Bence-Jones Protein

Article Author:
Neeraj Ramakrishnan

Article Editor:
Ishwarlal Jialal

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Ishwarlal Jialal

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Sarosh Vaqar
Mark Pellegrini
James Hughes
Beenish Sohail
Hajira Basit
Phillip Hynes
Sandeep Sekhon

4/11/2019 3:18:09 PM


Bence Jones protein (BJP) was first described in a patient admitted to St. George’s Hospital in London under the care of Drs. Watson and MacIntyre for vague continuous pain to the chest, back, and pelvis in 1845. Dr. Henry Bence Jones tested this urine and found a substance in it that was precipitated by the addition of nitric acid. Jones proceeded to call this substance “hydrated deutoxide of albumen.” It is important to remember that at that time all urine proteinuria was referred to as albuminuria. After this patient died, his sternum, as well as various parts of his vertebrae, were noted to be soft and fragile, while multiple hemorrhagic cavities were present in the bone. The patient’s cause of death was called “atrophy from albuminuria.” The actual term, Bence Jones Proteins, was used in 1880 by Dr. Fleischer. Its peculiar characteristics on heating first characterized BJP: precipitation of the urine at 40 to 60 degrees C and re-dissolving of the precipitate at 100 degrees C.[1][2]

While this was a seminal observation that led to the first description of multiple myeloma (MM), it did not hold up to scrutiny over time. Today BJP is known as the light chain of immunoglobulins without the accompanying heavy chain and can be accurately quantified by electrophoretic techniques including immunofixation electrophoresis (IFE). The other crucial teaching point is that BJP is undetectable by dipsticks used to detect proteinuria since they detect albumin and not BJP. In this brief review, we describe the evolution of BJP, its biochemistry, and high clinical relevance.

In 1939 electrophoresis was applied for the first time to study multiple myeloma by Longsworth. In 1953 Grabar and Williams first described the methods of immunofixation and direct immunoelectrophoresis, which increased detection of small monoclonal light chains not detectable on basic gel electrophoresis, which gave rise to the classic “M spike” seen in the gamma region in patients with multiple myeloma. In 1956, Korngold identified the two classes of BJP: Kappa and Lambda light chains. In 1962, Drs. Edelman and Porter received the Nobel Prize in Medicine for their work in elucidating the chemical structure of antibodies. The M-spike of a particular patient with multiple myeloma was broken down into heavy and light chains. Edelman demonstrated that the light chains of this M-spike were identical to the BJP that the patient excreted in his urine. He expanded on this work by comparing reduced myeloma proteins from different patients. When each of these proteins was reduced, alkylated, and put through starch gel electrophoresis, they exhibited a unique migration pattern. Similar to BJP, when Edelman heated samples containing light chain from normal human serum gamma globulins, they became insoluble and re-solubilized with continued heating. In 1967, Dr. Putnam demonstrated that different BJP had distinct peptide sequences. This differentiation further supported Dr. Edelman’s observation that no two BJP “had the same mobility pattern.”[2]

BJP, or free light chains, are found in the urine as low molecular weight monomers, dimers, or high molecular weight polymers. Contrastingly, Bence Jones proteins are present in the serum as tetramers. Their molecular weight is 22000, and the kidney metabolizes them through the processes of glomerular filtration, proximal renal tubular absorption, and renal catabolism. Bence Jones proteins spill into the urine once capacity for tubular absorption becomes saturated.[3]

Specimen Collection

A clean catch early morning urine sample is needed to screen for Bence Jones proteins by urine protein electrophoresis and immunofixation electrophoresis. A 24-hour urine catch is desirable to accurately quantify the amount of BJP excreted since it aids in the diagnosis of multiple myeloma. This specimen requires no preservative and can be kept at room temperature.[4]


Methods for detection of Bence Jone proteins include conventional high-resolution electrophoresis or capillary zone electrophoresis followed by confirmation through immunofixation electrophoresis (preferable) or immunoelectrophoresis.

The various electrophoretic methods are conducted on a medium of agarose and cellulose acetate. Differences between these methods arise regarding the volume of sample loaded and sensitivity of protein stain utilized. Depending on these characteristics, patient urine must be concentrated before analysis to ensure appropriate sensitivity. Coomassie Blue and Colloidal Gold serve as the stains used to detect Bence Jones proteinuria. Colloidal Gold is the most sensitive stain but has drawbacks such as demanding technique and lack of stoichiometry between uptake by BJP and by other proteins.[3]

After gel electrophoresis, follow-up immunofixation electrophoresis serves as immunologic confirmation of potential Bence Jones proteinuria type ( kappa or lambda) and is the most sensitive measure of BJP. Immunofixation electrophoresis follows with antisera to IgG, IgA, IgM, total kappa, and total lambda. In the instances of a monoclonal protein visualized without a corresponding heavy chain, additional immunofixation gel with antisera to delta and epsilon heavy chains can be performed but is not cost effective. The issue of antigen excess with immunofixation electrophoresis can be minimized through decreased initial loading of antigen. Immunoblotting is an alternative to immunofixation electrophoresis but is more cumbersome and is not common practice in clinical laboratories.[5][3]

An advance in the field is the ability to accurately measure free light chains by immunoassay and express them as a ratio. This test, the free light chain assay, is now used routinely to diagnose and monitor patients with plasma cell dyscrasias. The International Myeloma Working Group now suggests using serum free light chain (FLC) assay instead of urine protein electrophoresis and immunofixation electrophoresis if multiple myeloma is suspected. Quantitative free light chain assays utilize antisera directed against epitopes that are exposed only when light chains are free. Kappa and lambda free light chain concentrations are quantified, and the ratio of kappa to lambda is used to determine unbalanced light chain synthesis. The free light chain assay is a significant addition to the clinical management of plasma cell dyscrasias due to its increased sensitivity for the determination of serum free light chains compared to previous methods. It also is a better marker of early remission and early relapse of multiple myeloma and prognostic marker for identification of risk of progression of monoclonal gammopathy of undetermined significance and smoldering myeloma. Its short physiologic half-life allows the free light chain assay to be a useful tool for real-time monitoring for response to treatment and disease progression.[6] However the FLC assay has pitfalls including the prozone phenomena of antigen excess, excess variability, and non-linearity and hence IFE of urine is a valuable test for light chains.[7]

Overall, in light of the discussion of these various procedures, utilization of serum protein electrophoresis and free light chain assay will diagnose 100% of cases of multiple myeloma. This combination of testing poses as a cost-effective and accurate modality for many laboratories worldwide.[6] However, the immunofixation electrophoresis detection of BJP, commonly lambda light chains, is still required to detect amyloidosis.[5]

Normal and Critical Findings

Quantitation of Bence Jones proteins occurs through electrophoresis and densitometry. Laboratory analysis of patient urine can identify Bence Jones proteins at a threshold of 10 mg/L. It is important to note, however, that detection limits for Bence Jones proteins can only be approximate. The presence of Bence Jones proteins in the urine is usually considered abnormal and should raise suspicion for multiple myeloma or other plasma cell disorders.[3]

The free light chain (FLC) assay is quickly becoming the test of choice for detecting FLC. This assay measures free kappa or lambda light chains. The reference range for the k/l light chain ratio is 0.26 to 1.65. An increase in the ratio denotes increased kappa light chains and a decreased ratio of an excess of lambda.[6] However as stated above, this assay has several pitfalls also.


An issue is the phenomenon of light chain ladders in samples. Polyclonal light chains, usually kappa, tend to produce a characteristic banding pattern after electrophoresis and immunofixation electrophoresis. These light chain ladders are not Bence Jones proteins but commonly appear in urine samples of elderly populations suffering from tubular proteinuria due to inflammatory disease. Moreover, Bence Jones Proteins can sometimes co-migrate with the bands in these ladders. These ladders must be carefully examined to ensure that there are no concomitant Bence Jones proteins. However, the clinical presentation and other investigative findings help confirm the presence of BJP.[3]

Clinical Significance

Testing for Bence Jones proteinuria is indicated when plasma cell disorders such as multiple myeloma are suspected. Patient signs and symptoms that may prompt testing include anemia, hypercalcemia, and renal impairment. Bone manifestations such as painful lytic lesions and vertebral crush and long bone fractures are also indications. Increased suspicion for disorders such as multiple myeloma may also be prompted by symptoms of hyperviscosity like blurred vision, headaches, and epistaxis, as well as increased susceptibility to infection. Unfortunately, patients will initially present with nonspecific symptoms such as fatigue and back pain, which can often delay diagnostic testing such as assessing urine for Bence Jones proteinuria.[8]

At least 60% of patients with classical myeloma have BJP in their urine. More importantly, 20% of patients with multiple myeloma produce Bence Jones Proteins (light chains) only in the serum and urine without any heavy chains. Excessive secretion of Bence Jones Proteins causes acute kidney injury from tubular obstruction and tubulointerstitial inflammation, termed tubular nephropathy. Tubular damage results in the Fanconi syndrome with glycosuria, aminoaciduria, phosphaturia, and renal tubular acidosis type 2.[8]

Presence of Bence Jones proteinuria significantly raises suspicion for multiple myeloma and warrants referral to a hematology clinic. Bone marrow aspiration for plasma cells and investigation for BJP are also significant markers for prognostic progression regarding the treatment of multiple myeloma. Plasma cells in multiple myeloma can present in various forms, including as a mature, normal plasma cell, demonstrating features of immaturity such as a low cytoplasmic-nuclear ratio or loose chromatin, or present as bizarre “flame cells” with red cytoplasm. Bone marrow in patients with multiple myeloma generally is hypercellular and infiltrated diffusely by plasma cells[9]. Clonal bone marrow plasma cells equal to or greater than 10% or biopsy-proven or extramedullary plasmacytoma are criteria for the diagnosis of multiple myeloma.[10] More specifically, changes in the level of urinary light chain excretion can assess response to therapy. Direct positive effects on the treatment of multiple myeloma are defined as a reduction to 50% or less of the pretreatment value of urinary M-protein.[8] In patients with Waldenstrom macroglobulinemia and amyloidosis, BJP can also be present and crucial in the diagnosis. In amyloidosis, immunofixation electrophoresis is superior to the serum FLC assay for detecting light chains.[5]

Testing for BJP and interpreting its clinical implications is best accomplsihed by an interprofessional team that includes multiple health care professionals including physicians, NPs, PAs, nursing (particularly onconolgy specialy nurses), and laboratory persoinnel. [Level V]

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Bence-Jones Protein - Questions

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Bence-Jones protein is consistent with which one of the following?

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A 57-year-old female with an unremarkable past medical history is found to have Bence Jones proteins in her urine through gel electrophoresis when being screened for multiple myeloma. The consulting pathologist recommends a follow-up study to confirm the presence of Bence Jones proteins in this patient’s urine. Which of the following is a drawback that must be considered with this particular follow-up study?

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A 65-year-old patient with a past medical history of hypertension and diabetes mellitus type 2 presents with complaints of fatigue and lower back pain that has progressively worsened. The physical exam is significant for tenderness on palpation of the middle portion of the lower back. Follow-up labs are significant for calcium levels of 14 mg/dL, and a hemoglobin level of 10 g/dL. A diagnosis of multiple myeloma is considered. Which of the following tests should be performed next to check for Bence-Jones proteinuria?

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A 65-year-old male with a past medical history of hyperlipidemia, obstructive sleep apnea, and hypertension presents to the clinic with symptoms of progressively worsening blurred vision, headaches, and fatigue. The patient also complains of lower back pain that prevents him from engaging in exercise. Her metabolic panel reveals calcium of 11 mg/dl. Additionally, a free light chain assay is ordered. Suspecting the likely diagnosis, what is the benefit of this test compared to the standard combination of gel electrophoresis with follow-up immunoelectrophoresis?

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Bence-Jones Protein - References


Ribatti D, A historical perspective on milestones in multiple myeloma research. European journal of haematology. 2018 Mar;     [PubMed]
Clamp JR, Some aspects of the first recorded case of multiple myeloma. Lancet (London, England). 1967 Dec 23;     [PubMed]
Bhole MV,Sadler R,Ramasamy K, Serum-free light-chain assay: clinical utility and limitations. Annals of clinical biochemistry. 2014 Sep;     [PubMed]
Kaplan JS,Horowitz GL, Twenty-four-hour Bence-Jones protein determinations: can we ensure accuracy? Archives of pathology     [PubMed]
Willrich MA,Katzmann JA, Laboratory testing requirements for diagnosis and follow-up of multiple myeloma and related plasma cell dyscrasias. Clinical chemistry and laboratory medicine. 2016 Jun 1;     [PubMed]
Jialal I,Pahwa R,Beck RC,Schmotzer CL, Therapeutic Monoclonal Antibodies and the Value of the Free Light Chain Assay in Myeloma. American journal of clinical pathology. 2018 Oct 1;     [PubMed]
Beetham R, Detection of Bence-Jones protein in practice. Annals of clinical biochemistry. 2000 Sep;     [PubMed]
Smith D,Yong K, Multiple myeloma. BMJ (Clinical research ed.). 2013 Jun 26;     [PubMed]
Albagoush SA,Azevedo AM, Cancer, Multiple Myeloma 2019 Jan;     [PubMed]
Rajkumar SV, Multiple myeloma: 2016 update on diagnosis, risk-stratification, and management. American journal of hematology. 2016 Jul;     [PubMed]


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