Serology refers to considering serum and fluid of the body. In this part, antibodies are detected. So, in order to detect an infected diseases, microbes are separated from the patient. Factors such as, age of the patient, using medication, stage of disease and so on affects on result of the tests.
Separating microbe from patient. But due to using anti- bacterial drugs or problems related to separating and cultivating the microbe, this process will be difficult, and makes the diagnosis problematic. in such ways, the best way is to finding specific anti- bacterial antibodies in patient’s serum.
Now, serology, is one of the most simple and fast methods of diagnosis diseases. This test is so important, due to changes that occur in titr of the antibodies during the disease, and doctors can control any changes in your body. In other word, decreasing titr of the antibody, is an indication of proper treatment.
When result of test is not match with clinical condition of the patient, this test should be done 2 times, with 2 weeks intervals to compare titr of the antibody.
Following factors affect on interpretation of the test:
- Patient’s age and background
- Immune system of the patients
- Patient’s job
- Time of vaccine
- Converge of the antigen
- Using drugs
- Stage of the disease
Tests which are done in this part are as:
CRP (Latex agglutination)
A. Factor (RF latex)
Widal (agglutination test)
RPR (Rapid plasma reagin)
PPD Skin Test
Types of Serological Reactions
Sedimentary reactions or perspiration: In these reactions, antigens are soluble molecules. The soluble antigen is a periphytonogen, an antibody against it is called persipitin, and this reaction is called perspiration.
Agglutination reactions: If the antigen is either insoluble or particle, the antigen is called agglutinogen, an antibody is agglutinin, and the reaction is called agglutination. If the anti-insulin antigen is red, the reaction between the red blood cells and its anti-antibody is called hemagglutination.
Haas flocculation reaction: When the antigen is colloidal (such as cardiolipin, which is made from a beef heart extract and used in the VDRL test), the reaction is called flocculation.
The Vidal test is for the diagnosis of typhoid and paratyphoid disease. The cause of this is the infection with salmonella bacilli from the intestinal bacilli. These bacilli enter the human body through the digestive system and cause the following diseases:
1- Typhoid fever and parathyroid gland known as intestinal fever
2- Septicemia and infection of the blood
3- Food poisoning
Typhoid and parathyroid bacilli can be isolated from blood, feces, and urine, respectively, before the Vidal test is positive. Gradually, the antibody titer rises. Positive microbial cultures are also reduced if, after four weeks of germicide entry, 90 to 95% of the patients will experience a Vidal test.
The microorganism or antigen of H is measurable. The antigen of the O gene is also known to have a greater degree of lipopolysaccharide and its anticorrosive antigens than the IgM class and plays a role in the pathogenesis of the microbe. The antigen of the H gene is derived from protein, IgG class.
Another anti-venous polysaccharide Vi is found in salmonella typhi and para-typhi bacilli. The antibody against it is of the IgM class and appears later than the antigens O and H.
It is better to be fasting for this test.
Avoid hemolysis of the sample because hemolysis interferes with the test results.
Diagnosis of febrile illness based on active or direct agglutination. These experiments are carried out in two quick ways on the tube and the tube in the tube.
- If the test result is false positive or false negative for any reason, the test should be repeated with the tube method.
- If the patient’s serum head is 1.80 in the somatic antigen and at least 1:40 in the case of the anti-HG-1, it is suspected to be typhoid or parathyroid.
- The agglutination of H alone is not worthless if the agglutination of O is due to disease.
- In cases where the test response on the lam does not match the test tube response, the Vidal test is confirmed by a tube.
Wright’s test is used to detect the serology of brucellosis or bruising which is the result of the brucella microbes in humans and most animals. Human beings are more likely to be infected by direct contact with sick animals or taking milk and milk products contaminated with brucellosis. Antibodies that appear in serum against the surface antigens of brucella are IgG, IgM and IgA. At the end of the first or second week of the acute phase, IgM and then the specific IgG are raised, and after a few weeks, the titre also increases with IgM.
Typically, during the fourth to eighth weeks, the acute phase of the disease will reach its maximum level of antibody titre. If the appropriate treatment is performed, the IgG titre is reduced, but there is no change in the IgM titre. However, if the treatment is not appropriate, the disease enters the chronic phase, and in this case, the positive serum head is often related to the IgG antibody.
- Wright test is based on direct agglutination. These experiments are carried out in two quick ways on the tube and the tube in the tube.
- It is better to be fasting for this test.
- Avoid hemolysis of the sample, because hemolysis interferes with the test results.
- If my serum complex is disabled before the test, it may be falsely negative.
Vaccination against cholera may result in false positive results in the Burn test.
- A zone phenomenon (Pro-zone, post-zone) can be created in the Wright experiment due to the mismatch of antigen with an antibody.
Coomb’s wright test
This test is recommended if the result of the Wright test is negative and the physician suspects that patient is entering to the chronic phase of the disease. In the chronic phase of the disease, sometimes anti-brucella antibodies, which do not have agglutination ability, are found in the patient’s serum. These are from the class of IgG or sometimes IgA, termed the term antibody blocking or defective.
In such cases, antigens are known as human collectively known as human canine antibodies to detect these antibodies. Serum Kombes is an anti-human antibody that attaches to the FC region of human antibodies and helps to carry out agglutination. If this test is positive, the person is at a chronic stage of the disease.
2ME wright test
This test is performed after positive Wright test to identify the antibody class. The most important use of this test is the differential diagnosis of inactive active brucellosis in a person with clinical manifestations of the disease, but his blood culture is negative and the tithe of his burn test is also low. In addition, by performing this test, the appropriate antibiotic effect can be studied in the treatment of the disease.
- The 2ME test is used as a complementary assay to distinguish acute chronic brucellosis or previous contact with Brucella antigen.
- Heading 160: 1 or higher this test, which continues for more than a year after the onset of the disease, suggests a lack of improvement in brucellosis.
- The 160: 1 titre, or more, in the 2 ME test indicates an asymptomatic current infection and, if there are clinical symptoms, indicates current active infection, but the 80: 1 and 40: 1 titles may rarely indicate recent major infections and ultimately in patients After three weeks, they still have 2ME of about 20: 1 or less. The probability of Brucella involvement as a causative agent, the disease is largely neglected.
Rheumatoid arthritis is a chronic disease of the local immune complex and is considered as an autoimmune disease. The disease is more common in middle-aged and elderly people, but it’s also known as Juvenile Rheumatoid Arthritis in children. In the serum, about 60-88% of patients with rheumatoid arthritis, an autoantibody of IgM, called rheumatoid factor, appear against the Fc region of human and rabbit IgG molecules.
The concentration serum of RF depends on the degree of inflammation and disease, and increases in patients with chronic rheumatoid arthritis. This factor is positive in patients with lupus erythromatosis, Sjogren syndrome, infectious mononucleosis, dermatomyositis, subacute bacterial endocarditis, chronic active hepatitis, influenza, syphilis, sarcoidosis, biliary cirrhosis, leukemia and pyelonephritis.
- RF test is the most common and easiest way to diagnose rheumatoid arthritis.
- This test is based on the agglutination of the patio.
- This test is also carried out in two layers of quality on a slide and a little in the tube.
- The hemolysis and severe lipemic samples interfere with the results and cause false positives to be tested.
- High levels of serum cryoglobulins cause false positives.
- Since 20% of patients with rheumatoid arthritis are not diagnosed with this method, the negative effect of this test is not due to the definitive health of a person.
CRP is a non-specific, reactive protein and is used to diagnose bacterial infections and inflammatory disorders such as rheumatic fever. From acute phase proteins, the measurement of CRP due to its rapid increase at the onset of tissue lesion and its rapid reduction immediately after recovery is the best way to detect tissue lesions.
- The most sensitive test that can be associated with signs of necrosis or inflammation of the tissue.
- The best test to remain consistently throughout the disease.
- The CRP test is more sensitive and faster than ESR. In acute inflammation, the CRP value increases faster and more than ESR.
- The false positive result in this experiment is so rare.
- Different serologic methods exist for the diagnosis of CRP, among them, prostaglandin-agglutination is more common than other methods. This test is reversible based on inactivated agglutination.
- Sometimes the amount of CRP in the serum is very high, and as a result of regional phenomena, a positive serum may be negatively reported. Therefore, if serum CRP is negative, the test should be repeated before dilution of 1: 5 or more before it is reported.
- The amount of CRP is directly related to the severity of the infection.
- If serum lipid or serum rheumatoid factor is high, CRP may be falsely tested in rare cases.
- In the following diseases, the amount of CRP is raised:Bacterial infections
Active rheumatic fever
Acute myocardial infarction
Malignant cancers spread
Active Rheumatoid Arthritis
O (Antistreptolysin o titer)
The ASO titre is a serologic test that proves the body’s response to the infection caused by beta-hemolytic streptococcus group A. Streptococci generates an enzyme called Streptolysin O, which can leak red blood cells. Because Streptolysin O is antigenic, the body reacts with the production of an anti-antibody to the ASO.
- The ASO test is based on enzyme neutralization tests.
- The ASO headline should be interpreted with caution, usually the headline is considered to be less than 200 units of natural tat. But in 10 to 15% of the normal subjects, the titres are higher than 200 tatts.
- Various factors such as patient’s age, severity of infection, previous history of a patient with streptococcal infections, and immunological status of the patient in relation to antibody production are involved in the interpretation of the test.
- The baby’s ASO headline depends on the blood titre of the mother’s blood serum ASO.
- In patients with glomerulonephritis, bacterial endocarditis, the amount of this enzyme is increased.
- High levels of cholesterol, valprooproteins, can neutralize streptolysin, leading to a false negative response to the ASO titre.
VDRL (Venereal Disease Research Laboratory)
PRP (Rapid Plasma Reagin)
Treponema pallidum is cause of syphilis. Human is the only host of this germ. The transmission of this disease is 90% of cases of sexual contact. But germs can get through the skin or mucus if the epithelial layer is damaged, resulting in contact with a syphilis wound and causing the disease. The disease is divided into four acute, secondary, latent and third stages. The acute phase with canker is indicated on the skin near the infection (usually the genital tract).
The disease is divided into four acute, secondary, latent and third stages. The acute phase with chancre is indicated on the skin near the infection (usually the genital tract). The chancre is created about 3-6 weeks after inoculation and remains about 4-6 weeks old. Pregnant women can transfer the disease to the fetus at this stage. The second stage is characterized by rash (often on palms and legs) and diffuse lymphadenopathy. This takes about three months. The latent phase is an inactive phase of the disease and can last for 5 years. Some patients recover at this stage. Most of these patients go to the third stage, which is associated with CNS, cardiovascular and eye symptoms. In the usual immunohistochemistry tests, two antibody groups are evaluated.
- VDRL and RPR show false positive result due to the search property for a non-specific antibody.
- The VDRL test is positive to the patient about two weeks after trupone inoculation, and in the first and second stages of the disease, and in two thirds of the patients, it becomes positive to the third stage of the disease and returns to normal after appropriate treatment.
- Occasionally, in the secondary syphilis test, due to the high amount of antibodies, the prozone region may occur and the result is falsely negative.
- Treponema pallidum can pass through the placenta after the fourth month of gestation and cause the fetus to spread the hive and thus cause abortion.
- In Syphilis, although an antibody is produced against palpitum, but no serum or humoral or cellular immunity is observed, and a person may be infected several times.
Coombs Test, Direct
The coombs test is done to detect antibodies that stick to the red blood cells and make them susceptible, but are not capable of agglutinating the cells, to these antibodies, defective antibodies, or blocking antibodies. Except for incomplete antibodies, C3, C4 components are also able to sensitize red blood cells.
This test is done to detect hemolysis or to investigate a hemolytic transfusion reaction. In the event of a blood transfusion reaction, the coombs test can detect the patient’s antibodies that cover the surface of the injected RBCs. The coombs test proves that RBCs are attacked by antibodies in the patient’s bloodstream.
- Anti-phospholipid antibodies can cause false positives.
- Drugs such as levodopa, methyl dopa, ampicilin, insulin, penicillin, phenytoin, rifampin, streptomycin, sulfonamides, cephalosporins, indomethacin and tetracyclines can cause false positive results.
- This test is used to diagnose fetal erythroblastosis, unconscious transfusion reactions, autoimmune hemolytic anemiaand red blood cell sensitivity due to medication use.
Coombs Test, Indirect
This test is used to detect anti-RBC antibodies inside the serum for screening blood donors. The difference between direct and indirect coombs is that in the direct coombs, the red blood cells become susceptible to the body.
While in indirect coombs, red blood cells with known antigens are adjacent to the patient’s serum in the laboratory and if the serum contains antibodies against one or more antigens, the red blood cells become sensitive in the laboratory environment. The next steps are almost the same as the straight coombs.
- This test is a routine screening test and is used to diagnose and determine blood compatibility.
- If the agglutination is observed, it means that the blood receptor has antibodies against the red blood cell donors. As a result, the blood is not compatible with the receptor.
This test is done in the following cases:
– Rh negative mothers who are susceptible to Rh antigen.
– Cross-wrist test
– Detection of RhDu Antigen
– Detection of defective antibodies and the presence of specific antibodies to the specific type of agglutinin.
Total Hemolytic Complement Assay
Complement system is a regular system with more than 20 serum proteins that play a very important role in protecting the body against external factors, like microorganisms that attack the body, as well as inflammatory lesions. Complex activity is bound to break through the components of the complement, which strengthens this final response and produces many products resulting from the breakdown of the complement, some of which carry out important biological activities.
The overall performance of the complement system is evaluated by CH50, which evaluates the patient’s serum strength for lysing foreign blood cells. The hemolytic power of the complement is represented by a unit of CH50 unit (50% Hemolytic Unit of complement or unit) (HD50 unit (Hemolytic Dose). One unit of CH50 is the amount or titre of the serum that is required for lysis of 50% of the red blood cells.
PPD ((Mantoux test
A tuberculin skin test is done to examine the likelihood of a person’s contact with tuberculosis tuberculosis bacterial mycobacterium tuberculosis. The most commonly called tuberculin test is called Manto. In this test, the amount of five international tuberculin units from the refined protein solution (PPD) is injected into the forearm with a needle of tuberculin in 0.1 cubic centimeter of buffer.
Tips that should be taken into account after a tuberculin skin test:
- The test result should be read after 48 to 72 hours, since the test is done in the body, it is clear that the person himself must go to the laboratory after 48 to 72 hours.
- After the test, rigidity and redness appear in the skin of the injection area.
- Do not inhale if you feel itching at the injection site.
- Do not apply the injection site for 24 to 72 hours and do not bathe during this time.
- PPD testing should not be performed for patients with active TB or patients who have received BCG vaccine.