Plasma protein tests

Definition

Plasma protein tests are laboratory tests used to evaluate the levels of specific proteins in the blood. A decrease or increase in the concentration of the protein is associated with one or more clinical conditions. Prior to measuring a specific protein, a comprehensive metabolic profile is usually performed. This profile includes tests for the total protein and albumin. If either of these tests is abnormal, serum protein electrophoresis may be performed in order to determine the cause. Abnormalities seen on the protein electrophoresis test as well as other clinical information are used to determine the necessity for specific protein tests. The most frequently measured plasma proteins include alpha-1 antitrypsin (AAT), ceruloplasmin (CER), C3 and C4 (complement proteins), Creactive protein (CRP), haptoglobin (Hp, HAP), immunoglobulins (IgG, IgA, IgM), transferrin (TRF), and transthyretin (TTR).

Purpose

Plasma protein tests are used to determine whether a specific protein or proteins have increased or decreased in concentration. An abnormal level of total protein or of a specific protein indicates particular diseases or conditions associated with the respective change. Some protein tests are useful for establishing a diagnosis, while others are useful in determining the extent of a condition such as dehydration or inflammation.

Precautions

A nurse or phlebotomist who collects the sample for a plasma protein test should follow standard precautions for the prevention of transmission of bloodborne pathogens. Total protein and albumin concentrations are approximately 10% higher in ambulatory persons. Plasma total protein is approximately 0.2 - 0.4 g/dL higher than serum. Some drugs, especially estrogens and corticosteroids , may increase the concentration of specific proteins.

Description

Proteins are vital to the proper functioning of the body. There are more than 3,000 proteins found in a single human cell. Some proteins, such as enzymes, hormones, coagulation factors, lipoproteins, and hemoglobin, are measured routinely and are described in other topics. The proteins considered here are those which are sufficient in concentration to affect results of the total protein or protein electrophoresis tests or are measured by immunonephelometry. Each of the proteins described below, with the exception of total protein and albumin, are measured by this method. In immunonephelometry, plasma or serum is added to a buffer containing an excess of a specific antibody (e.g., anti-haptoglobin). The anti-bodies will bind to the haptoglobin molecules (antigen), forming small soluble immune complexes. Monochromatic light (usually 450-550 nm) passing through the reaction tube will be scattered by the immune complexes as they form. Forward-angle light scatter is measured by a light detector (photomultiplier tube) placed at an angle (usually 70 degrees) to the incident light. The combination of antibody and antigen molecules occurs rapidly, and the photodector current increases with time until a peak signal is reached. This peak is proportional to antigen concentration. To insure that the antibody molecules are in excess, an addition of reagent antigen is injected into the reaction mixture after the peak rate is obtained. This addition will produce an increase in light scattering provided that sufficient antibody remains. The clinical significance of each protein test is described below.

Total protein: The total protein of plasma or serum is measured by a colorimetric reaction called the biuret method. The sample is added to an alkaline solution of copper II sulfate. The copper ions form coordinate bonds with the carbonyl and imine groups of the protein. This causes the reagent to change from a sky blue to a purple color. The absorbance of the solution at 540 nm is proportional to protein concentration.

Total protein is increased in conditions causing dehydration. They includes vomiting, diarrhea , diabetes insipidus, diabetes mellitus , and Addison's disease. Total protein is increased in conditions that cause inflammation. These include cancer , autoimmune diseases, and chronic or severe infections. Total protein is also increased by monoclonal immunoglobulin production caused by benign or malignant proliferation of antibodysecreting cells (plasma cells) such as multiple myeloma. Low total protein can result from protein loss, as occurs in hemorrhage, glomerulonephritis, nephrosis, proteinlosing gastroenteropathy, and burns ; excessive hydration as occurs in salt retention syndromes and the syndrome of inappropriate antidiuretic hormone (SIADH); or decreased synthesis resulting from starvation and chronic liver disease.

Albumin: The albumin in plasma comprises 50-65% of the total protein. In addition to holding water in the vascular bed (maintenance of oncotic pressure) albumin is primarily a transport protein and is responsible for the protein binding of most drugs. It is measured by a dyebinding method using either bromcresol green (BCG) or bromcresol purple (BCP). These dyes selectively bind to albumin, forming a colored complex. The color formed is proportional to albumin concentration. Plasma albumin is increased by dehydration or intravenous infusion of albumin as a blood volume expander. It is decreased in hemorrhage, renal disease, salt retention, SIADH, liver disease, starvation, inflammation, malignancy, and infection .

Alpha-1 antitrypsin: ATT is a glycoprotein (molecular weight 52,000) made by the liver. ATT is an inhibitor of the enzyme trypsin as well as of other proteolytic enzymes (serine proteases) that are released from phagocytic white blood cells during inflammation. ATT reduces the damage to tissues caused by the immune response , and plasma levels increase up to two-fold in acute and chronic inflammatory conditions. Proteins such as ATT that are increased in response to inflammation are called acute phase proteins. They are sensitive markers for tissue injury, myocardial infarction , infection, malignancy, and autoimmune diseases.

A deficiency of AAT results in excessive inflammation in tissues that are exposed to bacteria and other foreign cells, commonly the lungs and gastrointestinal organs. AAT deficiency is usually inherited. Expression of the genes for AAT is codominant. The most common normal phenotype is MM. The phenotype ZZ (homozygous for the Z variant) produces only about 10% of normal activity and is most commonly associated with immunological damage. The most frequent complications involve the lungs (emphysema ) and the hepatobiliary tract (hepatitis and cirrhosis). A deficiency of AAT is suspected when the alpha-1 band of serum protein electrophoresis is absent or below 1% of the total protein. The AAT concentration is measured by immunonephelometry and the variant proteins can be identified by high-resolution gel electrophoresis.

Ceruloplasmin: CER is a protein (molecular weight 120,000) produced by the liver. It is an acute phase protein, and increased CER may contribute slightly to the size of the alpha-2 band on electrophoresis. Ceruloplasmin is measured as an aid to the diagnosis of Wilson's disease. Wilson's disease is an autosomal recessive disease in which the binding of copper by CER and the excretion of copper into the bile are impaired. Copper accumulates in the tissues, principally the liver, central nervous system , and eyes. Deposition in the eyes produces Kayser-Fleischer rings (green to brown rings around the edge of the cornea), a classic sign on physical exam. Copper deposition damages tissues, causing cirrhosis of the liver and damage to the lenticular area of the brain (hepatolenticular degeneration). It may also cause osteoporosis , renal, joint, cardiac, and other damage. Plasma levels below half the lower limit of normal and low plasma copper are suggestive but are not conclusive in the absence of clinical signs. CER is measured by immunonephelometry.

C3 and C4: C3 and C4 are glycoproteins that act along with other complement proteins to facilitate lysis of antibody-coated cells. The complement system consists of nine proteins in the classical pathway and an additional five that act in the alternative pathway. Deficiencies of 10 complement proteins have been described. A deficiency of C3 or C4 is associated with systemic lupus erythematosus (SLE) and other autoimmune diseases. The deficiency of either complement component may be responsible in part for the development of the disease by preventing the effective removal of immune complexes. C3 and C4 deficiency are also associated with severe recurrent infections. In active SLE, glomerulonephritis, cirrhosis, and sepsis C3, C4 and complement activity may be reduced due to consumption by immune complex formation. In rheumatoid arthritis, rheumatic fever , and some chronic dermatologic

KEY TERMS

Edematous —The state of having swelling (edema) caused by the collection of excess fluid within tissues.

Hematoma —Swelling and subsequent bruising when blood leaks from a vein into local tissues; can be caused by improper venipuncture when the needle has gone through a vein or when the needle has been inserted incorrectly.

Hemolytic —Destructive to red blood cells.

Venipuncture —Puncture of a vein with a needle for the purpose of withdrawing a blood sample for analysis.

Wilson's disease —A genetic disorder that leads to excessive amounts of copper in the body. Patients often have damaging deposits of copper in the brain, liver, and other organs, and a green discoloration from copper around the iris of the eyes. This disease is also called hepatolenticular degeneration.

diseases, C3 and C4 levels are elevated owing to increased complement activation.

C-reactive protein (CRP): C-reactive protein is a protein consisting of five subunits (molecular weight 120,000) produced mainly in the liver. Its name is derived from the fact that it binds to the C-polysaccharide of the capsule of Streptococcus pneumoniae. Increased levels are seen in patients with pneumococcal pneumonia as well as other acute infections and inflammatory conditions. In the absence of inflammation, CRP levels in plasma are very low (< 4 mg/L). Levels in inflammation can reach several hundred-fold above normal, causing a small but distinct band in the gamma zone on electrophoresis. Recent studies have shown that a CRP near the upper limit of normal (5-10 mg/L) in persons with a history of chronic inflammation is a risk factor for coronary artery disease . A new test, called high-sensitivity CRP, can measure CRP levels below 1 mg/L; and is being used by some cardiologists to predict the risk of coronary artery disease in persons with normal total cholesterol who have no history of angina or heart disease. CRP is measured by immunonephelometry or enzyme immunoassay.

Haptoglobin: Hp is a glycoprotein (molecular weight 85,000-100,000) made by the liver. Hp binds to free plasma hemoglobin, transporting it to the liver, where the complex is removed by the reticuloendothelial cells. Low levels are seen in persons with intravascular hemolysis (e.g., following an intravascular transfusion reaction). Haptoglobin is also an acute phase protein. Hp and alpha-2 macroglobulin are responsible for the increased density of the alpha-2 band on electrophoresis seen in acute and chronic inflammatory states. Hp is measured by immunonephelometry.

Immunoglobulins G, A and M (IgG, IgA, and IgM): Immunoglobulins are antibodies produced by B lymphocytes and secreted by plasma cells. They will be increased in response to infections, malignancy, and autoimmune diseases—all of which produce a polyclonal response (i.e., a general increase in all three immunoglobulin classes). This response causes a diffuse increase in the density of the gamma zone on electrophoresis. In malignant or benign plasma cell disorders, proliferation of a single clone of plasma cells results in the accumulation of identical antibody molecules called a monoclonal gammopathy. This result may be recognized on electrophoresis as an area of restricted mobility in the gamma zone. Decreased plasma concentration of one or more immunoglobulin classes may be associated with immunologic impairment and result in both recurrent and opportunistic infections. Both increases and decreases of each immunoglobulin class can be detected by immunonephelometry, using antibodies specific for each. For example, rabbit anti-human IgG can be used to measure the plasma concentration of IgG.

Transferrin: TRF is a glycoprotein (molecular weight 77,000) made by the liver. It is responsible for transport of iron from the gut to the bone marrow. The concentration of transferrin in the plasma is directly related to the total iron binding capacity (TIBC). In iron deficiency, the transferrin level is increased, causing an increase in the density of the beta globulin band on electrophoresis. Transferrin levels are decreased in nephrosis, liver disease, starvation, and chronic illness. Like albumin, transferrin is reduced in acute and chronic inflammation and is referred to as a negative acute phase protein.

Transthyretin: Transthyretin or prealbumin is a glycoprotein (molecular weight 54,000) made in the liver. TTR has a very short half-life, making it a useful marker for protein calorie malnutrition. In persons with deficient protein intake, the plasma level of TTR decreases before those of most other proteins. The level of TTR can be monitored to assess the efficacy of dietary intervention in malnourished persons. TTR is measured by immunonephelometry.

Preparation

Prior to performing the venipuncture, the nurse or other health care professional should document any medications the patient is currently taking, and any medical conditions that could influence the protein tests. For example, oral contraceptives, estrogen-containing drugs, or pregnancy can increase the level of ceruloplasmin.

Aftercare

The patient may feel discomfort when blood is drawn from a vein. Bruising may occur at the puncture site, or the person may feel dizzy or faint. Pressure should be applied to the puncture site until the bleeding stops, to reduce bruising. Warm packs can also be placed over the puncture site to relieve discomfort.

Complications

In normal circumstances, a blood draw for protein tests takes only a few minutes, and the patient experiences only minor discomfort from the puncture.

Results

The physician will carefully consider the results from the specific protein test within the context of the patient's current health status to make decisions on further testing, diagnosis, and treatment. The protein test results must be interpreted by comparison with standard reference ranges provided by the laboratory that has performed the test. The normal ranges shown below are frequently cited for the methods described previously.

• total protein: 6.0-8.0 g/dL nonambulatory; 6.5-8.5 g/dL ambulatory
• albumin: 3.0-5.0 g/dL nonambulatory; 3.5-5.5 g/dL ambulatory
• alpha-1 antitrypsin: 90-200 mg/dL
• ceruloplasmin: 20-60 mg/dL
• C-reactive protein: 0.7-8.2 mg/L
• Hs-CPR: 0.08-3.1 mg/L
• C3: 90-180 mg/dL
• C4: 10-40 mg/dL
• haptoglobin: 30 - 200 mg/dL
• immunoglobulins: IgG: 700-1600 mg/dL; IgA: 70-400 mg/dL; IgM: 40-230 mg/dL
• transferrin: 200-360 mg/dL
• transthyretin: 20-40 mg/dL

Health care team roles

In accordance with the physician's orders, the nurse, phlebotomist, or laboratory professional usually prepares the patient, performs the blood draw, and readies the specimen for transport to the laboratory for analysis. A clinical laboratory scientist, CLS(NCA)/medical technologist, MT(ASCP) or clinical laboratory technician CLT(NCA)/medical laboratory technician MLT(ASCP) performs the testing. Results are interpreted by a physician.

Resources

BOOKS

Fischbach, Frances. "Diagnostic Testing." In A Manual of Laboratory & Diagnostic Tests, 6th ed. Philadelphia: Lippincott Wiiliams & Wilkins, 2000, pp. 1-25.

Johnson, A. Myron, Elizabeth M. Rohlfs, and Lawrence M. Silverman. "Proteins." In Tietz Textbook of Clinical Chemistry, 3rd ed., edited by Carl A. Burtis and Edward R. Ashwood. Philadelphia: W. B. Saunders Company, 1999, pp. 478-523.

Kee, Joyce LeFever. Laboratory & Diagnostic Tests with Nursing Implications, 5th ed. Stamford, CT: Appleton & Lange, 1999, pp. xv-xix, 27-29, 111-112, 214-215, 269-271, 362-364.

Sacher, Ronald A., Richard A. McPherson, with Joseph M. Campos. "Special Tests in Evaluation of Liver Disease." In Widmann's Clinical Interpretation of Laboratory Tests, 11th ed. Philadelphia: F. A. Davis Company, 2000, pp. 586-593.

ORGANIZATIONS

The American Society for Clinical Laboratory Science. 7910 Woodmont Ave., Suite 523, Bethesda, MD 20814. (301) 657-2768. <http://www.ascls.org>.

Wilson's Disease Association. 4 Navaho Dr., Brookfield, CT 06804. (800) 399-0266. <http://www.wilsonsdisease.org>.

OTHER

"Normal Laboratory Values, Table 296-2." In The Merck Manual of Diagnosis and Therapy. 17th ed., internet edition, <http://www.merck.com/pubs/mmanual>, 1999.

Linda D. Jones, B.A., PBT (ASCP)