Dinesh Chandra Gupta

 Diagnostics: Diagnosis means to diagnose or find out the rout cause of disease. It is further bifurcated mainly in 2 branches a. Pathology b. Radiology PATHOLOGY is a branch of science which deals with the study of disease condition. In pathology various laboratory techniques are applied to find out the cause of disease. The functional sub branches of clinical pathology are listed below: a. Hematology b. Clinical Biochemistry& Urine Analysis c. Clinical Microbiology & serology d. Cytology e. Histopathology f. Molecular Diagnostic Type of samples used in pathology: a. Blood b. Urine c. Sputum d. Saliva e. CSF f. Stool g. Tissue h. Seamen Important Functional areas of Pathology Lab. Blood: Blood may be described as a composite red color liquid connective tissue, which circulates in the body and performs all functions necessary to keep the body alive. Blood mainly consist of Liquid part: Contains organic compounds like Carbohydrates, Lipids, Proteins etc. The inorganic compounds of the liquid part of blood are electrolytes, calcium, phosphorous etc where as the gases like oxygen, carbon dioxides etc are also found as liquid constituents of the blood. Types of blood sample in diagnostic application: For diagnostics application three types of samples from blood are derived. Whole Blood: The Blood sample collected from the patient and used as it is with out any processing. Plasma = Whole Blood – Solid part. ( It contains fibrinogen in liquid form hence it can be separated by mixing the whole blood with anticoagulant) Serum = plasma – fibrinogen (This can be separated by collecting the blood without Anticoagulant and then allowing it to get solidify. The solid part along with fibrinogen will settle at the bottom and clear supernatant will remain in the upper part which will be yellow in color which is serum) Anticoagulants: When blood is collected it clots after some time. Anticoagulants are the chemicals, which prevents clotting of blood when mixed with blood in proper proportion. Thromboplastin Prothrombin--------------------→Thrombin Ca++ Thrombin Fibrinogen --------------------→ Fibrin (Soluble) Ca++ (Insoluble) Fibrin + Blood Cells → Clots (Fine threads) Clinical Biochemistry: Biochemistry is a science concerned with the chemical constituents of living cells with the reaction and processes they undergo. Clinical biochemistry deals with the biochemistry laboratory application to find out the cause of disease. In biochemistry the substance is measured with the using colorimeter. Colorimetry: In the colorimetric determinations specific reagent or specific reagent are used which react with the specific component to form a colored complex. The concentration of the colored complex is directly proportional to the concentration of the component in the specimen. The depth of colored complex is measured on a photometer or colorimeter. The photometric readings compared with primary known standard. Light travels in wave. The color of light judged by the wavelength, which is the distance between the to identical points i.e. peak-to-peak or crest to crest to crest. It expressed in λ and the unit of wavelength is nm. 1 nm = 10-9 meters. Visible range of light is from 380 nm to 760nm The working of colorimeter or photometer is based on Beer and Lambert’s Law. The simplest expressions of this law are as follows. Beer’s Law: It says that the optical density (Absorbance) of a colored solution is directly proportional to the concentration of the solution. A C Lambert Law: it says that the optical density (Absorbance) of a colored solution is directly proportional to the length of light path, (diameter of cuvette i.e. if the diameter of cuvette doubled the O.D. of solution will be doubled. Al So, by combining both the laws A C. l Where length o light path kept 1cm fixed A C A =C. a Basic parts of a colorimeter: Lamp: A bulb, which emits radiant energy for the phonetic determination. A tungsten lamp emits the light in the visible range (400 to 770 nm). A halogen lamp or deuterium lamp emits the wavelength range 200nm to 900 nm, which covers the part of UV-range, complete visible range and part of infrared range. Condenser: a biconvex lens is used as condenser to focus the light rays in a straight direction. Slit: To adjust the light intensity Filter: A glass or interference filter is used to select the required wavelength of light. Cuvette: It is a round or square tube, made-up of glass, quartz, silica or special type of plastic. It holds the colored solution meant for the determination of absorbance. Photocell: these are photo detector or photodiode made-up of light sensitive material such as. These generate the electrical current when the light falls on it. Display: the final optical density or absorbance of colored solution shown over here. Lamp Condenser Slit Filter Cuvette Photocell ay Display Type of Biochemistry: a. End point b. Kinetic c. Fixed Time d. Multipoint Endpoint Chemistry: When we measure the optical density or absorbance of final colored complex solution at the end of reaction or completion of the reaction known as the End point chemistry. When the specific reagent and sample added at specified temperature the reaction take place and after the specific time the reaction gets completed, we take the optical density of colored solution. O.D. of test Con of test = ---------------- X Std Conc. O.D. of Std. Abs. Time Kinetic Chemistry: In kinetic chemistry we measure the change in O.D. when reaction is in the motion. In case of kinetic determination when, a specimen is mixed with a specific reagent the reaction is continuously monitored while it is in progresses. ▲A1= A1 – A2 A4 ▲A2= A2 – A3 ▲A3= A3 – A4 A3 A2 A1 ▲A1 + ▲A2 + ▲A3 ▲A/ min = ---------------------------- 3 Conc. Of test = ▲A/ min X Factor Fixed Time: This chemistry also known as two point chemistry or Initial rate chemistry. In this type of chemistry after the delay time, change in O.D. monitored or measured between two fix intervals. ▲A1 = A2 – A1 A2 A1 T1 T2 ▲A1 = A2 – A1 ( Test) Conc. Of Test = ------------------------------ X Conc. Of Std. ▲A1= A2 – A1 ( Std.) Multi Point Test In Multipoint calibration the varying concentration of the Control / Calibrators are used and the corresponding OD of the same is plotted on Graph in which “X” Axis comprising of Concentration and on “Y” Axis the Corresponding OD Increment is Plotted. After this an unknown sample will be run and it’s OD will be measured and based on the Graph the concentration would then be calculated. ADENOSINE DEAMINASE Adenosine Deaminase (ADA) is an enzyme widely distributed in animal and human tissues. Adenosine Deaminase is found in most cells, but its chief role concerns the proliferation and differentiation of lymphocytes and has been looked on as a marker for cell-mediated hypersensitive reactions. Increased levels of Adenosine Deaminase have been observed and documented in certain infectious diseases with an active participation of cell-mediated immune responses. Increased serum activity of this enzyme have been found in many infectious diseases caused by microorganisms infecting the macrophages, in leprosy, brucellosis, HIV infections, viral hepatitis, infectious mononucleosis, liver cirrhosis and tuberculosis. Diagnosis of extrapulmonary tuberculosis requires investigation of pleural aspiration, fluid biochemistry, cytology and biopsy. Further, it has been observed that cultures for acid fast bacilli are positive in 20-30% of pleural fluid samples and in 50-80 % of pleural biopsy specimens, making pleural tuberculosis often difficult to diagnose. Relatively new techniques such as Adenosine Deaminase, Interferon Gamma and Polymerase Chain Reaction have been reported to help in diagnosis of tuberculosis. However, the sensitivity of Polymerase Chain Reaction (PCR) for tuberculosis is relatively low (0.42-0.81) and the test is expensive. Interferon Gamma appears to exhibit a better sensitivity (0.89-0.99) but there are relatively few studies of its use. Adenosine Deaminase, has been proposed to be a useful surrogate marker for tuberculosis in pleural, pericardial and peritoneal. Studies have confirmed the high sensitivity and specificity of Adenosine Deaminase for early diagnosis of extrapulmonary tuberculosis and meningitis. ADA Kit Liquid stable format requires no reagent preparation saving time and reducing sample handling Fast test results for a rapid turnaround time Wide range of instrument parameters available for facilitating and simplifying implementation LIPOPROTEIN THE PROTEIN THAT CARRIES LIPID IN BLOOD CALLED LIPOPROTEIN. EXAMPLE: HDL,LDL,VLDL AND CHYLOMICRON HDL-C AND LDL-C Cholesterol Triglycerides can't dissolve in the blood. They have to be transported to the cells by special carriers called Lipoproteins. There are several kinds, but the following are significant because of their Clinical importance. Chylomicrons High-Density Lipoprotein (HDL) , Low-Density lipoprotein (LDL), very Low Density Lipoprotein (VLDL) ADVANTAGEE OF HDL-DIRECT METHOD OVER PRECIPITATION 1. No Centrifugation needed. 2. No Pre-Treatment of samples necessary. 3. Less time required. 4. Accurate result. 5. Fully Automated assay. FEATURES OF BEACON HDL-C Direct Quantitative Assay for HDL-C. 2. Facilitates full automation , eliminate Risk of Human Error. 3. Ready to use two Reagent chemistry, stable till expiry. 4. Certified by CDC. 5. Excellent Correlation with CDC Reference Method. 6. Negligible Interference in high Icteric, Lipemic and Hyper Immunoglobumic. 7. Linear up to 150mg/dl. 8. Convenient pack size.