4.**Multiple myeloma**

	- C—hypercalcemia
	- R—renal abnormalities (elevated creatinine)
	- A —anemia
	- B—bone lesions

There are exceptions in SPE findings in certain cases of multiple myeloma wherein the SPE does not reveal any significant alteration or a clue toward the diagnosis.

These variants of multiple myeloma characterized by an abnormal bone marrow (increased plasma cells) but a normal SPE are termed as nonsecretory myelomas

**9**

*Serum Protein Electrophoresis and Its Clinical Applications*

which account to 1–2% of multiple myelomas. In such cases, an immunoassay of free light chains (FLC) in serum provides a diagnostic clue toward NSMM which show a significant disproportionate elevation of usually a clone of light chains (kappa or lambda) with an alteration in kappa/lambda ratio (normal Ratio is between 0.60 and 1.65). A commonly encountered phenomenon with laboratory testing of FLC includes "prozone" effect or "hook" effect which occurs due to antigen excess and requires appropriate dilution to obtain reliable results.

Bence-Jones protein estimation in urine is an antique piece of laboratory evidence toward multiple myeloma, which is characterized by detection of light chains in urine. But since the methodology of testing is manual and does not provide standardization, this has been replaced by urine FLC analysis in laboratories practicing

One more valiant laboratory investigation which is an essential requisite for

SPE is a semiquantitative investigation which involves technical expertise to recognize the specific electrophoretic patterns and associate with various clinical conditions. This requires a laboratory practice integrated across various divisions of laboratory and with respective clinical and ancillary divisions of clinical medicine [1]. With respect to SPE, the laboratory professionals shall act as consultants to the clinical consultants. This is possible in scenarios where the clinician does not arrive at a provisional diagnosis of a gammopathy and the laboratory picks up the diagnostic clue toward gammopathy through an increased serum total protein level (<8 g/dl) and an altered serum albumin globulin (AG) ratio (which is usually altered in gammopathy). A normal AG ratio ranges between 1.2 and 1.8, while there is a significant reduction in the ratio in patients with gammopathy. This becomes an incidental finding which leads to a concept of "reflex" testing for multiple myeloma work-up including SPE, upon consent from the treating clini-

These are some of the common SPE patterns associated with various clinical

• **Nephrotic syndrome:** The albumin band is decreased due to hypoalbumin-

• **Cirrhosis or chronic liver disease:** A low albumin band due to significant hypoalbuminemia with a prominent beta-2 band and beta-gamma bridging

emia. In addition, the alpha-2 band may be more distinct.

• **Inflammation:** Increased intensity of alpha-1 and alpha-2 with a sharp leading edge of alpha-1 may be observed, but with chronic inflammation the albumin band may be decreased with increased gamma zone due to the polyclonal

One common principle employed in immunoelectrophoretic technique involves the use of specific antihuman immunoglobulins (e.g., Anti-IgG, Anti-IgA, Anti-Kappa, etc.) as a preprocessing step which results in precipitation of immunoglobulins if present and disappearance of the band/peak contributed by that specific immunoglobulin. Hence this technique is also known as immunosubtraction. This technique aids in typing the specific type of immunoglobulin (including the type of light chain) contributing to myeloma. This technique is supplemented by quantifi-

*DOI: http://dx.doi.org/10.5772/intechopen.88367*

good clinical laboratory practices (GCLP).

**4.1 SPE and its clinical significance**

cian and the patient.

gammopathy.

conditions:

multiple myeloma work-up includes immunoelectrophoresis.

cation of serum immunoglobulins by an immunoassay.

#### *Serum Protein Electrophoresis and Its Clinical Applications DOI: http://dx.doi.org/10.5772/intechopen.88367*

*Biochemical Testing - Clinical correlation and Diagnosis*

• No clinical symptoms/signs

• Normbal free light chain ratio in serum

• M protein (monoclonal band)—<3 g/dl

• Normal Free light chain ratio in serum

• M protein (monoclonal band)—<3 g/dl

• Abnormal free light chain ratio in serum

• M protein (monoclonal band)—>3 g/dl

• Abnormal free light chain ratio in serum

• Bone marrow biopsy—>10% plasma cells seen

○ R—renal abnormalities (elevated creatinine)

There are exceptions in SPE findings in certain cases of multiple myeloma wherein the SPE does not reveal any significant alteration or a clue toward the

These variants of multiple myeloma characterized by an abnormal bone marrow (increased plasma cells) but a normal SPE are termed as nonsecretory myelomas

○ C—hypercalcemia

○ A—anemia

4.**Multiple myeloma**

○ B—bone lesions

the four shall be present): ○ C—hypercalcemia

○ A —anemia

○ B—bone lesions

• Bone marrow biopsy—>10% plasma cells seen

which stands for (one of the four shall be present):

○ R—renal abnormalities (elevated creatinine)

• Clinically significant. Clinical diagnosis includes a tetrad of "ÇRAB"

• Clinical diagnosis includes a tetrad of "ÇRAB" which stands for (one of

3.**Smoldering myeloma**

2.**Monoclonal gammopathy of renal significance (MGRS)**

• Bone marrow biopsy—<10% plasma cells seen

• Renal disease with elevated serum creatinine

**8**

diagnosis.

which account to 1–2% of multiple myelomas. In such cases, an immunoassay of free light chains (FLC) in serum provides a diagnostic clue toward NSMM which show a significant disproportionate elevation of usually a clone of light chains (kappa or lambda) with an alteration in kappa/lambda ratio (normal Ratio is between 0.60 and 1.65). A commonly encountered phenomenon with laboratory testing of FLC includes "prozone" effect or "hook" effect which occurs due to antigen excess and requires appropriate dilution to obtain reliable results.

Bence-Jones protein estimation in urine is an antique piece of laboratory evidence toward multiple myeloma, which is characterized by detection of light chains in urine. But since the methodology of testing is manual and does not provide standardization, this has been replaced by urine FLC analysis in laboratories practicing good clinical laboratory practices (GCLP).

One more valiant laboratory investigation which is an essential requisite for multiple myeloma work-up includes immunoelectrophoresis.

One common principle employed in immunoelectrophoretic technique involves the use of specific antihuman immunoglobulins (e.g., Anti-IgG, Anti-IgA, Anti-Kappa, etc.) as a preprocessing step which results in precipitation of immunoglobulins if present and disappearance of the band/peak contributed by that specific immunoglobulin. Hence this technique is also known as immunosubtraction. This technique aids in typing the specific type of immunoglobulin (including the type of light chain) contributing to myeloma. This technique is supplemented by quantification of serum immunoglobulins by an immunoassay.

#### **4.1 SPE and its clinical significance**

SPE is a semiquantitative investigation which involves technical expertise to recognize the specific electrophoretic patterns and associate with various clinical conditions. This requires a laboratory practice integrated across various divisions of laboratory and with respective clinical and ancillary divisions of clinical medicine [1].

With respect to SPE, the laboratory professionals shall act as consultants to the clinical consultants. This is possible in scenarios where the clinician does not arrive at a provisional diagnosis of a gammopathy and the laboratory picks up the diagnostic clue toward gammopathy through an increased serum total protein level (<8 g/dl) and an altered serum albumin globulin (AG) ratio (which is usually altered in gammopathy). A normal AG ratio ranges between 1.2 and 1.8, while there is a significant reduction in the ratio in patients with gammopathy. This becomes an incidental finding which leads to a concept of "reflex" testing for multiple myeloma work-up including SPE, upon consent from the treating clinician and the patient.

These are some of the common SPE patterns associated with various clinical conditions:


is a characteristic feature. In addition, polyclonal hypogammaglobinemia is observed.


#### **4.2 Quality assurance in SPE**

Quality assurance in SPE is an essential prerequisite to ensure reliability of an SPE result [2]. There are two major aspects of analytical quality including precision (measure of precision) and accuracy (measure of trueness).

Good clinical laboratory practices demand processing of an internal quality control (IQC) for assessment of precision and external quality assurance (EQA)/ proficiency testing (PT testing) for accuracy assessment. IQC is a material which can be prepared in house (patient sample) or available commercially and is to be processed before a patient sample is taken up for processing.

The clinical laboratory has its responsibility to select and use an IQC which has a matrix comparable to patient sample, preferably covering the clinical decision point (cut off value that differentiates between a normal and abnormal result). EQA is an external assessment of the analytical quality wherein the laboratory processes a blinded sample and the results are compared against a reference method and/or against the consensus value of other participant laboratories for that specific sample.

The laboratory has to hold responsibility in selecting a suitable EQA provider who shall preferably be accredited to ISO 17043. If an EQA program is not available, the laboratories shall participate in exchange of samples with referral laboratories with a similar methodology and a comparable quality of testing standard.

### **5. Reporting of results and its standardization**

Reporting SPE requires interpretation of the electrophoretic pattern which is followed by comments of such an interpretation along with the piece of advice to the clinician if indicated. There is a big lacuna in the format of reporting of SPE, each laboratorian using his/her own means of interpreting and communicating. It is the need of the hour to have a standardized format of reporting SPE for ensuring patient safety and clinician follow-up. There are no international guidelines, though the working party on standardized reporting of protein electrophoresis which is an initiative of the Australasian Association of Clinical Biochemists has come out with a standardized format of reporting SPE.

#### **6. Conclusion**

In the current scenario, it becomes the responsibility of each and every laboratory to ensure that all relevant information is available in a SPE report, easily read, understood, and interpreted by a clinician. This becomes the core of a clinical laboratory practice.

**11**

**Author details**

Satish Ramanathan1

\* and Chakravarthy Narasimhachar Srinivas2

1 Division of Clinical Biochemistry, MIOT Hospitals, Chennai, Tamil Nadu, India

© 2019 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium,

2 Laboratory Medicine, MIOT Hospitals, Chennai, Tamil Nadu, India

\*Address all correspondence to: satishraksha1980@gmail.com

provided the original work is properly cited.

*Serum Protein Electrophoresis and Its Clinical Applications*

team for their invaluable contribution by sharing clinical cases.

We would wish to acknowledge and thank the management of MIOT hospitals for providing us with the infrastructure and technology to explore, learn, and

We would wish to thank our technical staff, Mr. Mathivanan Durairaj, and his

We wish to acknowledge Trivitron technologies for their valuable support in

installation and continual service and application support for Sebia CZE.

*DOI: http://dx.doi.org/10.5772/intechopen.88367*

contribute to our patients' well-being.

**Acknowledgements**
