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Serum Protein Electrophoresis

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Presentation on theme: "Serum Protein Electrophoresis"— Presentation transcript:

1 Serum Protein Electrophoresis
Dr. Nitin Inamdar Department of Biochemistry Tata Memorial Center, Parel, Mumbai.

2 Electrophoresis Phoresis: Separation or migration or movement
Electro: Under influence of electric field

3 Constant current 5mAM/Slide
Serum Electrophoresis Mixture of proteins Amino acids: Amino group (NH2) Carboxyl group (COOH) Two amino acids join each other with polypeptide bond to form polypeptide chain & many polypeptide chains make a protein. Ph 8.6/0.075M Barbitone buffer 1%Agarose Constant current 5mAM/Slide

4 Electrophoresis separates proteins based on
their physical properties. Serum is placed on a specific medium, and a charge is applied. The net charge (positive or negative) and the size and shape of the subsets of these proteins are used in interpreting the results.

5 Serum Electrophoresis
Paper electrophoresis Starch gel electrophoresis Agarosre gel electrophoresis Cellulose acetate gel electrophoresis Capillary gel electrophoresis

6 Serum protein electrophoresis on agarose gel is a type of horizontal gel electrophoresis

7 Equipment used for the gel electrophoresis
power supply (direct current) containers for staining and destaining gel electrophoresis chamber applicator

8 Various electrophoresis
Serum/protein electrophoresis: plasma cell dyscrasia, nephrotic syndrome, etc. Hemoglobin electrophoresis: thalassemia, hemoglobinopathies like sickle cell anemia, HbC, HbD, etc. Urine electrophoresis: BJP in plasma cell dyscrasia

9 Indication of serum electrophoresis
Diagnostic: Diagnosis of plasma cell dyscrasia Diagnosis of Waldenstrom’ macroglbulinemia Monitoring of disease: Monitoring of plasma cell dyscrasia

10 Method of detection Serum protein electrophoresis
Quantitative immunoglobulins (nephelometry) Immunofixation, immunoelectrophoresis and immunodiffusion Urine studies light chain

11 Albumin Globulins: α1, α2, β1, β2, gamma A/G Ratio: 2.0

12 Normal serum electrophoresis
Anode +ve Cathode -ve Normal serum electrophoresis pH=8.6, 1% agarose gel, sodium barbitone buffer

13 Abnormal serum electrophoresis showing M band
Anode +ve Cathode -ve Abnormal serum electrophoresis showing M band

14 Normal serum electrophoresis

15 Abnormal serum electrophoresis showing M band

16 WHO Classification of Plasma Cell Neoplasms
Plasma cell myeloma (MM) Plasma cell myeloma variants Non secretory myeloma Indolent Myeloma Smoldering Myeloma Plasma cell leukemia Plasmacytomas Solitary plasmacytoma of bone Extramedullary plasmacytoma Immunoglobulin deposition diseases Primary Amyloidosis Systemic light and heavy chain deposition disease Osteosclerotic myeloma (POEMS syndrome) Heavy chain disease Gamma /Mu / Alpha

17 Diagnostic criteria for plasma cell dyscrasia
Symptomatic plasma cell myeloma: BM clonal plasma cells or Plasmacytoma Marrow plasmacytosis (>30%) M-component in serum or urine Serum: Ig G >3 g/dL, Ig A >2.5 g/dL or Urine: >/= 1g/ day of λ or κ [BJP] Asymptomatic plasma cell myeloma: M band in serum at myeloma levels > 3 gm/dL and /or 10% or more clonal plasma cell in BM No related organ or tissue impairment (end organ damage or bony lesion) or myeloma-related symptoms MUGS: M-Component, present but less than 3 gm/dL Marrow Plasmacytosis < 10% No Lytic bone lesions or related organ impairment etc

18 Immunoglobin (Ig) structure and types
Malignant Plasma Cell (Clonal Proliferation) produces one type of Ig either A,M, G, D or E with kappa or lambda light chains in excess amount.

19 Various patterns



22 Analbuminemia



25 Haptoglobin phenotype






31 Bis-albuminemia


33 Nephrotic syndrome

34 Hypergammaglobulinemia

35 M band in β region

36 M band in γ region



39 Nephelometry Antigen-antibody complexes, when formed at a high rate, will precipitate out of a solution resulting in a turbid or cloudy appearance. Nephelometry: is indirect measurement of light scattered by the antigen-antibody complexes.

40 Immunofixation electrophoresis
After separation of serum proteins by electrophoresis separated serum in different lanes is incubated with monospecific anti sera against IgA, M G and κ λ











51 Intravascular hemolysis


53 light chain myeloma A type of multiple myeloma in which plasma cell tumors produce only monoclonal light chain proteins. Persons with light chain disease may develop lytic bone lesions, hypercalcemia, impaired kidney function, and amyloidosis.

54 Freelite test Free Kappa Free Lambda Kappa/ Lambda ratio Normal serum
mean median % concentration concentration range Free Kappa 8,36 mg/l ,30 mg/l ,30 – 19,40 mg/l Free Lambda 13,43 mg/l ,40 mg/l ,71 – 26,30 mg/l Kappa/ Lambda ratio mean median total range 0, , ,26 – 1,65


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