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No Capsule: No Clear halos zone Examples of Capsule Positive and Negativeīacillus anthracis, Klebsiella pneumoniae, Streptococcus pneumonia Neisseria meningitidis Clostridium spp, Rhizaobium spp, etc.

Examine the smear microscopically (100X) for the presence of encapsulated cells as indicated by clear zones surrounding the cells.Ĭapsule: Clear halos zone against dark background.

Drain the crystal violet by tilting the slide at a 45 degree angle and let stain run off until it air dries .

Flood the smear with crystal violet stain (this will stain the cells but not the capsules) for about 1 minutes.

Use the other slide to drag the ink-cell mixture into a thin film along the first slide and let stand for 5-7 minutes.

Using sterile technique, add a loopful of bacterial culture to slide, smearing it in the dye.

Nigrosin may need to be kept very thin or diluted.

India Ink generally works, but it has tiny particles that display Brownian motion that must be differentiated from your bacteria.

Place a small drop of a negative stain (India Ink, Congo Red, Nigrosin, or Eosin) on the slide.Ĭongo Red is easier to see, but it does not work well with some strains.Reagents used for Capsule Stainingĭistilled Water = 100 ml Procedure of Capsule Staining A drop of serum can be used during smearing to enhance the size of the capsule and make it more easily observed with a typical compound light microscope. Capsules appear colorless with stained cells against dark background.Ĭapsules are fragile and can be diminished, desiccated, distorted, or destroyed by heating. By counterstaining with dyes like crystal violet or methylene blue, bacterial cell wall takes up the dye. India ink is difﬁcult to obtain nowadays however, nigrosin is easily acquired.Ī positive capsule stain requires a mordant that precipitates the capsule. The background is formed with india ink or nigrosin or congo red. Negative staining methods contrast a translucent, darker colored, background with stained cells but an unstained capsule. Principle of Capsule StainingĬapsules stain very poorly with reagents used in simple staining and a capsule stain can be, depending on the method, a misnomer because the capsule may or may not be stained. Most capsules are composed of polysaccharides, but some are composed of polypeptides. The capsule differs from the slime layer that most bacterial cells produce in that it is a thick, detectable, discrete layer outside the cell wall. The capsule stain employs an acidic stain and a basic stain to detect capsule production. A capsule is a gelatinous outer layer secreted by bacterial cell and that surrounds and adheres to the cell wall. The main purpose of capsule stain is to distinguish capsular material from the bacterial cell.