INTRODUCTION TO AUTOIMMUNITY AND AUTOANTIBODY TEST

1. INTRODUCTION TO AUTOIMMUNITY AND AUTOANTIBODY TEST By MadihaHamid Horror autotoxicus: Literally, the horror of self-toxicity. • Tolerance to self Ags is maintained by central and peripheral mechanisms. • Dysregulation in these mechanisms will trigger autoimmune disease. • A family of 80 chronic and disabling diseases • Affects about 15-23 million people in the USA.

2. Autoimmunity Genetic Factors CAUSES • Three main sets of genes are suspected in many autoimmune diseases. These genes are related to: • Immunoglobulins • T-cell receptors • The major histocompatibility complexes (MHC). Autoimmunity Environmental Factors • Following factors can trigger autoimmunity: • Pathogens, • drugs, • hormones, • toxins Symptoms • Common symptoms may be Fever, muscle ache, fatigue, joint pain. • Each disease has specific symptom

3. Autoimmunity Can be classified into clusters that are either organ-specific or systemic Lungs of a patient with Goodpasture’s

4. Autoantibody test Mainly done for the diagnosis of autoimmune disease. The Antinuclear antibody (ANA) test is often ordered first. ANA is a marker of the autoimmune process – it is positive with a variety of different autoimmune diseases but not specific. If an ANA test is positive, a panel of 4 or 6 autoantibody tests called extractable nuclear antigens (ENA) is typically ordered. , it is often followed up with other tests associated with arthritis and inflammation, such as a rheumatoid factor (RF), an erythrocyte sedimentation rate (ESR), a C-Reactive Protein (CRP), and/or complement protein complement levels. The AMA test is ordered to help diagnose primary biliary cirrhosis (PBC).These abs are detedtced by indirect immunoflourescence in > 90% of patients.Reference range is <0.1 -1.0 units. APA testing is used to help determine the cause of : Inappropriate blood clot formation (unexplained thrombotic episode, excessive clotting) Recurrent miscarriage Low platelet count (thrombocytopenia) Prolonged PTT test A cyclic citrullinated peptide (CCP) antibody test may be ordered along with or following a rheumatoid factor (RF) test to help diagnose rheumatoid arthritis (RA) and to assess the severity and probable course of the disease (prognosis).

5. C-Reactive Protein Complement Levels GulSanober

6. C-reactive protein (CRP) test • a non-specific marker produced by the liver • increases during episodes of acute systemic inflammation • measured by blood tests. • Some forms of arthritis • Autoimmune diseases, such as lupus or vasculitis • a CRP level of 10 mg/L or lower is considered "normal."

7. Complement Levels • most commonly C3 and C4 • Synthesized in the liver • Measured through blood tests • Normal levels C3 • Males: 88-252 mg/dL • Females: 88-206 mg/Dl C4 • Males: 12-72 mg/dL • Females: 13-75 mg/dL • Levels decrease in autoimmune disorders(Hypocomplementemia)

8. Diagnose and monitor: •  acute or chronic autoimmune diseases such assystemic lupus erythematosus (SLE) • immune complex-related diseases and conditions such as glomerulonephritis, serum sickness, rheumatoid arthritis, and vasculitis • Individual complement components ordered when the total complement activity (CH50 or CH100) is abnormal to help determine which of the component(C1 to C9)s are deficient or abnormal

9. Considerations • Serum is preferred •  All samples must show no signs of deterioration and lipaemic samples should be avoided. • Samples may be stored at 4°C prior to analysis • Acute malarial infection can cause false-positive results • Complement component 3 has been shown to interact with Factor H.

10. Rheumatoid factor - RF By: Namra Haq

11. Rheumatoid factor explained Rheumatoid factor is an immunoglobulin (antibody) which can bind to other antibodies. Antibodies are normal proteins found in the blood which function within the immune system. Rheumatoid factor though is not normally found in the general population (only found in about 1-2% of healthy people). The incidence of rheumatoid factor increases with age and about 20% of people over 65 years old have an elevated rheumatoid factor. A blood test is used to detect the presence of rheumatoid factor. The blood test is commonly ordered to diagnose rheumatoid arthritis. Rheumatoid factor is present in 80% of adults who have rheumatoid arthritis but there is a much lower prevalence in juvenile rheumatoid arthritis. The incidence of rheumatoid factor increases with duration of disease in rheumatoid arthritis: at 3 months the incidence is 33%, while at one year it is 75%. Up to 20% of rheumatoid arthritis patients remain negative for rheumatoid factor (also known as "seronegative rheumatoid arthritis") throughout the course of their disease.

12. Other conditions in which RH is positive apart from rheumatoid arthritis Other autoimmune diseases can also be positive rheumatoid factor including: Sjogren’s Syndrome, Systemic Lupus Erythematosus, Scleroderma, Polymyositis, Dermatomyositis, Mixed Connective Tissue Disease Other infections or conditions which can be associated with positive rheumatoid factor include: Bacterial Endocarditis, Osteomyelitis, Tuberculosis, Syphilis, Hepatitis, Mononucleosis, Liver Cirrhosis, etc. Conditions not associated with RF Rheumatic conditions NOT associated with elevated rheumatoid factor include: Osteoarthritis, Gout, Reiter's syndrome / Reactive arthritis, etc.

13. How is RF factor measured? The amount of rheumatoid factor in blood can be measured by: Agglutination tests One method mixes the patient's blood with tiny latex beads covered with human antibodies (IgG). The latex beads clump or agglutinate if rheumatoid factor (IgM RF) is present. Another method mixes the patient's blood with sheep red blood cells that have been covered with rabbit antibodies. The red blood cells clump if rheumatoid factor is present. A titer is an indicator of how much the agglutination test blood sample can be diluted before rheumatoid factor is undetectable. A titer of 1:20 indicates that rheumatoid factor can be detected when 1 part of blood is diluted by up to 20 parts saline. The lab value for rheumatoid factor of 1:20 or less is considered normal.

14. Nephlometry test • This method mixes the patient's blood with antibodies that cause the blood to clump if rheumatoid factor is present. A light is passed through the tube containing the mixture and an instrument measures how much light is blocked by the mixture. Higher levels of rheumatoid factor create a more cloudy sample and allow less light to pass through, measured in units. The lab value for rheumatoid factor of 23 or less units is considered normal. • When analyzing lab results the following should be considered: • A rheumatoid factor more than 23 units and a titer more than 1:80 is indicative of rheumatoid arthritis but may also occur in other conditions. • False positive results can occur when the blood is high in fats. • Inaccurate results can be caused by improper handling of the blood specimen. • A negative test result for rheumatoid factor does not exclude the diagnosis of rheumatoid arthritis.

15. Erythrocyte Sedimentation Rate Test (ESR / Sed Rate) • Marker of Inflammation-autoimmune diseases like lupus or rheumatoid arthritis cause inflammation. • Nonspecific Test • Not used for screening purpose (low senstivity and specificity) • Inexpensive • Uses: a. Diagnosis(auto-immune diseases ,rheumatoid arthritis, chronic kidney diseases) b. Disease severity c. Monitoring response to therapy ( inflammatory diseases such as temporal arteritis, polymyalgia rheumatica and rheumatoid arthritis)

16. Erythrocyte Sedimentation Rate Test (ESR / Sed Rate) • Measure of the settling of red blood cells in a tube of blood during one hour. • When abnormal proteins are present in the blood, typically due to inflammation or infection, they cause red blood cells to clump together and sink more quickly.

17. Erythrocyte Sedimentation Rate Test (ESR / Sed Rate) Autoimmune Disorders Higher than Normal • Factors That May Influence ESR • Increase ESR • Old Age • Pregnancy • Anemia • Macrocytosis • Decreased ESR • Extreme leukocytosis • Polycythemia • Red blood cell abnormalities • Very high ESR levels • Allergic vasculitis • Giant cell arteritis • Polymyalgia rheumatica Reference Age Adults Upper limit of reference range (mm/hr) Age < 50 years Men 0 to 15 Women 0 to 20 Age > 50 years Men 0 to 20 Women 0 to 30

18. Secret Blood Test of Autoimmune Disease TH1 / TH2 Cytokine Test The immune system works like a seesaw. On one side you have Th1 cytokines that initiate the first line of defense. On the other side you have Th2 cytokines which help product antibodies to protect you from future invasions. However, when one side goes up, the other side goes down. This can contribute to a weak immune system and potentially promote autoimmune conditions. 

19. Complete blood count in primary care

20. PRINCIPLE • This test measures red blood cells and white cells. The test includes: red blood cell count, hemoglobin, hematocrit, MCV, MCHC, RDW, platelet count, white blood cell count, neutrophils, lymphocytes, monocytes, eosinophils and basophils.

21. Phisiology: • Whole blood collected in EDTA (purple top tube) is the only acceptable specimen. The • specimen must be kept and transported at room temperature. For best results, the • collection tube should be full and no less than half full. Mix the specimen slowly for 2 • minutes. • Unacceptable specimens include: • Any sample not collected in an EDTA tube. • Any tube less than half full • Any tube that is not labeled properly • Hemolyzed specimens • Specimens older than 24 hours • Specimens that are clotted METHOD: VCS Technology, Hemoglobinometry

22. Significance: • Screen for a wide range of conditions and diseases • Help diagnose various conditions, such as anemia, infection, inflammation, bleeding disorder or leukemia, to name just a few • Monitor the condition and/or effectiveness of treatment after a diagnosis is established • Monitor treatment that is known to affect blood cells, such as chemotherapy or radiation therapy • A CBC is a panel of tests that evaluates the three types of cells that circulate in the blood and includes the following: • Evaluation of white blood cells, the cells that are part of the body's defense system against infections and cancer and also play a role in allergies and inflammation • Evaluation of red blood cells, the cells that transport oxygen throughout the body • Evaluation of platelets, cell fragments that are vital for normal blood clotting

23. ULTRASOUND* • >20,000 Hertz OR more • Diagnostic Medical Ultrasound -use of high frequency sound waves to aid in the diagnosis and treatment • Frequency ranges -> 2 - 15 MHz • Piezoelectric Effect : The principle of converting energy by applying pressure to a crystal • Pulse-echo principle, ultrasound transducers; [convert 1 type of energy to another] – Electricity into sound = pulse {machine to tissue} – Sound into electricity = echo {tissue to machine} • Echoes are interpreted and processed by the ultrasound machine

24. X-RAYS* • Digital Radiography, High Frequency Radiography Unit help in diagnosis • Electro magnetic radiations {ELECTRONS} • High-speed flow of electrons bombarding the anode target surface, 99% converted to heat, to produce X-ray - 1% ; {(50Hz or 60Hz)} • Useful in detection of pathology of the skeletal system • Bones contain much calcium hence absorb x-rays efficiently ; reduces amount of X-rays reaching detector in the shadow of the bones {making them clearly visible on the radiograph} • In medical diagnostic applications, low energy (soft) X-rays are unwanted {since they are totally absorbed by the body, increasing the radiation dose without contributing to the image} • A thin metal sheet, often of “aluminium”, [an X-ray filter], usually placed over the window of the X-ray tube, absorbing the low energy rays. Center of the spectrum shifts toward higher energy {hardening the beam ; Hard X-rays}

25. BIOPSY* • Medical test performed by surgeon involving sampling of cells or tissues for examination. [TISSUE SAMPLING] • Medical removal of tissue from a living subject to determine the presence or extent of a disease. • Frequent biopsies are taken to assess activity of disease and to assess changes that precede malignancy. • Examined under a microscope or analyzed chemically by a pathologist. • Entire lump or suspicious area is removed {Excisional biopsy}. • Sample of tissue is removed with preservation of the histological architecture of the tissue’s cells {Incisional biopsy or core biopsy}. • Scanner's images help doctors determine exact position of the needle in the targeted tissue{CT-guided biopsy}. • An ultrasound scanner helps a doctor direct the needle into the lesion.{Ultrasound-guided biopsy}. • A needle withdraws material out of a mass. {Aspiration biopsy}.

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