MAJOR BIOCHEMICAL ESTIMATIONS

MAJOR BIOCHEMICAL ESTIMATIONS

Kannan.S.Das
M.Sc Biochemistry
Acid Phosphatase(AcP)
AcP is the name given to a group of Phosphatases with optimal activity below 7.0 and found in various tissues including Liver, Spleen, Prostate , Erythrocytes and Platelets.
Clinical interpretations.
Determination of prostatic AcP activity in serum is useful in monitoring carcinoma of the Prostate.
Normal values
Specimen used
Serum
Plasma – EDTA or Heparin.
Don’t use hemolysed specimens.
Albumin
Of all serum proteins albumin is present in the highest concentration. It maintains the plasma oncotic pressure and the transport of many substances.
Clinical interpretations.
Increased serum albumin may indicate dehydration or hyper fusion with albumin , a decrease is found in rapid hydration , over hydration , severe malnutrition , malabsorption , severe diffuse , liver necrosis , chronic active hepatitis and neoplasia.
Decreased serum albumin may indicate chronic alcoholism , pregnancy , renal protein loss, thyroid dysfunction ,peptic ulcer disease, chronic inflammatory diseases.
Specimen used.
Serum
Plasma –heparin
Alkaline Phosphatase(ALKP)
ALKP is present mainly in bone, liver, kidney, intestine, placenta and lung.
Clinical interpretations.
Serum ALKP may be elevated in increased bone metabolism (in adolescencets and during the healing of a fracture) primary and secondary hyperparathyroidism, Paget’s disease of bone, carcinoma metastatic to bone, Osteogenic Sarcoma, Hodgkin’s disease if bones are invaded. Hepatobiliary diseases involving Cholestasis, inflammation or cirrhosis increase alkaline phosphate activity, renal infarction and failure and in the complications of pregnancy.
Low ALKP activity may occasionally seen in hypothyroidism.
Specimen used.
Serum
Plasma –heparin
EDTA, Citrate fluoride oxalate will interfere with this enzyme, so don’t use this in plasma.
Alanine Aminotransferase (ALT)
ALT activity is present high in Liver, Skeletal muscle, heart and Kidney.
Clinical interpretations.
Serum ALT increase rapidly in liver cell necrosis, hepatitis, hepatic cirrhosis, liver tumors, obstructive jaundice, Reye’s syndrome, extensive trauma to skeletal muscle, myositis, myocarditis and myocardial infraction.
Specimen used
Serum
Plasma – EDTA or Heparin.
Ammonia
Ammonia is a waste product of protein catabolism. It is potentially toxic to the CNS.
Clinical interpretations.
Increased plasma ammonia may be indicative of hepatic encephalopathy , increased coma in terminal stages of liver cirrhosis , hepatic failure , acute and sub acute liver necrosis and Reye’s syndrome. Hyper ammonia may also be found with increasing dietary protein intake.
Specimen Collection. :Plasma – EDTA or Heparin.
Amylase
Amylase is an amylolytic digestive enzyme produced by the exocrine pancreas and salivary glands.
Clinical interpretations.
Amylase is increased in acute pancreatitis, pancreatic abscess or psedocyst, pancreatic trauma, amyloidosis, pancreatic neoplasm, common-bile duct obstruction, after thoracic surgery , mumps parotitis, renal insufficiency.
Specimen used.
Serum
Plasma –heparin
Urine
* plasma activities are approximately 20 U/L higher than serum activities.
Specimen not recommended if the plasma is mixed with Citrate, EDTA, and Fluoride oxalate. If the sample is urine, don’t add Boric acid/Sodium formate, Glacial acetic acid, concentrated hydrochloric acid.
Bilirubin
The term direct bilirubin means bilirubin conjugated to glucuronic acid (Bc) and the term Indirect bilirubin means Unconjugated bilirubin (Bu).The concentration of each of the different forms of serum bilirubin provides important additional diagnostic information when compared to the measurement of Total bilirubin (TBIL) alone.
Normal values
TBIL → 0.3 - 1.0 mg/dl
Bc → 0.1 – 0.3 mg/dl
Bu → 0.2 – 0.7 mg/dl
Direct bilirubin → 0.0 – 0.4 mg/dl
Neonatal bilirubin → 1.0 – 10.5 mg/dl
Clinical interpretations.
a) Unconjugated bilirubin fraction increased during
i) impaired conjugation due to physiological Jaundice Crigler-Najjar Syndrome.
ii) increased production due to Hemolytic jaundice ( Kernicterus in Neonates).
iii)decreased uptake is due to Gillbert’s disease.
iv) Ineffective erythropoiesis.
v) Presence of drugs competing for Glucuronide.
b)Conjugated bilirubin fraction increased during
i) Biliary obstruction due to biliary calculi.
ii) Impaired secretion due to Dubin-Johnson syndrome.
iii) Liver Cell damage.
iv) Rotor Syndrome.
Specimen used.
Serum
Plasma –heparin
Blood Urea Nitrogen.
The major pathway of nitrogen excretion is in the form of Urea that is synthesized in the liver , released into the blood and cleared by the kidneys.
Clinical interpretations.
A high serum urea nitrogen occurs in the Glomerulonephritis, Shock, Urinary tract obstruction, Pyelonephritis, acute or chronic renal failure, severe congestive heart failure, hyperalimentation, diabetic ketoacidosis, dehydration, bleeding from the gastrointestinal tract.
Low urea nitrogen occurs in normal pregnancy, with decreased protein intake, acute liver failure and with intravenous fluid administration.
Specimen Collection
Serum
Plasma – EDTA or Heparin.
Sodium fluoride inhibits the enzyme urease activity, so it is not used for specimen collection.
Note: If we want to convert the BUN value to Urea value, then multiply the BUN value with a factor 2.145.
Calcium.
Calcium is the major mineral component of bone , 99% of the body’s calcium is in bone. Calcium ions plays an important role in the transmission of nerve impulses and in maintaining the normal muscle contraction.
Clinical interpretations.
Abnormal concentrations of serum calcium may indicate malfunction of the parotid gland ,bone diseases, carcinoma, malabsorption syndrome and malnutrition, vitamin D deficiency, overdose with calcium containing antacids and renal diseases.
Specimen used
Serum
Plasma – Heparin.
Urine.
Caution : Protective gloves manufactured with calcium carbonate powders may cause an elevated test results because of the contamination of the sample handling supplies(pipette tips, transfer pipettes, sample cups, caps).Supplies that have come in contact with powdered gloves may subsequently contaminate the test specimen during sample metering. Don’t use blood from patients on EDTA therapy.
Cholinesterase
There are two types of Cholinesterase.
a) Acetylcholinesterase which is found in RBC and Nerve tissues.
b) Cholinesterase which is found in plasma, liver, heart and other tissues.
Clinical interpretations.
These measurements are useful in the diagnosis of pesticide poisoning, liver diseases and sensitivity to succinylcholine administration.
a)Pesticide Poisoning: Organophosphate and Carbamate pesticides are inhibitors of both cholinesterase and Acetylcholinesterase. Although the toxic effect is caused by the inhibition of Acetylcholinesterase in the nerve endings , cholinesterase is often used clinically because it is present in high activities and is easy to measure.
b)Liver Disease: Cirrhosis, hepatitis and carcinoma with metastasis to the liver are known to lower cholinesterase activity. A decrease in the cholinesterase activity is considered a sensitive measure of a drop in liver synthetic capacity , because high activities of cholinesterase are normally present in serum.
c)Sensitivity to succinylcholine administration: Succinylcholine is a short acting muscle relaxant administered during the surgery. It is a reversible inhibitor of acetylcholinesterase and is hydrolyzed by serum cholinesterase. Individuals without sufficient serum cholinesterase activity or with certain genetic variants may be unable to metabolize the drug quickly resulting in prolonged apnea. Low cholinesterase acclivities may be chronic for the individual or transient due to pesticide exposure, liver disorder, pregnancy or the use of oral contractives.
Specimen used
Serum
Plasma – Heparin.
Cholesterol
Cholesterol is present in tissues and in serum & plasma either as cholesterol or cholesterol esters bound to proteins. cholesterol is an essential structural component of cell membranes and the outer layers of the plasma lipoproteins and is the precursor of all steroid hormones, including sex and adrenal hormones ,bile acids and vitamin D.
Cholesterol measurements are used to evaluate the risk of developing coronary artery occlusion ,atherosclerosis ,myocardial infarction, cerebrovascular disease.
Clinical interpretations.
Cholesterol level is increased during coronary atherosclerosis, primary hyper cholestrolemia, secondary hyper lipoproteinemia ,nephritic syndrome, primary biliary cirrhosis, hypothyroidism, some cases of diabetes mellitus.
Low cholesterol level is found in malnutrition , malabsorption, advanced malignancy and hyper thyroidism.
Serum cholesterol level depends on many factors including age and sex.
Specimen used
Serum
Plasma – Heparin
Creatine kinase(CK) / Creatine phosphokinase.
CK is a cellular enzyme with a wide tissue distribution. CK is found mainly in skeletal and cardiac muscle. CK’s physiological role is associated with ATP generation for contractile or transport systems.
Clinical interpretations.
Serum Ck is almost always increased during myocardial infarction or skeletal muscle damage. The enzyme is commonly elevated in myocarditis of any cause, cerebrovascular accidents ,rhabdomyolysis, polymyositis and acute physical exertion, muscular dystrophies in Duchene’s muscular dystrophy.
Low CK is due to decrease muscle mass or muscle wasting. Low serum CK activities are common in elderly ,in the bedridden and in patients with advanced malignancy.
Specimen used
Serum
Plasma – Heparin
Creatine Kinase MB
The CK-MB isoenzyme is found primarily in cardiac muscle, trace amounts are present in skeletal muscle.
Clinical interpretations.
CK-MB is elevated in acute myocardial infarction. CK-MB usually peaks between 12 and 24 hours after myocardial infarction and returns to the normal in 48 to 72 hours in an uncomplicated case.
CK-MB is also increased in myocarditis ,Duchene’s muscular dystrophy, polymyositis, rhabdomyolysis and other myocardial or myopathic disorders.
Specimen used
Serum
Chloride( cl- )
Chloride is the major anion in the extra cellular water space, its physiological significance is in maintaining proper body water distribution, osmotic pressure, and normal anion-cation balance in the ECF compartment.
Clinical interpretations.
Chloride is increased in dehydration ,renal tubular acidosis( hyperchloremia metabolic acidosis) and in excessive infusion of isotonic saline.
Chloride is decreased in over hydration, chronic respiratory acidosis, salt losing nephritis, metabolic alkalosis and congestive heart failure.
Specimen used
Serum
Plasma – Heparin
Creatinine
Serum creatinine and urinary creatinine excretion is a function of lean body mass in normal persons and shows little or no response to dietary changes. Serum creatinine is higher in men than in women. Since urinary creatinine is excreted mainly by glomerular filtration, with only small amounts due to tubular secretion, serum creatinine and 24 hour urine creatinine excretion can be used to estimate the glomerular filtration rate.
Clinical interpretations.
Serum creatinine is increased in acute or chronic renal failure, urinary tract obstruction, reduced renal blood flow, shock, dehydration and rhabdomyolysis. Exercise may cause an increased creatinine clearance.
Low serum creatinine concentration include debilitation and decreased muscle mass.
The creatinine clearance rate is unreliable if the urine flow is low.
Creatinine Clearance Test (CCT) = Total Urine Volume X urine creatinine
1440 Serum creatinine It is useful to correct the clearance value with body surface area. This is important especially in children, persons with short or tall frame. Body surface area may be calculated from the formula
Log A = 0.425 log W + 0.725 log H – 2.144 { where A= Body surface in m2 , W= weight in Kg and H= height in cm. The standard body surface area is 1.73 m2
CCT= Total Urine Volume X urine creatinine X 1.73
1440 Serum creatinine Body surface in m2
Specimen used →Serum, Heparinized Plasma , Urine.
C-reactive protein
C-reactive protein is synthesized by the liver and is one of the acute phase proteins .In the acute phase response, increased concentrations of a number of plasma proteins including C-reactive protein are observed.
Clinical interpretations.
C-reactive protein concentration measurements are useful in the detection and elevation of inflammatory disorders, tissue injury and infections.
Specimen used
Serum
Plasma – Heparin /EDTA.
Direct Bilirubin.
Direct Bilirubin is used for evaluating liver and biliary diseases. An increased direct bilirubin occurs in both intrahepatic and extrahepatic biliary tract obstructions. Hepatocellular causes of elevation include hepatitis, cirrhosis and advanced neoplastic states. It is also increased in Dubin-Johnson syndrome and in Rotor Syndrome.
Direct Bilirubin = Total bilirubin – Bilirubin Unconjugated.
Direct HDL Cholesterol.
Clinical interpretations.
HDL cholesterol is used to evaluate the risk of developing coronary heart disease(CHD).
The risk of CHD increases with lower HDL cholesterol concentrations.
Specimen used
Serum
Plasma – Heparin /EDTA.
Carbon dioxide.
The Carbonic acid – Bicarbonate buffer system is one of the important buffer systems that maintains the pH of the blood. Total CO2 measurements are used for the evaluation of acid—base disorders.
Clinical interpretations.
Total CO2 is increased in respiratory acidosis, metabolic acidosis and excessive alkali intake.
CO2 decreased in compensated respiratory alkalosis, metabolic acidosis and in renal disorders where H+ ions cannot be excreted.
Specimen used
Serum
Plasma – Heparin
Iron (Fe)
Most body iron is found in Hemoglobin. The serum measurement of iron is useful in the differential diagnosis of anemia, iron deficiency anemia, thalassemia, possible sideroblastic anemia and iron poisoning.
Clinical interpretations.
hemosiderosis ,hemolytic anemia’s, Thalassemia, Sideroblastic anemia’s, hepatitis, acute hepatic necrosis, Hemochromatosis, inappropriate iron therapy and iron poisoning.
Serum iron is decreased in cases of insufficient dietary iron, chronic blood loss, inadequate absorption of iron, impaired release of iron stores, infection and chronic diseases.
Specimen used
Serum
Plasma – Heparin
Gamma Glutamyl Transferase.( GGT )
GGT plays a major role in glutamine metabolism and in resorption of amino acids from the glomerular filtrate. It is also important in the absorption of amino acids from the intestinal lumen. GGT is found mainly in the liver, pancreas and kidney, although lower activities can be demonstrated in most of organs.
Clinical interpretations.
Serum GGT is a sensitive indicator of Hepatobiliary disease and is useful in the diagnosis of obstructive jaundice and chronic alcoholic liver disease, in the follow up of chronic alcoholics undergoing treatment and in the detection of hepatotoxicity.
GGT is more responsive to biliary obstruction than AST, ALT, and ALKP. GGT is also increased in hepatoma, carcinoma of pancreas and carcinoma metastatic to the liver.
Specimen used
Serum
Plasma – Heparin /EDTA.
Glucose.
Glucose is the primary cellular energy source.
Clinical interpretations.
Fasting plasma glucose concentrations and tolerance to a dose of glucose are used to establish the diagnosis of diabetes mellitus and disorders of carbohydrate metabolism. Glucose measurements are used to monitor therapy in diabetics and in patients with dehydration, coma, hypoglycemia, insulinoma, acidosis and ketoacidosis.
Specimen used.
Serum
Plasma – Heparin /EDTA/Sodium fluoride/Potassium oxalate.
Urine
CSF
Potassium ( K+ )
Potassium is the major cation of the intracellular fluid.
Measurements of serum potassium are used for the electrolyte imbalance, cardiac arrhythmias, muscular weakness, hepatic encephalopathy, renal failure and for the monitoring of ketoacidosis in the diabetes mellitus and intravenous fluid replacement therapy.
Clinical interpretations.More than 90% of hypertensive patients with aldosteronism have a low K+, a low K+ is also common in vomiting, diarrhea, alcoholism and folic acid deficiency. High K+ values occur in rapid K+ infusions, end stage renal failure, hemolysis, trauma, Addison’s disease, metabolic acidosis, acute starvation, dehydration and acute medical emergency.
Normally K+ is freely filtered by the glomerulus’s but tends to be conserved if the serum K+ is low. Urinary potassium may be elevated with dietary increase, hyperaldosteronism, renal tubular acidosis and at the onset of alkalosis.
Specimen used.
Serum
Plasma – Heparin
Urine
Lactate
Lactate is the end product of the anaerobic metabolism of glucose. The concentration of lactate in the blood is dependent on the rate of production in muscle cells and erythrocytes and the metabolism in the liver. Lactic acidosis usually results from overproduction or underutilization of lactate.
Clinical interpretations
Elevated lactate levels can occur as a result of tissue hyoxia,diabetes mellitus, phenformin therapy,malignancies,glycogen storage disease,ethanol,methanol or salicylate ingestion and metabolic acidosis.
Specimens used
Plasma-Fluoride oxalate
Heparinized plasma is acceptable, but precautions must be taken to retard glycolyisis by keeping the whole blood on ice.
LDH
LDH is an enzyme present in the cytosol of all human cells. It catalyses the reversible reduction of pyruvate to lactate using NADH.
Clinical interpretations
Causes of high LDH include neoplastic states, hypoxic cardio respiratory diseases, myocardial infractions, hemolytic anemias,megaloblastic anemias,hepatic cirrhosis, renal infraction,trauma,muscle damage, muscular dystrophy, shock and hypotension. In myocardial infraction cases,LDH begins to rise within about 12 hrs after infraction and usually returns to normal after two to five days.
Specimens used.
Serum
Plasma – Heparin
Serum is the right specimen for estimation of the LDH.
Specimens not recommended
Do not use hemolysed specimens.
Lipase
Lipase is a digestive enzyme that is mainly produced by the acinar cells of the exocrine pancreas. its physiological role is to hydrolyses the long-chain triglycerides in the small intestine.
Clinical interpretations.
Serum lipase uses rapidly in patients with acute and recurrent pancreatitis, pancreatic cancer, common bile duct obstruction and ingestion of drugs hat are toxic to the pancreas. It is increased by most inflammatory conditions in the abdominal cavity, biliary tract disease, abdominal abscesses and renal failure. Lipase is more specific than total amylase in the digestion of acute pancreatitis.
Specimens used. :Serum
Magnesium (Mg)
Mg is predominantly an intracellular cation and is essential in enzymatic reactions.
Clinical interpretations.
Mg deficiency may cause weakness, tremors, tetany and convulsions. Hypomagnesaemia is associated with hypocalcaemia, alcoholism, some types of malnutrition, malabsorption, chronic hemodialysis and pregnancy.
Increased serum Mg concentrations occur in patients with renal failure, dehydration, Addison’s disease.
Specimens used.
Plasma-EDTA /fluoride oxalate/ citrate.
Urine-- Boric acid with Sodium formate as a preservative.
Sodium(Na)
Sodium is the major cation of ECF.The kidneys regulate sodium content of the body.
Clinical interpretations.
Low sodium concentrations may be due to excessive urine loss, diarrhea, Addison’s disease and renal tubular disease. High Na levels may occur in severe dehydration, some types of brain injury, diabetic coma, excessive intake of sodium salts.
Specimens used.
Serum
Plasma-Heparin
Urine.
Neonatal Bilirubin (NBIL).
NBIL is the sum total of Unconjugated bilirubin and Conjugated bilirubin is increased in the erythroblasosis fetalisis(Hemolytic disease)of the new born which causes Jaundice in
the first two days of life. Other causes of neonatal jaundice includes physiological jaundice, hematomal/hemorrhage, hypothyroidism, obstruction jaundice.
Specimens used.
Serum
Plasma-Heparin
Phosphorus.
Phosphorus, as phosphate is distributed throughout the body. Cause of high serum P include dehydration, hyperparathyroidism, hypervitaminosis D, metastasis to bone sarcoidisis, pulmonary embolism, renal failure, diabetes mellitus with ketosis.
Clinical interpretations.
Low serum phosphorus is found in primary hyperparathyroidism and other causes of serum calcium elevation, sepsis, vitamin D deficiency, renal tubular acidosis, chronic hemodialysis, vomiting and decreased dietary phosphate intake.
Specimens used.
Serum
Plasma-Heparin
Urine.
Specimens not recommended:
Plasma – EDTA / Fluoride oxalate / Citrate.
Urine – Preservatives.


Cerebro Spinal Fluid (CSF) protein.
CSF proteins are those that remain in CSF following the ultracentrifugation of plasma through the choroidal capillary wall. Some proteins that are unique to the CSF are synthesized in the CNS.
Clinical interpretations.
In general diseases that interrupt the integrity of the capillary endothelial barrier lead to an increase in total CSF protein.CSF protein is generally increased in all types of meningitis, cerebral infarction, brain abscess, meningovascular syphilis, subarachinoid hemorrhage, some brain tumors, trauma to the brain, multiple sclerosis, encephalomyelitis, degenerative neurological disease.
Decreased CSF protein may occur in water intoxication, CSF leak and hyperthyroidism.
Specimen used
CSF
Total Bilirubin(TBIL)
TBIL in serum and plasma is the sum of Unconjugated bilirubin ,mono and di-glucuronide conjugated bilirubin and delta bilirubin a fraction covalently bound to albumin.
With the exception of anicteric jaundice, total serum bilirubin is invariably increased in jaundice.
Causes of jaundice are prehepatic, resulting from various hemolytic disease, hepatic; resulting from Hepatocellular injury or obstruction and post hepatic resulting from obstruction of the hepatic or common bile duct.
Specimen used
Serum
Plasma- Heparin
Total Iron Binding Capacity.(TIBC)
Most body iron is found in Hb. The serum measurement of iron is useful in the differential diagnosis of anemia, iron deficiency anemia, thalassemia, possible sideroblastic anemia and iron poisoning. TIBC in serum representing transferring concentration in Iron binding capacity is a useful index of nutritional iron status. Iron deficiency anemia is characterized by a decreased serum iron, increased TIBC or transferrin and a decreased transferrin saturation.
Clinical interpretations
Serum TIBC is increased in iron deficiency. Serum TIBC is decreased in anemia of chronic disease.
Specimen used
Serum.
Total Protein.(TP)
Serum proteins transport drugs ,metabolites and maintain plasma osmotic pressure. Most of the serum proteins are synthesized in the liver, with the exception of gamma globulins. One of the most important serum proteins produced in the liver is albumin. Total serum protein concentration can be used for the evaluation of nutritional status.
Clinical interpretations
Causes of high TP concentration include dehydration, Walden Strom’s macroglobulinemia, multiple myeloma, hyperglobulinemia, Granulomatous disease and some tropical diseases. TP concentration is occasionally increased in collagen diseases, Lupus erythematosus and other instances of chronic infection or inflammation.
Causes of low TP concentration includes pregnancy, excessive intravenous fluid administration, cirrhosis, or other liver diseases, chronic alcoholism, heart failure, nephritic syndrome, Glomerulonephritis, neoplasia, protein-losing enteropathies, malabsorption and sever malnutrition.
Specimen used
Serum
Plasma- heparin.
Triglyceride(TG)
TG, fatty acid esters of glycerol, represent the major form of fat found in the body. Their primary function is to store and provide cellular energy. The concentration of TG in the plasma at any given time is a balance between the rates of entry and removal. TG concentration in the plasma varies with age and gender.
Moderate increases occurs during growth and development.
Clinical interpretations.
TG is used for the evaluation of hyperlipidemias. High concentration may occur with hypothyroidism, nephrotic syndrome, glycogen storage disease and diabetes mellitus. Extremely high TG is common in acute pancreatitis.
Specimen used
Serum(It is the good choice) {12 hour fasting}
Plasma- heparin.
Urine Protein
The filtration and resorption of plasma proteins in the formation of urine are important functions of the intact , healthy kidney. The presence of elevated concentration of protein in urine is a key finding in primary renal disease of glomerular, tubular or mixed origin.
Clinical interpretations.
Proteinuria is also seen in patients with synthetic disorders that affect the kidneys such as diabetes mellitus, hypertension, vascular disease , neoplasia, drug toxicity and certain infectious diseases. Protein may also exist as either a benign or transient condition.
Specimen used
Urine.
Uric Acid
Uric acid is the end product of purine metabolism.
Clinical interpretations
Elevation of uric acid occurs in renal failure, prerenal azotemia, gout, lead poisoning, excessive cell destruction (example: following Chemotherapy) , hemolytic anemia, congestive heart failure and after myocardial infarction. Uric acid is also increased in some endocrine disorders, acidosis , toxemia of pregnancy, hereditary gout, and Glycogen storage disease Type I.
A low uric acid concentration may be found following treatment by some drugs (example: low dose Aspirin ) with low dietary intake of purines, in the presence of renal tubular defects and in Xanthinuria.
Specimen used
Serum
Plasma- heparin.
Low Density Lipoprotein ( LDL ) = Total Cholesterol ─ TG + HDL.{ VLDL = TG}
5 5

Prothrombin Time (PT)
PT is a rapid sensitive screening test for coagulation disorders in the domain of the extrinsic system[ factors II, V,VII & X ].Due to its high sensitivity for these coagulation factors is especially well suited for;
The induction and monitoring of oral anticoagulant therapy.
Diagnosing the genetically caused deficiencies in the coagulation factors of extrinsic system.
Diagnosing the required deficiencies in the coagulation factors.
Checking the synthesis performance of the liver in hepatic diseases.
Principle: The coagulation process is triggered by incubation of plasma with the optimal reaction time of Normal plasma in seconds
Result : the result is reported in seconds, Prothrombin ratio(PR), International normalized Ratio(INR)
To obtain the Prothrombin ratio, the reaction time of the sample is divided by the reaction time of the normal plasma.
PR = Reaction time of the sample (seconds)
Reaction time of normal plasma (Seconds)
If the Prothrombin ratio is determined using a normal plasma which does not have a PR of 1.0, the PR of this plasma has to be taken into account in the calculation.
PR = Reaction time of sample in seconds X PR of Normal Plasma.
Reaction time of Normal Plasma in seconds.
Prothrombin Ratio can be converted into Internationally comparable values by means of International Sensitivity Index (ISI).Result obtained is in INR.
INR = PT X a log factor
Control
Reference Values.
Normal → 12 to 15 seconds.
[Deep Vein Thrombosis, Pulmonary embolism, Arterial diseases, Myocardial infarction] → 2.0 to 3.0 seconds.
Recurrent systemic embolism, artificial cardiac valves → 2.5 to 4.5 seconds.
Activated Partial Thromboplastin Time ( APTT ).
APTT is a global screening procedure used primarily to evaluate coagulation abnormalities in the Intrinsic pathway, will also detect severe functional deficiencies in the factors II,V,X or fibrinogen. It is also used to monitor the effectiveness of heparin therapy, where the clotting time is prolonged in proportion to the level of heparin.
In summary the APTT is applicable for diagnosing Coagulant disorders and therapeutic monitoring of both hemorrhagic and thrombolic disease.
Principle: Factors of intrinsic coagulation system are activated by incubating the plasma with the optimal amount of Phospholipids and two surface activators. The addition of Ca ions triggers the coagulation process and the clotting time is then measured.

Immunoglobulin Assays
The method described here is immuno turbid metric assay. The anti human Ig antibodies ( for A,G,M,D & E) when mixed with samples containing the corresponding Ig’s will form insoluble complexes. These complexes cause an absorbance change depending upon the corresponding Ig concentration of the patient sample , that can be quantified by comparison from the calibrator of the corresponding known Ig concentration.
Clinical Significance of Ig’s.
IgA
IgA represents approximately 10 to 15 % of the total serum immunoglobulins. Its structure is monomeric , similar to the IgG molecule , but 10 to 15 % of the IgA in serum is polymeric , particularly IgA2 which is more resistant to destruction by some pathogenic bacteria. Another more important form of IgA is called secretory IgA which is found in skin , pulmonary , kidney infections and hepatic cirrhosis.
Increased monoclonal IgA concentrations may be found in multiple myeloma and other disturbances of plasmatic cells.
Normal range = 70 to 400 mg/dl.
IgG
IgG is the most important immunoglobulin produced by the plasma cells and represents about 75% of the total immunoglobulins. Its main function is to neutralize toxins in tisular spaces.
IgG deficit may be due to a congenital primary disturbance and is a special risk in children.
Polyclonal hyperimmunoglobulinemia is the normal response to infections, especially in hepatitis and cirrhosis as well as autoimmune diseases.
Increases of monoclonal IgG are found in multiple myeloma , lymphocytic leukemia and Waldenstrom macroglobulinemia.
Normal range = 700 to 1600 mg/dl.
IgM
IgM is the only immunoglobulin that a neonate normally synthesis and in adults it represents the 5 to 10 % of the total immunoglobulins. Its structure is a pentamer of five IgG molecules and its high molecular weight prevents its passage into the extra vascular spaces.
IgM concentrations is decreased in diseases related with hereditary or acquired deficiencies of the immunoglobulin production. Polyclonal increases in serum immunoglobulins are the normal response to infections. The IgM generally increases as a primary response to virus infections and blood stream infections such as malaria and primary biliary cirrhosis. In multiple myeloma , if the paraprotein proves to be IgM, the diagnosis is probably Waldenstrom macroglobulinemia.
Normal range = 40 to 230 mg/dl.

Clinical significance of Some other important Assays.
Alpha-fetoprotein ( AFP ).
AFP is a glycoprotein with a high molecular weight (approx: 68,000 D) consisting of a single polypeptide chain. AFP ,which belongs to the group of oncofetal proteins which is produced by the yolk sac and in the fetal liver.
In oncology, AFP is determined in patients with liver cell carcinoma or germ-cell tumours( non-seminomatous tumors of the testes , endodermal sinus tumors of the ovaries).
AFP plays an important role in pregnancy monitoring. During pregnancy, AFP levels in the maternal blood continuously increased. Between weeks 28 to 32 a maximum is reached , after this period a decrease can be observed until 15th week of gestation. Elevated AFP levels in early pregnancy indicate neural tube defects ( spina bifida, anencephaly ). Lower AFP concentrations in the maternal serum are indicative of Down’s syndrome.
The determination of serum AFP during therapeutic monitoring provides valuable information about the success or failure of treatment as well as the occurrence of recidivation.
Normal Range= healthy men and non-pregnant women normally shows AFP values below 5.5 IU/ml.
Thyroid Stimulating Hormone ( TSH ).
TSH is a glycoproteohormone with a molecular weight in the range of 28,000 to 30,000 Dalton and is composed of the two non-covalently bound subunits hTSHα and hTSHβ.
A characteristic feature of the glycoproteins TSH , Luteinising hormone (LH), Follicle Stimulating Hormone (FSH) and Human Chorionic Gonadotropin (HCG) is their relative carbohydrate content as well as the nearly identical sequential homology of their α-subunits .On the other hand the β-subunit has a different amino acid sequence in all four hormones.
TSH releases and synthesized in the anterior pituitary is stimulated by the hypothalamic thyrotropin-releasing Hormone (TRH).The TSH released stimulates the thyroidal release of the hormones Thyroxin(T4) and Triiodothyronine(T3) whose binding proteins are physiologically active in the peripheral tissues and regulates the thyroidal function via a pituitary feedback mechanism.Determination of basal TSH is generally sufficient in the monitoring of suppression or substitution therapy.
CA125 antigen.
Determination of CA125 antigen is used in the follow-up of patients with primary invasive ovarian carcinoma.
The two antigen determinants defined by the monoclonal antibodies OC125 and M11 are fond on a heterogeneous group of high molecular weight 200,000 to 1,000,000 glycoproteins.
They can be detected in a high percentage of nonmucinous epithelial ovarian tumours. Furthermore they are found in some fetal tissues (amnion, periderm, derivatives of the coelomic epithelium) and in adult tissues in the epithelium of the Fallopian tubes , apocrine sweat glands, breast glands, endometrium and endocervix.
Elevated CA125 assay values in serum are found in most of the patient with active epithelial ovarian cancer in early stages of the disease already and can therefore be used for therapeutic monitoring of such patients.
Normal range = 35 IU/ml. this cut-off can vary depending upon the age and menstrual cycle.
Hepatitis C Virus ( HCV ).
HCV is formerly described as the parenterally transmitted form of non-A, non-B hepatitis (NANBH), causes chronic liver disease in 50% of the cases. After blood transfusion from donors testing positive for HCV antibodies , 88% of recipients develop NANBH and seroconvert to positive anti-HCV test.
HCV can also be transmitted through intravenous drug abuse , sexual contact and administration of contaminated blood or blood products.
HCV is a single stranded RNA virus with some structural relation to the Flavivirus family. The HCV has been linked to cases of Cryptogenic Cirrhosis, Hepatocellular Carcinoma, Auto immune liver diseases and with a variety of extrahepatic disorders such as Glomerulonephritis, Polyarteritis and Cryoglobulinemia.
By using recombinant DNA techniques, the HCV genome is encoded. The genome encodes for three structural proteins Capsid (core) protein, Envelop Glycoproteins E1,E2 and other non structural proteins NS2, NS3, NS4, NS5.
Common tests done for diagnosing a disease.
Diabetes.
Glucose
Amylase
HDL
Pancreas function.
Lipase
Triglycerides
Hyperlipidemia.
Cholesterol
Cholesterol
Triglycerides
HDL
LDL
Liver Function Test ( LFT).
Bilirubin
GGT
Alkaline phosphate
Glucose
Albumin
Cholinesterase
AST
ALT
Ammonia
Gout
Urea
Uric acid
Creatinine
Myocardial infraction
Creatine kinase
Creatine kinase-MB
AST(GOT)
LDH
ALT(GPT)
Tumor Diagnosis
LDH
Alkaline Phosphate
Cholinesterase
Calcium
GGT
Kidney Function Test.
Creatinine
Uric acid
Urea
Total protein
Albumin
Sodium
Potassium
Calcium
Magnesium
Hemoglobin

Courtesy : Manual of Ortho diagnostic instrument.