Serum protein electrophoresis

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Serum protein electrophoresis
Normal serum protein electrophoresis diagram with legend of different zones.
MeSHD001797
protein electrophoresis
gel

Serum protein electrophoresis (SPEP or SPE) is a laboratory test that examines specific

alpha-2 globulins, beta 1 and 2 globulins, and gamma globulins
.

Acetate or gel electrophoresis

Proteins are separated by both electrical forces and electroendoosmostic forces. The net charge on a protein is based on the sum charge of its amino acids, and the pH of the buffer. Proteins are applied to a solid matrix such as an agarose gel, or a cellulose acetate membrane in a liquid buffer, and electric current is applied. Proteins with a negative charge will migrate towards the positively charged anode. Albumin has the most negative charge, and will migrate furthest towards the anode. Endoosmotic flow is the movement of liquid towards the cathode, which causes proteins with a weaker charge to move backwards from the application site. Gamma proteins are primarily separated by endoosmotic forces.[4] The drawing of the electrophoretic bands provided by the laboratory may be difficult to remember, and medical students, residents, nurses, and non-specialized medical practitioners may find visual mnemonics useful to recall the five main bands and the shape of normal serum electrophoresis.[5]

Visual mnemonics for serum protein electrophoresis, right hand

Capillary electrophoresis

In capillary electrophoresis, there is no solid matrix. Proteins are separated primarily by strong electroendosmotic forces. The sample is injected into a capillary with a negative surface charge. A high current is applied, and negatively charged proteins such as albumin try to move towards the anode. Liquid buffer flows towards the cathode, and drags proteins with a weaker charge.[6][7]

Serum protein fractions

Serum protein electrophoresis normal results

Albumin

acetylsalicylic acid, and occasionally from tryptic digestion in acute pancreatitis.[citation needed
]

Absence of albumin, known as analbuminaemia, is rare. A decreased level of albumin, however, is common in many diseases, including liver disease, malnutrition, malabsorption, protein-losing nephropathy and enteropathy.[9]

Albumin – alpha-1 interzone

Even staining in this zone is due to alpha-1 lipoprotein (

high density lipoprotein – HDL). Decrease occurs in severe inflammation, acute hepatitis, and cirrhosis. Also, nephrotic syndrome can lead to decrease in albumin level; due to its loss in the urine through a damaged leaky glomerulus. An increase appears in severe alcoholics and in women during pregnancy and in puberty.[citation needed
]

The high levels of AFP that may occur in hepatocellular carcinoma may result in a sharp band between the albumin and the alpha-1 zone.[citation needed]

Alpha-1 zone

alpha 1 antitrypsin (AAT) constitutes most of the alpha-1 band. Alpha-1 antitrypsin has an SG group and thiol compounds may be bound to the protein altering their mobility. A decreased band is seen in the deficiency state. It is decreased in the nephrotic syndrome[10] and absence could indicate possible alpha 1-antitrypsin deficiency. This eventually leads to emphysema from unregulated neutrophil elastase activity in the lung tissue. The alpha-1 fraction does not disappear in alpha 1-antitrypsin deficiency, however, because other proteins, including alpha-lipoprotein and orosomucoid, also migrate there. As a positive acute phase reactant, AAT is increased in acute inflammation.[citation needed
]

Bence Jones protein may bind to and retard the alpha-1 band. [citation needed]

Alpha-1 – alpha-2 interzone

Two faint bands may be seen representing

acute phase protein.[citation needed
]

Alpha-2 zone

This zone consists principally of

phagocytes). Haptoglobin is raised as part of the acute phase response, resulting in a typical elevation in the alpha-2 zone during inflammation. A normal alpha-2 and an elevated alpha-1 zone is a typical pattern in hepatic metastasis
and cirrhosis.

Haptoglobin/haemoglobin complexes migrate more cathodally than haptoglobin as seen in the alpha-2 – beta interzone. This is typically seen as a broadening of the alpha-2 zone.

Alpha-2 macroglobulin may be elevated in children and the elderly. This is seen as a sharp front to the alpha-2 band. AMG is markedly raised (10-fold increase or greater) in association with glomerular protein loss, as in nephrotic syndrome. Due to its large size, AMG cannot pass through glomeruli, while other lower-molecular weight proteins are lost. Enhanced synthesis of AMG accounts for its absolute increase in nephrotic syndrome. Increased AMG is also noted in rats with no albumin indicating that this is a response to low albumin rather than nephrotic syndrome itself[11]

AMG is mildly elevated early in the course of diabetic nephropathy.[citation needed]

Alpha-2 - beta interzone

Cold insoluble globulin forms a band here which is not seen in plasma because it is precipitated by heparin. There are low levels in inflammation and high levels in pregnancy.[citation needed]

Beta lipoprotein forms an irregular

hypercholesterolaemia, hypertriglyceridemia
, and in the nephrotic syndrome.

Beta zone

iron deficiency anemia, pregnancy, and oestrogen therapy. Increased beta-1 protein due to LDL elevation occurs in hypercholesterolemia. Decreased beta-1 protein occurs in acute or chronic inflammation.[citation needed
]

Beta-2 comprises C3 (

complement protein 3). It is raised in the acute phase response. Depression of C3 occurs in autoimmune disorders as the complement system is activated and the C3 becomes bound to immune complexes and removed from serum. Fibrinogen, a beta-2 protein, is found in normal plasma but absent in normal serum. Occasionally, blood drawn from heparinized patients does not fully clot, resulting in a visible fibrinogen band between the beta and gamma globulins.[citation needed
]

Beta-gamma interzone

skin disease, or rheumatoid arthritis (increased IgA). Fibrinogen from plasma samples will be seen in the beta gamma region. Fibrinogen, a beta-2 protein, is found in normal plasma but absent in normal serum. Occasionally, blood drawn from heparinized patients does not fully clot, resulting in a visible fibrinogen band between the beta and gamma globulins.[citation needed
]

Gamma zone

The

hypogammaglobulinaemia), or have an abnormal peak or peaks. Note that immunoglobulins may also be found in other zones; IgA typically migrates in the beta-gamma zone, and in particular, pathogenic immunoglobulins may migrate anywhere, including the alpha regions.[citation needed
]

Hypogammaglobulinaemia is easily identifiable as a "slump" or decrease in the gamma zone. It is normal in infants. It is found in patients with X-linked agammaglobulinemia. IgA deficiency occurs in 1:500 of the population, as is suggested by a pallor in the gamma zone. Of note, hypogammaglobulinema may be seen in the context of MGUS or multiple myeloma.[citation needed]

If the gamma zone shows an increase the first step in interpretation is to establish if the region is narrow or wide. A broad "swell-like" manner (wide) indicates polyclonal immunoglobulin production. If it is elevated in an asymmetric manner or with one or more peaks or narrow "spikes" it could indicate clonal production of one or more immunoglobulins,[13]

Polyclonal gammopathy is indicated by a "swell-like" elevation in the gamma zone, which typically indicates a non-neoplastic condition (although is not exclusive to non-neoplastic conditions). The most common causes of polyclonal hypergammaglobulinaemia detected by electrophoresis are severe infection, chronic liver disease, rheumatoid arthritis, systemic lupus erythematosus and other connective tissue diseases.[citation needed]

A narrow spike is suggestive of a monoclonal gammopathy, also known as a restricted band, or "M-spike". To confirm that the restricted band is an immunoglobulin, follow up testing with

Waldenström's macroglobulinaemia (IgM), monoclonal gammopathy of undetermined significance
(MGUS), amyloidosis, plasma cell leukemia and solitary plasmacytomas also produce an M-spike.

Oligoclonal gammopathy is indicated by one or more discrete clones.[citation needed]

tumour cells.[citation needed
]

References

  1. PMID 21374283
    .
  2. ISBN 978-1-933864-75-4
    .
  3. PMID 4158264
    .
  4. ^ Harris 2012, pp. 9–16.
  5. ^ Medina-De la Garza CE, García-Hernández M, Castro-Corona MA. Visual mnemonics for serum protein electrophoresis https://www.tandfonline.com/doi/full/10.3402/meo.v18i0.22585
  6. ^ Harris, 2012 & pages 117–123.
  7. ISBN 0340-81213-3
    .
  8. PMID 10437648
    .
  9. ^ Peralta, Ruben; Rubery, Brad A (July 30, 2012). Pinsky, Michael R; Sharma, Sat; Talavera, Francisco; Manning, Harold L; Rice, Timothy D (eds.). "Hypoalbuminemia". Medscape. Retrieved 2 October 2013.
  10. PMID 19870946
    .
  11. PMID 9453001
    .
  12. ^ Keren 2003, pp. 93–97.
  13. ^ Tuazon, Sherilyn Alvaran; Scarpaci, Anthony P (May 11, 2012). Staros, Eric B (ed.). "Serum protein electrophoresis". Medscape. Retrieved 2 October 2013.
  14. PMID 27028734
    .
  15. ^ Harris 2012, p. 60.
  16. PMID 20713974
    .

External links

 
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