January 2001

Accurate measurement and interpretation of plasma protein concentration and electrophoresis are important diagnostic tools in avian medicine. Protein electrophoresis is a practical and useful test in assessing  health status of psittacine birds. Dramatic changes in plasma protein fractions are evident in several diseases, and may help in establishing a diagnosis when other tests are equivocal. Serial electrophoresis is valuable in monitoring response to therapy.

Although differences between plasma and serum protein concentration and electrophoresis patterns are small, fibrinogen is virtually nonexistent in serum. As fibrinogen concentration is an important indicator of inflammation, plasma is the preferred sample for these tests. Since lithium heparinized blood samples are preferred for avian chemistry analysis, this plasma can also be used for protein electrophoresis.

At Antech Diagnostics, determination of total plasma proteins is performed by a biuret method. This is regarded as the most accurate way to measure total protein concentration in birds. Albumin concentrations, when measured as part of a chemistry profile, are determined using a wet chemistry dye-binding technique. The accuracy of albumin concentration measured by chemical methods is disputed by the claim that this methodology has no correlation to albumin concentration measured by electrophoresis. To investigate this concern, albumin concentrations were compared using wet chemistry and electrophoresis in 37 birds (25 psittacine and 12 non-psittacine). Although the wet chemistry albumin concentrations tended to be lower, results correlated well with those found on protein electrophoresis (see Figure 1). Thus, while electrophoretic measurement of albumin concentration in birds is considered by some to be the "gold standard", the wet chemistry dye-binding technique provides an accurate assessment of avian albumin concentration.

Avian plasma proteins (albumin and globulins), except for immunoglobulins, are manufactured in the liver. Albumin is the largest single plasma protein fraction in the healthy patient. It serves as the major reservoir of protein, is a main contributor of colloidal osmotic pressure, is involved in acid-base balance, and acts as a transport carrier for small molecules such as vitamins, minerals, hormones, fatty acids, and many drugs. Increases in albumin concentration may be an artifact of sample hemolysis or may be due to dehydration or hemoconcentration.

 Hypoalbuminemia may be due to decreased synthesis (chronic liver disease, dietary protein deficiency or chronic inflammation), increased loss (renal disease, intestinal parasitism or gastrointestinal disease), or sequestration (decreased oncotic pressure or increased hydrostatic pressure). Decreases can also occur with blood loss, severe inanition and chronic infection. Pre-albumin is a separate and distinct plasma protein fraction that precedes albumin on electrophoresis, and is involved in the transportation of  thyroid hormones.

The globulins are composed of three fractions, designated alpha, beta and gamma. In birds, one or two subfractions of alpha globulins have been identified as well as a single fraction for each of the beta and gamma globulins.

Alpha globulins are a group of proteins manufactured almost entirely by the liver. As concentrations increase during inflammation, their measurement is helpful in the diagnosis and monitoring of many infectious diseases as well as other causes of acute and chronic inflammation. Alpha globulins increase with acute nephritis, severe hepatitis, active systemic inflammation, malnutrition and in the nephrotic syndrome. Decreases can occur with hepatic insufficiency, severe inanition, blood loss and protein-losing gastrointestinal (GI) diseases. Beta globulins include carrier proteins, complement, ferritin, C-reactive protein, lipoproteins and fibrinogen, and many are also acute phase proteins. Increases in beta globulins occur with acute inflammation, hepatitis, malnutrition, lipemia artifact, systemic mycotic disease, and the nephrotic syndrome. Decreases occur with hepatic insufficiency, severe inanition, blood loss and protein-losing enteropathies.

The gamma fraction in birds contains most of the immunoproteins, including IgA, IgE, IgM and IgG. In some species, nonimmunoglobulin proteins, including transferrin, complement and fibrinogen, are found within the gamma globulin fraction.

Gamma globulins increase with ongoing antigenic stimulation, often from infectious agents. Broad increases (polyclonal gammopathies) in gamma globulins occur with acute or chronic inflammation, infection, chronic hepatitis and immune-mediated disorders. Sharp increases (monoclonal gammopathies) occur with tumors of the reticuloendothelial system and plasma cell dyscrasias. Deficiencies can occur with immunodeficiency states, blood loss, overwhelming infection, protein-losing GI diseases and severe inanition. The half-life of gamma globulins in birds is relatively short.

Panhypoproteinemia, which occurs when both albumin and globulins are decreased, may occur with severe inanition or malnutrition, severe hepatic insufficiency, overwhelming infection, protein-losing states (especially GI or kidney), external blood loss, or blood loss into the body cavity.

There are some instances when the electrophoretic pattern may be difficult to interpret. Avian neonates and juveniles likely show age-related differences in serum proteins, but this has not been fully investigated. Thus, caution is advised when interpreting electrophoretic patterns of young birds. In some psittacine species, a monoclonal spike may be present in the beta fraction during periods of egg-laying, when there is transport of egg proteins to the ovary. This increase in an egg-laying hen has been attributed to transferrin, and to increases in estrogen-induced yolk protein precursors, vitellogenin and lipoproteins. In raptors, there are normal variations in electrophoresis patterns that differ from those found in psittacines.

A unique electrophoretic pattern has been reported during acute chlamydiosis. Typically, there is a moderate to severe hypoalbuminemia, mild to moderate elevation in beta globulins,and moderate to severe hypergammaglobulinemia. In chronic chlamydiosis, there may be no changes in the inflammatory proteins, or only a mild hyperbetaglobulinemia. In mycotic diseases, especially aspergillosis, an increase in the beta fraction is seen during the acute phase. There may be beta or gamma globulin elevations during the chronic phase of aspergillosis. Eventually, birds chronically infected with aspergillus may lose the inflammatory protein response altogether, becoming hypoproteinemic. Mycobacterial infections may show increases in either beta or gamma globulins, whereas sarcocystis infections usually show an elevation in both beta and gamma fractions. Birds with hepatitis or nephritis often show a decrease in albumin and increases in beta globulins. Gross hemolysis may show a severely restricted (usually gamma) spike composed of hemoproteins that can be misinterpreted as being clinically important.

This newsletter was contributed by Marge Wissman DVM, Dipl ABVP (Avian Practice) and Robin Lane DVM, Dipl ABVP (Avian Practice).

RECHECK PROFILES FOR AVIANS AND EXOTICS

To facilitate patient follow-up, Antech Diagnostics now offers recheck profiles on avians, exotic mammals, and reptiles at considerable savings.

Recheck profile pricing affords ~ 37% savings from the original comprehensive profile price. To qualify for the reduced price, recheck profiles must arrive at the lab within 30 days of the original profile, and the accession number for the original profile needs to be written on the Test Request Form.

Recheck avian comprehensive profile is listed on the current A & E Test Request Form (see under "Avian Profiles"). Recheck profiles for small exotic mammals and reptiles are not listed on the Test Request Form and need to be entered, along with the accession number for the original comprehensive profile, in the "Custom Panel" space in the top left corner of the form. Further questions can be directed to our specially trained Avian and Exotics Customer Service Representatives or our Veterinary Consultants.