For proper management of a disease, it is important to diagnose it early and know its pathophysiology.
However, conventional diagnosis based on serum and urine analysis cannot do so. The disease is diagnosed when the pathogen has multiplied in large number in the body. It is then quite late and harm to body has been caused by then. Therefore, detection of disease in its early stages is important.
Remedial measures can then be taken and the disease treated without harming the human body. Early diagnosis has now become possible with the help of molecular probes. Molecular probes are labelled (radio- and non-radiolabelled) DNA segments, RNA segments, antibodies and antigens. They are used to detect the disorder through presence of complementary structure in defective gene, 'pathogen, their antigens or antibodies produced against them.
A mixture of DNA, RNA and proteins is treated over gel electrophoresis to separate them. The separated bands can be stained and observed. For determining their identity, they are first transferred tonitrocellulose membrane. The process is called blotting. In Southern blotting, DNA segments an diagnosed by hybridization them with radioactiyely labelled DNA or RNA probes. In Northern blotting RNA is identified by labelled DNA or RNA probe. In Western blotting, protein is identified with the help of labelled antibody.
PCR or polymerase chain reaction is used in case the amount of DNA is small. DNA probes are then used to detect the different defects. The technique is employed to detect HIV in suspected AIDS patient, detect mutations in genes in suspected cancer patients, study, proneness to breast cancer, antenatal diagnosis of congenital diseases, sexually transmitted diseases, parasitic protozoa and helminths etc.
ELISA (Enzyme Linked Immunosorbent Assay Test). It is a technique of detecting very small amount of protein (antibody or antigen) with the help of enzyme peroxidase or alkaline phosphatase and stain producing substrate like 5-aminosalicylic acid or orthophenylenediamine.
Serum is sorbed to the surface of ELISA plate. Antibody specific to antigen for diagnosis is place over immobilised antigen. The spot is washed to remove free antibody. Anti-antibody bound to enzyme is poured over the spot so as to react with complexed anyibody. The area is washed again to remove free anti-antibody enzyme complex. Chromagen is added. It will produce stain if the antigen was present. ELISA is quick methods pf diagnosis of pregnancy (by detection pf hCG in urine), AIDS, hepatitis, STDs, thyroid disorder and rubella Virus.
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