Immunology and Serology Lab Preliminary 01 PDF

IMMUNOLOGY AND SEROLOGY PRELIM 01 INTRODUCTION AND PIPETTING TECHNIQUES LABO...

IMMUNOLOGY AND SEROLOGY PRELIM 01 INTRODUCTION AND PIPETTING TECHNIQUES LABORATORY SERUM VS PLASMA TESTS KITS Must know the principles behind every test kits/procedure such as: - Agglutination - Precipitation - Lateral Flow Assay - Immunofluorescence Antibody Tests - Western Blot - Enzyme-linked Immunosorbent Assay (ELISA) QUALITATIVE VS QUANTITATIVE TEST Qualitative Sensitive analysis Testing - produce a binary result, such as positive or HOW TO DIFFERENTIATE SERUM FROM PLASMA? negative SERUM PLASMA - Indicates the presence or absence of a Tube used Red top Purple/Lavender top substance or condition Presence of None EDTA (Ethylenediaminetetraacetic Quantitative Specific analysis anticoagulant acid) Testing - Involves numerical values which you must Presence of None. Clotting factors are Present. Due to the presence of quantitate everything. clotting already consumed and anticoagulant that prevent the - Gives you the exact viral load of the test. factors exhausted during entire blood to clot which makes the clotting process for clotting factor remain. serum to be collected. WHAT ARE THE CLOTTING FACTORS PRESENT IN PLASMA SAMPLE? Factor I – Fibrinogen – important/main factor PIPETTING TECHNIQUES Factor 2 – Prothrombin MANUAL PIPETTING Factor 7 – Proconvertin Factor 8 – Antihemophilic Factor A o Hand pipetting using fixed volume is fast in small applications and only Factor 9 – Christmas Factor requires the hand of a practiced lab tech instead of extra hardware. Factor 10 – Stuart-Prower Factor o Time-consuming, results can be unreliable, and the repetitive actions can Factor 11 – Plasma Thromboplastin Antecedent lead to injury Factor 12 – Hageman Factor o 5-10 samples per hour Fibrinogen Level SEMI PIPETTING 200–400 mg/dl = Normal range 400 mg/dl = Hyperfibrinogenemia Increase in fibrinogen level, means you heal faster. Conditions o Most valuable in high-throughput applications that benefit from completely associated to Hyperfibrinogenemia: removing human movements. 1. Platelet Thrombosis o Can process hundreds of samples at a time and follow highly complex 2. Pulmonary Embolism methods without deviation. Most common is unwanted clot formations in blood vessels. o Protection from hazardous/infectious samples NOTE: Having this conditions means you’re prone to cardiovascular accidents. REMEMBER! Buffy Coat Layer Composed of (1) White blood cells and (2) Platelets. Mouth pipetting – the greatest potential hazard is when is done instead of mechanical suction. Mouth pipetting is never acceptable in Granular Neutrophils Fights bacteria and fungi the clinical laboratory. Leukocytes Basophils Respond to inflammation Eosinophils Fights parasites and BUBBLES = WRONG PROCEDURE HANDLING respond to allergic reaction Agranular Monocytes Clean/remove dead cells Bubbles and viscous solutions can cause problems with the measurement and delivery of samples and solutions. Leukocytes Lymphocytes Fights viruses and make antibodies If some procedures require inactivated serum. - Macrophages – are simply monocytes in the tissue, they engulf large particles and pathogens. Complement can be inactivated by heating to 56° C for 30 minutes or, Acquired after centrifugation and found in the middle of the sample. after 4 hours, re-inactivated by heating for 10 minutes Increase buffy coat levels = associated to inflammation/infection, common condition is LEUKEMIA. TYPE OF PIPETTES 1|P a g e YURRY EMMANUEL D. DOLAR  1. TRANSFER PIPETTE Ø Volumetric Pipette Ø Ostwald Folin Pipette 2. GRADUATED PIPETTE Ø Serological Pipette Ø Mohr Pipette 3. MICROPIPETTE TRANSFER PIPETTE Disposable plastic pipettes used to transfer small volumes of liquids VOLUMETRIC PIPETTE o “To deliver" (TD) types that have the bulb closer to the center and accurately deliver a fixed volume of aqueous solution. OSTWALD FOLIN PIPETTE o TD types that have the bulb closer to the delivery tip because they deliver viscous fluids. o These pipettes deliver an accurate volume by being “blown out" using a pipetting bulb. GRADUATED PIPETTE o A graduated pipette is a pipette with its volume, in increments, marked along the tube. It is used to accurately measure and transfer a volume of liquid from one container to another. o Tapered end and graduation marks on the stem SEROLOGIC PIPETTES o The orifice, or tip opening, is larger in the serologic pipette than in other pipettes. The rate of fall of liquid is much too fast for great accuracy or precision. o Calibrated to the tip and must be "blown out" to deliver entire volume. The need to blow out is indicated by the etched rings at the top of the pipette. MOHR PIPETTE o Calibrated between marks. o Cannot be “blown out. MICROPIPETTE o Allow rapid repetitive measurements and delivery of predetermined volumes of reagents and specimens. o Piston-operated devices that allow repeated, accurate, reproducible delivery of specimens, reagents, and other liquids requiring measurement in small amounts. PROPER PIPETTING TECHNIQUES To maximize precision and minimize contamination 1. Adjust pipette to the correct volume 2. Check if you are using the correct pipette tip 3. Make sure no bubbles are produced 4. Always use the pipette in a vertical position 5. No reusing of pipette tips COMMON PIPETTING ERRORS 1. Loose pipette tip 2. Tilting the pipettor 3. Plunger quick release (will cause air bubbles) 4. Second stop draw 2|P a g e YURRY EMMANUEL D. DOLAR