposal has subsequently been confirmed complete immunoglobulin (H2L2).9 Control and expanded through elucidation of the of some of the intermediate synthetic events primary structure of many BJ proteins by appears to be a function (at least for IgG amino acid sequence analysis. These se­ synthesis) of the individual H chain quence analyses have shown that a BJ pro­ subgroup, each of which has a characteristic tein (or a normal light chain) is composed of number and location of disulfide bonds that approximately 214 amino acid residues. The are essential to interchain linkage.9 initial one-half (residues 1 to 107) or amino Initial investigations suggested that the terminal end represents the variable portion formation of immunoglobulin molecules was of the molecule (VL). The carboxy terminal characterized by the production of an excess portion (residues 108 to 214) corresponds to of light chains41 and this was considered the the constant portion of the polypeptide source of the small amount of L chains pres­ chain (CL). Thus the C L portion of a par­ ent in normal serum and urine. Other ticular kappa type BJ protein is essentially workers, however, have shown that balanced identical to the CL region of all other type synthesis of H and L chains is most charac­ kappa BJ proteins and normal immunoglob­ teristic of nonmalignant immunoglobulin ulins containing kappa light chains, but production.1 These investigators also dem­ differs considerably from the CL region of all onstrated the existence of a small, rapidly type lambda proteins. An exception to this utilized intracellular pool of free light chains constancy is the occurrence of the genetic that may be important in controlling the marker Inv at position 191 on the kappa assembly of immunoglobulins.1 Their results chain.3 Various amino acids substituted in did not exclude the possibility that a small this position account for the different Inv amount of light chain might appear in the allotypes.48 In contrast to the constant por­ circulation as a result of cell death, cyto­ tion of the BJ protein or light chain is the plasmic shedding or unbalanced synthesis by variable region where there appears to be a a limited number of cells. 4 0 DEEGAN In malignant plasma cells of human and maleate or in clinical disorders such as the animal origin, a wide spectrum of synthetic Fanconi syndrome, Wilson’s disease and events has been shown to occur. Zolla and chronic cadmium poisoning.30 In these con­ coworkers55 demonstrated three basic pat­ ditions the processes of protein uptake and terns of immunoglobulin synthesis by ma­ catabolism diminish and increasing amounts lignant plasma cells: (1) production of only of small molecular weight proteins appear in L chains, (2) balanced synthesis where H the urine. In conditions characterized by a and L chains are made in equal amounts and selective proximal tubular disease, there is (3) an unbalanced synthesis in which intact usually little or no decrease in the immunoglobulin molecules and excess L glomerular filtration rate. However, when chains are made and secreted. It was also renal damage is more severe and involves the demonstrated that the formation of an HL glomeruli and tubules, there is a decrease of intermediate was common in the synthesis of the glomerular filtration rate. This results in a complete IgG myeloma protein and that less protein being filtered and presented to dimerization of L chains occurred intracellu- the tubular epithelium; therefore, ca­ larly.55 Excretion of the monoclonal L chain tabolism and excretion are diminished. In (i.e., BJ protein) into the urine as a monomer addition, many small proteins (e.g., L or a dimer was usually the same as the state chains) that would have undergone ca­ of polymerization when secreted by the ma­ tabolism or been excreted by the kidneys lignant plasma cell. However, in a number remain in the circulation and have a of cases where free light chains were clearly significantly prolonged half-life. demonstrated to be secreted by the cells, Individuals with normal renal function they were not detected in the urine. This remove labelled BJ proteins at a rate of 10 to suggests that the presence or absence of BJ 42 percent of the intravascular pool each protein in the urine is dependent on the rate hour; the half-life of a BJ protein or L in the and amount of free light chain synthesized circulation is approximately four hours.29,42 and the patient’s renal status (vida infra). Miettinen and Kekki29 injected 131I-labelled The investigations of Solomon,42 Woch- BJ protein into normal subjects and found ner51 and Miettinen29 and their collaborators that only 3.6 percent of the labelled protein have all contributed to establishing the was recovered intact in the urine. The bulk kidney as the major organ responsible for of the injected BJ protein was degraded the catabolism of normal light chains and within the body. The kidney was clearly BJ proteins. In a normal individual, the demonstrated to be the site of this degra- glomerular filtration apparatus effectively dative process in the studies of Wochner and controls the amount and type of protein that others.51 They showed that the survival time enters the tubular lumen usually allowing of human BJ protein was quite short in both only trace amounts of albumin to be ex­ unoperated and ureterligated mice; but in creted while significantly larger amounts nephrectomized animals it was markedly of proteins with molecular weights less than prolonged. The location of this active ca­ 50,000 daltons are presented to the tubular tabolism within the kidney was the epithelial epithelium. The proximal tubular epithelium lining of the convoluted tubular cells. Tan actively takes up and catabolizes a large and Epstein48 studied homogenates of number of these smaller proteins and, thus, normal human kidney tissue and found that only a limited amount of the filtered protein the highest catabolic activity directed at L is ultimately excreted in the urine. The im­ chains was in the lysosomal rich fraction. portance of this tubular reabsorptive process This process was most active at pH 5.0 and has been shown by an analysis of the type of in the presence of a reducing agent. Interest­ proteinuria that results when the proximal ingly, the first step in the degradative tubules are selectively poisoned with sodium process resulted in the formation of in­ BENCE JONES PROTEINS 41 soluble polymers. This was followed by the (3) electrophoresis and immunoelectro- appearance of half molecules with either phoresis.26 Methods for accurate quanti­ constant or variable antigenic features upon tation of the amount of BJ protein have also continued incubation. Morphologic confir­ been described.5,54 mation of this work has demonstrated that For many years, BJ proteins were defined 80 percent of injected kappa chains are by their ability to precipitate when heated to reabsorbed by the proximal tubular cells 45 to 60° followed by their redissolution on where they form crystal-like structures boiling. Although the mechanism of this within phagolysosomes.12 phenomenon was not appreciated, these The concentration of L chains in normal unusual thermal properties provided labora­ urine is three to four micrograms per ml and tories with a relatively simple technique for the kappa to lambda ratio of 1.9 closely their identification. Putnam and his collabo­ simulates that of serum.49 The urinary L rators34 demonstrated that the extent of ag­ chains may exist as monomers, dimers and gregation and precipitation by these proteins tetramers with a greater tendency for was dependent on the pH, the ionic strength polymer formation as the concentration in­ and the electrolyte composition of the creases.6,7 Fragments of light chains which reaction mixture. The pH was the most im­ retain the antigenic features of the constant portant variable, with an optimum pre­ or variable portion of the molecule have also cipitation of all proteins occurring in the been detected in urine.44 range 4.6 to 5.4. Edelman and Gaily16 used Individuals with a compromised renal spectrofluorometric analysis to demonstrate status clear BJ proteins and L chains at a rate that BJ proteins and L chains underwent of 1.8 to 3.9 percent of the intravascular characteristic reversible molecular transi­ pool per hour.42 This is considerably lower tions when heated and that pH changes than their normal counterparts. The half-life markedly influenced these events. Solomon of a labelled BJ protein is also much longer and McLaughlin44 analyzed the thermal (8 to 32 hours vs. 4 hours in the normal sub­ solubility properties of a BJ protein and its ject29) in these individuals. Patients with isolated CLand VL fragments. They found multiple myeloma and normal renal that the whole molecule and its VL portion function have a great capacity to catabolize had identical heat precipitating properties BJ proteins in spite of the large amounts of while the C L portion had none. Differences monoclonal L chains that may be syn­ in the primary structure of the VL portion of thesized.42 However, declining renal status the molecule were thought to be responsible from any cause sets a vicious cycle in motion for the variable thermal solubility properties with rising serum levels of BJ protein, addi­ of individual BJ proteins. Ghose18 also found tional cast formation and further decline of that the VL portion of the molecule was the renal function.8 The insoluble polymers determinant of a BJ protein’s heat precipi­ formed during the normal sequence of cata­ tating qualities. This suggested that the bolic events take on added significance in formation of a precipitate is the result of this context and may contribute to the renal thermally induced conformational changes injury seen so often in patients with multiple with exposure of hydrophobic bonds leading myeloma and collectively termed “myeloma to increased aggregation and precipitation. kidney,” a lesion whose etiology continues In spite of our increased understanding of to defy clarification. the mechanism(s) responsible for the heat precipitation of BJ proteins, the use of this Detection test is justifiably declining. The reasons for Procedures for the identification of BJ pro­ this include (1) the necessity for rigid pH teins include (1) heat precipitation,34 (2 ) control,34 (2) the inability of the test to de­ precipitation with acids and/or salts13,31 and tect BJ protein at less than 145 mg per dl,26 4 2 DEEGAN (3) the recognition that other proteins (e.g., tained by layering the urine over con­ transferrin) found in the urine may closely centrated hydrochloric acid and noting the simulate the thermal solubility properties of formation of a precipitin ring. Interestingly, BJ protein7 and (4) the recognition that not Hobbs reported a 52 percent failure rate all BJ proteins, as defined immuno- when using the toluene-sulfonic acid chemically, possess the characteristic heat method of Cohen and Raducha.13 Stone and precipitating qualities.7,44 Lindstrom et al26 Frenkel46 using the same method obtained a reviewed the ability of a standard heat test positive result in 30 of 35 patients with L to detect BJ proteins in urines from patients chain myeloma. The failures all occurred in with multiple myeloma. They found the patients with less than 400 mg of protein per heat precipitation test positive in only 13.6 24 hours. In general, these tests are not percent of patients and recorded several sensitive to low levels of BJ protein, or they instances of BJ protein being present by im­ fail to discriminate BJ protein from other uri­ munologic methods when screening tests nary proteins. Attention should also be were repeatedly negative. This phenomenon called to the misleading results obtained has been confirmed in our laboratory in­ with the Dipstix“ test in patients with BJ cluding one instance of a patient with L proteinuria. This procedure is notable for its chain disease who was losing 10 to 14 g inconsistent and frequent negative results in of protein every 24 hours in his urine and the presence of significant BJ protein­ had several negative heat tests. Urine uria.20,48 In fact, a negative Dipstix test and a electrophoresis and immunoelectrophoresis positive sulfosalicylic acid test have been revealed that virtually all of his urinary pro­ proposed as a presumptive test for BJ protein tein was a lambda type monoclonal (BJ) pro­ in the urine.17 Laboratories using any of the tein. Recent publications confirm the inade­ above techniques should be aware of the quacy of heat tests to screen urines effec­ shortcomings inherent in most if not all of tively for BJ protein. Stone and Frenkel46 these procedures. noted that only six of the 18 urines obtained Immunochemical analyses have clearly from patients with L chain myeloma had a demonstrated the presence of gamma glob­ positive heat precipitation test. This is par­ ulin and gamma globulin subfragments in ticularly undesirable in a disorder like L urine from normal individuals.49 The latter chain disease where BJ proteinuria may be include intact IgG, IgG Fc fragment, the only laboratory manifestation of a free L chains and fragments and IgA monoclonal protein. The heat precipitation molecules of the secretory type. The IgG Fc test for BJ protein cannot be recommended fragment may account for 15 percent of the for the routine screening of urine. In the gamma globulin present in normal urine. It future, it will play a less important role in is the product of catabolic events involving the laboratory evaluation of patients with the complete IgG molecule. The L chains suspected immunoproliferative disorders. present in normal urine account for more Numerous methods that rely on the ability than half the globulin fraction. Their kappa of various acids and/or salts to precipitate to lambda ratio remains approximately two the BJ protein in urine have been proposed. to one and their concentration may vary They are reviewed by Naumann.31 Hobbs21 from 2.5 to 4.0 mg per liter.5,6 Immunoelec- evaluated six of the more commonly used trophoretic examination of the urinary L methods and compared the results with two chains reveals a diffuse electrophoretic mo­ variations of the heat precipitation test and bility comparable to that seen with normal immunoelectrophoresis. He reported failure serum immunoglobulins.21 The L chains in rates ranging from five to 52 percent, with the best results (5 percent failure rate) ob­ °Ames Division Miles Laboratories, Elkhart, IN BENCE JONES PROTEINS 4 3 normal urine may be monomers, dimers or new antiserum before it is put into routine trimers. use. Electrophoresis and immunoelectropho- The incidence of BJ proteinuria associated resis of a concentrated urine specimen is the with multiple myeloma and other disorders most sensitive method currently available is dependent on the method used for de­ for the detection of BJ proteinuria.21,26'46 tection. In large series, the heat test is The use of high quality anti-kappa and anti­ positive in only 50 percent of the tested lambda antisera in immunoelectrophoretic patients.25 The use of a concentrated urine techniques provides a dimension of specimen, electrophoresis and immunoelec­ specificity that is unavailable with any other trophoresis increases the number of positives method. Materials and technical methods to 70 or 80 percent.25,28 are readily available and will not be considered.10 However, some remarks on anti-light chain antisera appear warranted. Significance Kappa and lambda chains are a hetero­ A search for BJ protein is most often genous protein population. Antigenic dif­ associated with the laboratory evaluation of ferences have allowed the division of both a patient suspected of having multiple my­ light chain types into several subgroups.37,43 eloma. However, monoclonal L chains may Antisera prepared against certain BJ proteins be found in the urine of patients with other have been found to contain antibody popu­ immunoproliferative disorders15,28 (Wal- lations unique to antigenic sites on the VL denstrCm’s macroglobulinemia, chronic portion, the CL portion and the hinge region lymphocytic leukemia), amyloidosis,23 carci­ between CLand VL.43 Not all BJ proteins noma33 and perhaps in a few patients with no are capable of generating these various underlying disease.24 specificities. In addition, L chain antisera As previously noted, concentrated urine prepared using whole immunoglobulin or electrophoresis and immunoelectrophoresis Fab fragments that contain bound kappa or leads to the detection of BJ protein in ap­ lambda chains will have different properties proximately 70 percent of patients with my­ than antisera prepared with BJ protein as the eloma. The absence of BJ proteins in the immunogen.47,52 In particular, antisera other 30 percent of patients may reflect a against monoclonal L chains may react very lack of sensitivity in the method or a well with other BJ proteins of the same type balanced synthesis by the malignant plasma but poorly or not at all with a similar L chain cells so that no free monoclonal L chain is bound to an intact immunoglobulin. This available for excretion.55 antiserum may be useful in determining the Light chain disease is a myelomatous type of BJ protein present in serum or urine disorder in which BJ protein in the serum but may fail to detect monoclonal proteins and/or urine is the only monoclonal protein (especially IgA) present in serum and should detected.46,50 This entity comprises ap­ not be used to evaluate the kappa/lambda proximately 20 percent of the cases in large ratio in routine sera. Therefore, any labora­ series of myeloma. It is often a rapidly tory involved in the detection and analysis of progressive disease and is frequently asso­ BJ proteins by immunoelectrophoresis ciated with renal disease and amy­ should require complete information re­ loidosis. In other patients with multiple garding the preparation of antisera pur­ myeloma a phenomenon known as “BJ es­ chased from commercial sources. In ad­ cape” may occur.22,46 These are usually dition, panels of serum and urine known to patients with an IgG or IgA myeloma and contain certain monoclonal proteins should are initially responsive to chemotherapy but be frozen ( — 70°) and used to evaluate any later become refractory. This increased re­ 44 DEEGAN sistance to therapy is heralded by a rapid in­ the patients with benign monoclonal gam­ crease in the amount of monoclonal L chain mopathy will be necessary to assure that excreted but no corresponding increase of they were not in a premyelomatous state. the monoclonal IgG or IgA. “BJ escape” Thus one should be cautious in using BJ pro­ may occur as a result of clonal selection by teinuria to discriminate a benign from a ma­ the therapeutic agent. Experimental support lignant monoclonal gammopathy. for this concept is available.32 Amyloidosis and a plasma cell dyscrasia Waldenstrom’s macroglobulinemia is an­ occur more often in the same patient than other disorder in which the reported inci­ coincidence would allow.4,23 In most if not dence of BJ proteinuria varies from zero to all of these patients there is a monoclonal 60 percent as a result of different methods of protein—frequently a BJ protein. Until very testing.14,15,28 Urine protein electrophoresis recently, this relationship was unexplained. and immunoelectrophoresis will detect BJ However, in the past few years Glenner and proteins in fifty percent or more of these his colleagues19 have demonstrated, by patients. The urinary monoclonal L chain chemical methods, that the amyloid fibrils in and the L chain portion of the monoclonal a group of cases are complete immunoglob­ IgM found in the serum are the same in most ulin L chains or their variable (amino-ter­ instances. BJ proteinuria may also be found minal) fragments. This observation would in patients with lymphocytic lymphoma and explain the frequent occurrence of amy­ chronic lymphocytic leukemia.28,33 This is loidosis in patients with L chain disease.46 not surprising in light of the probable B cell Isobe and Osserman23 have detected a nature of most of these tumors.38 monoclonal protein in one hundred percent Monoclonal immunoglobulins, primarily of their patients with primary amyloidosis. IgG and IgA, are often present in patients Forty-six of their 50 patients had a BJ pro­ with nonmyelomatous diseases53 or perhaps tein, most often of lambda type. Stone and with no illness15 (benign monoclonal gam- Frenkel’s46 findings in their patients with mopathy). The reasons for this remain un­ amyloidosis and L chain disease are similar. known. However, the presence or absence of They observed that the patients who secrete BJ proteins in these patients has been used as monoclonal L chains with “amyloidogenic” a criterion for distinguishing the benign properties usually have a disease course gammopathy from those that are associated dominated by their amyloidosis rather than with a malignant immunoproliferative dis­ their myeloma. Similar observations have ease. Dammacco and Waldenstrom15 used been made in our patients. concentrated urine electrophoresis and im­ munoelectrophoresis and found that 23.8 percent of 42 patients with a benign Summary monoclonal gammopathy had a monoclonal Bence Jones proteins are monoclonal light L chain in their urine. The physicochemical chain proteins. They are made and secreted properties of the BJ proteins in these by plasma cells usually in association with a patients did not differ from those in the urine malignant immunoproliferative disorder. of myeloma patients. However, the amount They circulate in the plasma, are catabolized of BJ proteinuria never exceeded 60 mg by the kidney and excreted into the urine. per liter in the patients with a benign The heat tests traditionally used for the de­ monoclonal gammopathy. Concentrations tection of Bence Jones proteins have been varied widely in the myeloma patients and shown to lack specificity and sensitivity and most of them had significantly greater levels are being replaced by electrophoretic and with some patients excreting 10 g per liter of immunologic methods. Although BJ proteins monoclonal L chain. Longterm followup on are most often found in patients with mul­ BENCE JONES PROTEINS 4 5 tiple myeloma, they may be present in a va­ 17. Fine, J. D. and Rees, E. D.: Bence-Jones protein: Detection and implications. New Eng. J. Med. riety of other disorders. 290:106-107, 1974. 18. Ghose, A. C.: Thermally induced structural transi­ tions of the variable and constant halves of Bence References Jones proteins: Fluorometric studies. Biochem. Biophys. Res. Comm. 52:240-245, 1975. 1. Askonas, B. A. and Williamson, A. R.: Balanced 19. Glenner, G. G., Terry, W. D., and Isersky, C.: synthesis of light and heavy chains of immuno­ Amyloidosis: Its nature and pathogenesis. Seminars. globulin G. Nature 276:264-267, 1967. Hemat. 70:65-86, 1973. 2. Baglioni, C. and Cioli, D.: A study of immunoglob­ 20. Greenberg, M.S.: Detection of Bence Jones protein ulin structure II. The comparison of Bence-Jones in urine. New Eng. J. Med. 289:806-807, 1973. proteins by peptide mapping. J. Exp. Med. 21. Hobbs, J. R.: The detection of Bence-Jones pro­ 124:307-330, 1966. teins. Biochem. J. 99.15P, 1966. 3. Baglioni, C., Alescio-Zonta, L., Cioli, D., and Car- 22. Hobbs, J. R.: Growth rates and response to bonara, A.: Allelic antigenic factor Inv(a) of the treatment in human myelomatosis. Brit. J. Haemat. light chains of human immunoglobulins: chemical 76:607-617, 1969. basis. Science 752:1517-1519, 1966. 23. Isobe, T. and Osserman, E. F.: Patterns of 4. Barth, W. F., Willerson, J. T., Waldmann, T. A., amyloidosis and their association with plasma-cell and Decker, J. L.: Primary amyloidosis. Clinical, dyscrasia, monoclonal immunoglobulins and immunochemical and immunoglobulin metabolism Bence-Jones proteins. New Eng. J. Med. 290:473- studies in fifteen patients. Amer. J. Med. 47:259- 477, 1974. 273,1969. 24. Kyle, R. A., Maldonado, J. E., and Bayrd, E. D.: 5. Bell, C. E. and Chaplin, H.: Application of a Idiopathic Bence Jones proteinuria—a distinct modified technique to study urinary immunoglob­ entity? Amer. J. Med. 55:222-226, 1973. ulin light chain and whole IgG excretion in normal 25. Kyle, R. A.: Multiple myeloma: review of 869 cases. human subjects and patients with immune-related Mayo Clin. Proc. 50:29-40, 1975. disorders. J. Lab. Clin. Med. 75:636-653, 1970. 26. Lindstrom, F. D., Williams, R. C., Swaim, W. R., 6. Berggard, J. and Peterson, P. A.: Polymeric forms of and Freier, E. F.: Urinary light-chain excretion in free normal kappa and lambda chains of human im­ myeloma and other disorders—an evaluation of the munoglobulin. J Biol. Chem. 244:4299-4307, 1969. Bence-Jones test. J. Lab. Clin. Med. 77:812-825, 7. Bernier, G. M. and Putnam, F. W.: Polymerism, 1968. polymorphism, and impurities in Bence Jones pro­ 27. Mannik, M. and Kunkel, H. G.: Classification of teins. Biochim. Biophys. Acta 86:295-308, 1964. myeloma proteins, Bence Jones proteins, and 8. Bryan, C. W., Mclntire, K. R., and Princler, G. L.: macroglobulins into two groups on the basis of Effect of sustained diuresis on the renal lesions of common antigenic characters. J. Exp. Med. mice with BJ protein-producing tumors. J. Lab. 776:859-877, 1962. Clin. Med. 83:409-416, 1974. 28. McLaughlin, H.: The clinical significance of Bence 9. Buxbaum, J. N.: The biosynthesis, assembly, and Jones proteinuria. J. Clin. Path. 25:555-556, 1972. secretion of immunoglobulins. Seminars. Hemat. 29. Miettinen, T. A. and Kekki, M.: Effect of impaired 10:33-52, 1973. hepatic and renal function on 131I Bence Jones pro­ 10. Cawley, L. P.: Electrophoresis and Immunoelec­ tein catabolism in human subjects. Clin. Chim. Acta trophoresis, Boston, Little, Brown & Co., 1969. 78:395-407, 1967. 11. Clamp, J. R.: Some aspects of the first recorded case 30. Mogielnicki, R. P., Waldmann, T. A., and Strober, of multiple myeloma. Lancet 2.1354-1356, 1967. W.: Renal handling of low molecular weight pro­ 12. Clyne, D. H., Brendstrup, L. First, M. R., Pesce, teins. I. L-chain metabolism in experimental renal A. J., Finkel, P. N., Poliak, V. E., and Pirani, C. L.: disease. J. Clin. Invest. 50:901-909, 1971. Renal effects of intraperitoneal kappa chain in­ 31. Naumann, H. N.: Differentiation of Bence Jones jection. 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