Cryoglobulins Classification and Diagnosis

Cryoglobulins Classification and diagnosis

Sample for cryoglobulins

1. Venous blood is needed.
2. Keep the blood at 37 °C.
3. Separate the serum at 37 °C,  so the cryoglobulins remain in the serum.

Precautions for detecting the cryoglobulins

1. Maintain the temperature of the blood at 37 °C.
2. Also, maintain the temperature of the serum at 37 °C.

Purpose of the test (Indications) for detecting the cryoglobulins

1. This test is done to diagnose:
   1. Purpura.
   2. Arthralgia.
   3. Raynaud phenomenon.

Pathophysiology of Cryoglobulins

Definition:

1. These immunoglobulins precipitate in serum reversibly at low temperatures or may gel at cold temperatures.
2. Most cryoglobulins are polyclonal immunoglobulin complexes, and nearly half are monoclonal, usually IgM.
3. Cryoglobulins are abnormal immunoglobulins proteins produced due to various diseases.
4. Cryoglobulins:
   1. These are insoluble at 4 °C.
   2. May aggregate up to 30 °C.
   3. It can fix complement.
   4. It can initiate an inflammatory response.

Cryoglobulinemia is defined as a condition in which plasma or serum proteins (IgM) precipitate when cooled below body temperature.

1. These proteins precipitate at 4 °C and dissolve again at 37 °C.
2. These are the proteins that reversibly precipitate or gel at 4 °C.
   

   Cryoglobulins main features

These cryoglobulins are classified as follows:

1. Type I (Monoclonal).
2. Type II (Mixed cryoglobulin, a mixture of polyclonal immunoglobulins).
3. Type III (Polyclonal).
   1. Primary or idiopathic or essential.
   2. Secondary that is associated with diseases.
4. Type I monoclonal cryoglobulins are produced by the neoplastic lymphocytes and plasma cells. These are typically IgM types of immunoglobulin.
   1. May see IgG as well.
   2. Rarely see IgA only.
   3. These may constitute 25% of the cryoglobulins.
   4. Often present >5 mg/dL in the serum.
   5. Symptoms are severe, and often there may be gangrene without any other cause.
   6. Associated with multiple myeloma, macroglobulinemia, and rare neoplasm of the plasma cells and lymphocytes.
5. Type II cryoglobulins consist of two types:
   1. The monoclonal form is seen as the rheumatoid factor and usually is IgM with a light kappa chain.
   2. The second is polyclonal IgG which is always with RF.
   3. It is seen in 25% of the cases.
   4. It is most often seen with HCV infection and less often with HBV, EBV, and other diseases.
6. Type  III  cryoglobulins are mixed cryoglobulins and the most common combination of IgG and IgM (usually with RF).
   1. It is seen in ∼50% of the cases.
   2. It is usually present in a small amount of <1 mg/dL.
   3. More than 90% contains IgM, rheumatoid factor, and IgG.
   4. Mostly seen in autoimmune diseases.
      1. There are circulating immune complexes produced by various antigens like viral, bacterial, and autologous antigens.
7. Mixed cryoglobulinemia consists of type II and type III.

Signs and symptoms:

1. Cryoglobulinemia is a systemic immune-complex disease characterized by clinical syndrome-like:
   1. Palpable purpura. This is a constant feature and is present in the lower extremities.
   2. Arthralgias.
   3. Hepatosplenomegaly.
   4. Lymphadenopathy.
   5. Anemia.
   6. Glomerulonephritis. The most common is membranoproliferative glomerulonephritis ( MPGN type II).
   7. Most patients are hypocomplementemic, reflecting immune-complex disease.
2. These proteins precipitate in the blood vessels of fingers when exposed to cold.
3. These patients will have symptoms of:
   1. Vascular purpura. This is the most common symptom and is seen in 60% to 100% of the patients.
   2. Bleeding tendency.
   3. Arthralgia. It is the second common, most seen in 60% to 90% of the cases.
   4. Cold-induced urticaria.
   5. Raynaud phenomenon (pain, cyanosis, the coldness of fingers). It is seen in 50% of the cases.
4. These proteins are present in various diseases.
   1. Serum level > 5 mg/dL = Multiple myeloma, macroglobulinemia, and leukemia.
   2. Serum level between 1 to 5 mg/dL= Rheumatoid arthritis.
   3. Serum level < 1 mg/dL = SLE, RA, infectious mononucleosis, viral hepatitis, endocarditis, cirrhosis, glomerulonephritis.

Type of cryoglobulin	diseases
Type I	Multiple myelomas Macroglobulinemia Monoclonal gammopathy.
Type II	Vasculitis Glomerulonephritis SLE RA Sjogren’s syndrome Hepatitis CMV
Type III	May see diseases like type II and autoimmune diseases.

Normal Cryoglobulins

• The serum is negative for the cryoglobulins.

Procedure

1. Take the blood and keep it at 37 °C until it is clotted.
2. Clotted blood is centrifuged at 37 °C.
3. Keep the serum in the fridge at 4 °C.
   1. Please keep it for at least 3 days and again centrifuge at 4 °C.
4. Check the serum daily for white precipitate or gel.
   1. Checking the time period is disputed; some suggest 3 days, while others propose 7 days.
5. Check daily for at least 72 hours to 7 days.
   1. The precipitate is recorded and measured, which is cryoglobulin.
   2. Rewarm the sample if it dissolves, then it confirms the cryoglobulins.
6. Cryoprecipitate is washed and subject to Immunoelectrophoresis to identify the type of cryoglobulin (immunoglobulin).
7. Another procedure is:
   1. Collect the blood in EDTA or citrate and also make a serum sample.
   2. Incubate both at 4 °C.
   3. A precipitate in the plasma but not in the serum will indicate cryoglobulins.

Causes of Cryoglobulins

1. Autoimmune diseases like SLE, RA, Sjogren’s syndrome.
2. Malignancies like Multiple myeloma, Leukemia, Waldenstrom macroglobulinemia, and Lymphoma.
3. Acute and chronic infections like Infectious mononucleosis, Poststreptococcal glomerulonephritis, and endocarditis.
4. Liver diseases like Cirrhosis and Hepatitis.
5. Renal involvement manifests as either nephrotic or nephritic syndrome associated with hypocomplementemia.
   1. The most common finding is membranoproliferative glomerulonephritis type II.

Treatment of cryoglobulins

1. It is attempted on immune-modulation using corticosteroids.
2. Can try plasmapheresis.
3. α-interferon in patients with HCV.
4. Remissions are achieved in 75% of the cases.
5. Relapse is seen in 50% of the cases.

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