Laboratory Practice, Safety and Applied Skills
Domain 10 tests safe, accurate bench work: aseptic technique, microscopy, staining, media handling and waste disposal. Marks come from sequence and safety, not just naming equipment.
Core rules
Disinfect work surfaces before and after work; wash hands; wear a lab coat, gloves and goggles where needed. Never mouth pipette; never eat or drink in the laboratory; tie back long hair; keep flames away from flammable disinfectants. Flame loops to red heat and cool before touching culture. Open culture vessels briefly and close them; work close to the flame. Label plates with organism, date and initials. Leave autoclave bags loose, not sealed. Do not overcrowd incubators. Cover spills with absorbent material, flood with disinfectant, allow contact time, then clean; remove broken glass with forceps, not hands. Autoclave or incinerate biohazard waste.
Microscopy and staining
A compound microscope has a base, arm, stage, condenser, iris diaphragm, coarse and fine focus, and objective lenses. Oil immersion at 100× uses immersion oil to prevent refraction; avoid air bubbles. Gram staining uses crystal violet, Gram's iodine, decolouriser and safranin. Acid-fast staining uses carbol fuchsin, acid-alcohol and methylene blue. Lactophenol cotton blue is used for fungi.
Worked example
A Gram stain of a known Gram-positive coccus appears pink. Reasoning: the decolouriser was left too long or the smear was too thick, so the crystal violet–iodine complex washed out and safranin stained the cells. Repeat with a thin smear and controlled decolourisation before reporting Gram-negative.
Traps
Oil must not be used with the 40× objective. Autoclave tape is not proof of sterility. Do not flame plastic loops. Do not discard cultures down the sink. Aseptic technique protects the worker and the environment as well as the sample.