University:
College
Course:
Medical Laboratory Science
Academic year:

2023
Views:

163

Pages:

1
Author:

Carlo M.

o VIROLOGY LABORATORY METHODS FOR VIROLOGY SPECIMEN COLLECTION SPECIMEN COLLECTION • • • Appropriate time of collection o Early in disease process o Collected at acute phase of infection or within the first 72 hrs of disease Go where the manifestation of the viral disease shows o Aspirated secretions – maximum recovery o Swabs – use Cotton (Dacron or Nylon) o Aspirate are better than swab but swab is more convenient ▪ Calcium Alginate: Toxic to many viruses o Urine – neutralizing pH with 7.5% solution of sodium bicarbonate to remove vaginal flora o Blood ▪ Anticoagulated blood : CMV ▪ Citrate : Other Viruses ▪ EDTA: Nucleic Acid Testing ▪ Serum: Serologic Test Transport o Medium: buffered isotonic solution + some type of protein + antibiotic o Refrigerated or on ice (not frozen) o -4°C – temperature for storage for viruses o Delay of 4-7 days: -70°C DIAGNOSTIC METHODS IN VIROLOGY LABORATORY DIAGNOSIS • • • • • • Direct Microscopic Examination o For detecting the presence of viral inclusions o Commonly used for herpes simplex, herpes zoster and CMV detection Electron Microscope Antigen Detection o Immunofluorescence Microscopy (FAT) ▪ Direct – uses fluorescence-labeled specific antibody ▪ Indirect – a fluorescence-labeled antiglobulin is used to detect the unlabelled specific antibody o Immunoperoxidase Staining o Solid-Phase Immunoassay ▪ RIA ▪ ELISA ▪ Particle agglutination Animal Inoculation Test Nucleic Acid Hybridization Cultivation o Chick Embryo ▪ sites of inoculation ▪ signs of growth o Tissue Culture Media ▪ Primary Monkey Kidney (PMK) ▪ Human fetal diploid (HFD) ▪ Hep-2 continuous cell line ▪ HeLa cells ▪ African green monkey kidney, baby hamster kidney (BHK), Human embryonic kidney (HEK), • • • Signs of growth: ▪ Cytopathic effect (CPE) ▪ Plaque formations ▪ Production of hemagglutinins ▪ Foci formation Serologic tests o Direct detection ▪ Cytopathic effect (CPE) from cell scrapings Bright-field light microscopy ▪ Direct fluorescent antibody (DFA) staining ▪ Enzyme immunoassays (EIAs) ▪ Nucleic acid-based assays Molecular Methods o Based on the detection of viral genome o Classical Molecular Methods ▪ Depend on the use of specific DNA/RNA probes for hybridization. ▪ Dot-blot, ▪ Southern blot, ▪ In-situ hydridization o The sensitivity of these techniques is not better than conventional viral diagnostic methods. It is usually more tedious and expensive than conventional techniques. o Polymerase Chain Reaction (PCR) and Reverse Transcriptase-PCR